Effect of goserelin acetate on non-invasive indices of hepatic steatosis in women with endometriosis

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Anastasilakis, Athanasios D. Anastasilakis, Athina Gkiomisi, and 7 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7566446/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 29 Dec, 2025 Read the published version in Endocrine → Version 1 posted 7 You are reading this latest preprint version Abstract Purpose Menopause has been associated with metabolic dysfunction-associated steatotic liver disease (MASLD). This study primarily aimed to evaluate changes in non-invasive indices of hepatic steatosis in premenopausal women with endometriosis after treatment with goserelin resulting in pharmacologic menopause. Methods This was a post-hoc study of an interventional study. Non-invasive indices of hepatic steatosis and, secondarily, fibrosis were calculated in premenopausal women with endometriosis (intervention group, n = 21) before (baseline), 6 months after goserelin treatment (estrogen-deficient period), and 6 months after goserelin discontinuation (estrogen replenishment period). Matched premenopausal women were recruited for baseline comparisons (control group, n = 27). Steatosis was evaluated using the Hepatic Steatosis Index (HSI), Triglyceride/Glucose Index (TyG), TyG-Body Mass Index (BMI), Triglyceride/High Density Lipoprotein (TG/HDL), Metabolic Score for Insulin Resistance (MetS-IR), Dallas Steatosis Index (DSI), and Framingham Steatosis Index (FSI), and fibrosis with BMI – Aspartate Aminotransferase (AST)/Alanine Aminotransferase (ALT) Ratio Diabetes Mellitus (BARD). Results The non-invasive indices of hepatic steatosis and BARD were similar between patients and controls at baseline and did not significantly change after goserelin treatment, except for MetS-IR, which showed an overall significant change (p = 0.04); however, in pairwise comparisons after Bonferroni correction, no significant difference in MetS-IR was observed between baseline (34.8 ± 7.7), month 6 (33.3 ± 9.0), and endpoint (35.0 ± 8.1) values. As expected, the lipid profiles and insulin resistance worsened during the study. Conclusion Non-invasive hepatic steatosis and BARD indices were not significantly affected by estrogen deficiency caused by 6-month goserelin treatment in premenopausal women with endometriosis. menopause fibrosis goserelin metabolic dysfunction-associated steatotic liver disease metabolic dysfunction-associated steatohepatitis nonalcoholic fatty liver disease non-invasive indices steatosis Figures Figure 1 1. Introduction Metabolic dysfunction-associated steatotic liver disease (MASLD) [ 1 ], previously known as non-alcoholic fatty liver disease (NAFLD), is a highly prevalent disease (25–30% of the general adult population) [ 2 ]. However, the first medication, a selective thyroid hormone receptor-β agonist called resmetirom, was approved by the FDA specifically for its treatment in March 2024 [ 3 ]. Resmetirom, which has been approved for advanced disease [i.e., metabolic dysfunction-associated steatohepatitis (MASH)], previously known as nonalcoholic steatohepatitis, with fibrosis stage (F)2–3, represents a breakthrough for a disease in which only pioglitazone and vitamin E were previously recommended as off-label treatments [ 4 ]. The pathogenesis of MASLD seems to be multifactorial, and the prevailing concept is the “multiple hit” hypothesis, in which insulin resistance (IR), hormones, adipokines and cytokines constitute common contributors (“hits”) to a multifactorial disease [ 5 ]. Among the multiple contributors, estrogens affect hepatic lipogenesis in women directly through estrogen receptors (ERs) and indirectly through changes in the lipid profile [ 6 ]. A recent meta-analysis showed that menopause, whose hallmark is the lack of estrogen, is associated with a higher prevalence of MASLD [ 7 ]. However, there is a lack of interventional studies showing a causal association between estrogen and MASLD. In light of the above, a potential model to study the effect of the presence or deficiency of estrogen on hepatic steatosis and fibrosis could be essential to strengthen this hypothesis. Endometriosis is defined as the presence of extrauterine endometrial-like tissue that can be affected by the menstrual cycle and can potentially lead to estrogen-driven inflammation [ 8 ]. The diagnosis of endometriosis may be based on the patient’s history (chronic pelvic pain, dyspareunia, and/or dysmenorrhea) and ultrasound imaging (“chocolate” cysts); however, the gold standard for the diagnosis of endometriosis is histological confirmation after laparoscopic surgery and biopsy of the lesion [ 8 , 9 ]. Following histological confirmation and surgical excision of the endometrial tissues, adjunctive hormone therapy may be administered, including contraceptives and gonadotropin-releasing hormone (GnRH)-agonists [ 9 ]. More specifically, GnRH agonists, such as goserelin acetate, lead to pharmaceutical menopause by suppressing the hypothalamus-pituitary-ovaries axis, thus causing a type of pharmacologic, acute, albeit reversible, ovarian cessation, which lasts as long as the use of GnRH agonists. The adverse effects of GnRH are generally reversible after discontinuation of treatment [ 10 , 11 ]. Therefore, the treatment of endometriosis with GnRH-agonists can be considered a pharmacological model of the menopause. Considering the above, the primary aim of this post-hoc analysis of a previous prospective study was to evaluate changes in non-invasive indices of hepatic steatosis in premenopausal women with endometriosis before and after treatment with goserelin, reflecting estrogen deficiency and subsequent estrogen replenishment. 2. Materials and Methods 2.1 Patients This was a post-hoc analysis of a previous single-center, prospective, non-randomized, interventional study (ClinicalTrials.gov identifier: NCT04203212) [ 10 ]. The medical records of patients with endometriosis included in the study who were followed up on an outpatient basis at the Gynecology Clinic of the 424 General Military Hospital, Thessaloniki, Greece, were retrieved and re-evaluated. The inclusion criteria for the group of patients with endometriosis (intervention group) were as follows: 1) women of reproductive age; 2) diagnosis of endometriosis, histologically verified by a blinded pathologist; 3) 6-month treatment with goserelin acetate; and 4) available appropriate data and serum samples before the initiation of goserelin acetate (baseline), at 6 months after treatment initiation (estrogen deficiency period), and at 12 months (i.e., 6 months after goserelin acetate discontinuation and menstrual restoration; estrogen sufficiency period, endpoint). Specifically, for the needs of this post-hoc analysis, we recruited a new control group in September 2023, consisting of apparently healthy individuals undergoing check-ups at the 424 General Military Hospital in Thessaloniki, Greece. The inclusion criteria for the participants in the control group were as follows: 1) women of reproductive age who were age- and BMI-matched with the women in the intervention group; 2) normal menstruation; 3) no history, symptoms, or signs of endometriosis; and 4) no previous treatment with GnRH analogs. The exclusion criteria for both groups were as follows: 1) alcohol use > 20 g/day; 2) liver cirrhosis; 3) other liver diseases, such as viral hepatitis, autoimmune hepatitis, drug-induced liver injury, hemochromatosis, Wilson disease, α1 antitrypsin deficiency, or mixed liver diseases; 4) type I diabetes mellitus; 5) uncontrolled hypothyroidism or hyperthyroidism; 6) end-stage renal disease; 7) thrombophilia; 8) any malignancy; 9) pregnancy; 10) any acute infection or acute illness; 11) any drug addiction; and 12) any medication that could positively or negatively affect MASLD, received up to 12 months prior to the initiation of goserelin, including glucagon-like peptide-1 receptor agonists, dipeptidyl peptidase-4 inhibitors, pioglitazone, insulin, vitamin E, ursodeoxycholic acid, orlistat, interferon, tamoxifen, amiodaron, metronidazole, antiviral, antitubercular drugs, and antiepileptic drugs. All enrolled patients (intervention group) received monthly subcutaneous injections of goserelin acetate 3.6 mg (Zoladex®) for 6 months as per the standard of care protocol [ 9 ], which was in use at that time. The menstrual cycle was expected to be restored 30–40 days after goserelin acetate discontinuation. The patients were then monitored for another 6 months after menstrual restoration ( from the first day of the restored menstrual cycle). The women in the control group received no treatment, and their data were only available at baseline. 2.2. Methods At baseline, data on medical history were retrieved, including age, concomitant medications, smoking status, and alcohol consumption. Special focus was given to the history of diabetes mellitus, prediabetes, and the use of contraceptives. At the three time points of the study (i.e., baseline, 6 months after treatment, and endpoint), data on blood pressure, weight, and height were also retrieved, and the body mass index (BMI) was calculated. Morning fasting blood samples were obtained from all the included patients at baseline, 6 months, and endpoint, and from women in the control group at baseline. Several data from the intervention group have already been reported in the original article [ 10 ] and a previous analysis focusing on changes in lipidomics [ 12 ]. For the needs of this post-hoc analysis, we measured specific biochemical parameters using previously unthawed serum samples from women in the intervention group, as well as samples obtained from women in the control group at baseline. The following biochemical parameters were included: glucose, insulin, total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C), triglycerides (TG), albumin, aspartate aminotransferase (AST), alanine aminotransferase (ALT), and γ-glutamyl transferase (γ-GT) concentrations. Laboratory tests were performed at the same laboratory as the 424 General Military Hospital, Thessaloniki, Greece, using standard methods. The Homeostasis Model Assessment - Insulin Resistance (HOMA-IR) was calculated using the equation: glucose (mg/dL) × insulin (mU/L)/405. Hepatic steatosis was evaluated using the following non-invasive indices: Hepatic Steatosis Index (HSI) [ 13 ], Triglyceride/Glucose Index (TyG) [ 14 ], TyG-BMI [ 15 ], TG-HDL [ 16 ], Metabolic Score for Insulin Resistance (MetS-IR) [ 17 ], Dallas Steatosis Index (DSI) [ 18 ], and Framingham Steatosis Index (FSI) [ 19 ]. Hepatic fibrosis was a secondary endpoint of this study, which was evaluated using the non-invasive index of the BMI – AST/ALT Ratio Diabetes mellitus (BARD) [ 20 ]. The study was approved by the Ethics Committee of the 424 General Military Hospital. All procedures were performed in accordance with the Declaration of Helsinki and International Conference on Harmonization for Good Clinical Practice. Written informed consent was obtained from all patients included in this study. 2.3. Statistical analysis The Shapiro–Wilk test was used to evaluate the normality of distribution. Continuous variables are presented as mean ± standard deviation (SD), and categorical variables are presented as frequencies and/or percentages (%). Repeated measures analysis of variance (ANOVA) for normally distributed variables or the Friedman test for non-normally distributed variables were used to compare non-invasive indices over time. Bonferroni correction was used for multiple pairwise comparisons in the case of significant trends in the repeated measures ANOVA or Friedman test. An independent sample t-test for normally distributed variables or Mann-Whitney U test for non-normally distributed variables were used to compare continuous variables between the control and intervention groups at baseline. The chi-square or Fisher’s exact test was used to compare categorical variables. Repeated measures analysis of covariance (ANCOVA) was applied to adjust the results of repeated measures ANOVA for potential confounders, such as age and log(HOMA-IR). Linear regression analysis was also applied at baseline to adjust for differences between groups for the same potential confounders (e.g., age and log(HOMA-IR). For the needs of these adjustments, HOMA-IR was logarithmically transformed because it was not normally distributed. The level of statistical significance (two-tailed) was set at p < 0.05 for all tests. Statistical analyses were performed using R project, version 4.3.1 (The R Foundation, Vienna, Austria). 3. Results Twenty-one premenopausal women with endometriosis (intervention group) and 27 apparently healthy premenopausal women (control group) were included in the analysis. The demographic and biochemical data of the variables of interest are presented in Table 1 . Menstrual cessation was induced in all patients during the 6-month treatment with goserelin acetate. Thirty to 40 days after the last injection of goserelin acetate (median 33 days), the menstrual cycle was restored in all patients. At baseline, hepatic steatosis was presumed in 6/21 (29%) patients in the intervention group and in 7/27 (26%) participants in the control group when the diagnosis was based on a positive result of ≥ 3 non-invasive indices (p = 0.838). Hepatic fibrosis was presumed in 3/21 (14%) patients in the intervention group and in 4/27 (15%) participants in the control group (p = 0.959). Presumed hepatic steatosis (6/21 patients at month 6 and 5/21 patients at the endpoint) and fibrosis (4/21 patients at month 6 and 5/21 patients at the endpoint) rates did not significantly change in the intervention group during the follow-up period. Table 1 Baseline and endpoint data of variables of interest. Controls (n = 27) Patients (n = 21) P-value (between group comparison at baseline)* Trend for p-value (within intervention group comparison)** Baseline Baseline Month 6 Endpoint Age (years) 33.9 ± 8.8 34.9 ± 7.8 - - 0.75 - Smoking (N) 9 6 - - 0.74 - BMI (kg/m 2 ) 23.2 ± 3.9 24.0 ± 4.8 24.1 ± 5.3 24.8 ± 4.9 0.61 0.059 TC (mg/dL) 171.9 ± 28.0 166.1 ± 21.8 194.5 ± 31.4 a 188.1 ± 23.9 a 0.42 < 0.001 TG (mg/dL) 74.4 ± 40.3 82.9 ± 27.8 83.7 ± 52.9 71.3 ± 25.2 0.16 0.54 HDL-C (mg/dL) 55.0 ± 9.0 46.8 ± 10.3 55.9 ± 9.9 a 52.6 ± 9.1 0.004 0.008 LDL-C (mg/dL) 102.0 ± 28.9 103.1 ± 16.5 122.0 ± 27.3 a 121.3 ± 22.5 a 0.87 0.002 γ-GT (U/L) 16.7 ± 8.0 21.7 ± 19.1 17.5 ± 10.7 16.0 ± 6.5 0.83 0.76 AST (U/L) 18.1 ± 3.1 16.9 ± 25.2 10.7 ± 2.3 10.2 ± 2.6 < 0.001 0.09 ALT (U/L) 14.1 ± 5.6 12.1 ± 17.0 5.0 ± 2.2 3.9 ± 1.4 0.002 0.10 AST/ALT 1.41 ± 0.45 1.88 ± 0.75 2.47 ± 1.14 2.84 ± 0.99 a 0.01 0.005 Glucose (mg/dL) 91.7 ± 13.5 84.2 ± 9.5 82.0 ± 7.4 84.3 ± 7.0 0.009 0.57 Insulin (mU/L) 7.50 ± 5.02 5.91 ± 3.79 6.39 ± 3.09 7.95 ± 4.35 0.12 0.015 HOMA-IR 1.67 ± 1.00 1.23 ± 0.80 1.31 ± 0.72 1.68 ± 1.01 0.03 0.01 Data are presented as mean ± standard deviation. *: Independent sample t-test was applied for normally distributed variables, and Mann Whitney U test was applied for non-normally distributed variables, comparing patients with controls at baseline. **: Repeated measures analysis of variance (ANOVA) was applied for normally distributed variables, and Friedman test was applied for non-normally distributed variables, comparing variables within patients for trend. a : p < 0.05 compared to baseline (pairwise comparisons with Bonferroni correction). Abbreviations : ALT, Alanine Aminotransferase; AST, Aspartate Aminotransferase; BMI, Body Mass Index; γ-GT, γ-Glutamyl Transferase; HDL-C, High Density Lipoprotein-Cholesterol; LDL-C, Low Density Lipoprotein-Cholesterol; TG, Triglycerides; TC, Total Cholesterol. The comparisons between the control and intervention groups at baseline are presented in Tables 1 and 2 . HDL-C, AST, ALT, glucose, and HOMA-IR concentrations were lower, whereas the AST/ALT ratio was higher in the intervention group than in the control group. No significant differences were observed in any of the other variables between the control and intervention groups at baseline, including all non-invasive indices (Tables 1 and 2 ). In the linear regression analysis, after adjustment for age (model 1) or age and log(HOMA-IR) (model 2), the comparisons between the controls and patients at baseline remained non-significant for all non-invasive indices (Table 2 ). Model 2 was not applied to TyG, TyG-BMI, and MetS-IR to avoid overadjustment because these indices, as well as HOMA-IR, include glucose in their equations. Table 2 Baseline and endpoint of non-invasive indices of hepatic steatosis and fibrosis. Controls (n = 27) Patients (n = 21) P-value (between group comparison at baseline) Trend for p-value (within intervention group comparison) Unadjusted* model 1 model 2 Unadjusted** model 3 model 4 Baseline Baseline Month 6 Endpoint HSI 31.4 ± 4.3 31.3 ± 5.8 29.8 ± 6.0 30.1 ± 5.1 0.53 0.83 0.46 0.07 0.07 0.06 TyG 8.00 ± 0.53 8.10 ± 0.37 7.97 ± 0.61 7.95 ± 0.37 0.50 0.56 *** 0.17 0.15 *** TyG-BMI 185.8 ± 34.6 195.0 ± 43.8 193.3 ± 51.2 199.9 ± 44.2 0.48 0.49 *** 0.71 0.71 *** TG/HDL-C 0.65 ± 0.52 0.82 ± 0.36 0.70 ± 0.52 0.62 ± 0.27 0.18 0.23 0.10 0.08 0.06 0.07 MetS-IR 32.3 ± 6.4 34.8 ± 7.7 33.3 ± 9.0 35.0 ± 8.1 0.17 0.25 *** 0.04 0.04 *** DSI -3.01 ± 0.87 -2.80 ± 1.08 -3.14 ± 1.13 -3.10 ± 0.96 0.68 0.51 0.63 0.12 0.10 0.09 DSI-NAFLD risk (%) 6.5 ± 6.8 8.9 ± 10.6 7.2 ± 9.1 6.2 ± 6.2 0.66 0.39 0.07 0.37 0.36 0.33 FSI -3.22 ± 0.79 -2.96 ± 0.94 -2.99 ± 1.15 -2.90 ± 0.97 0.23 0.24 0.04 0.93 0.92 0.93 BARD 2.15 ± 0.36 2.05 ± 0.59 2.19 ± 0.40 2.26 ± 0.45 0.69 0.33 0.54 0.08 0.05 0.07 Data are presented as mean ± standard deviation. *: Student’s t-test was applied for normally distributed variables, and Mann Whitney U test was applied for non-normally distributed variables, comparing patients and controls at baseline. **: Repeated measures ANOVA was applied for normally distributed variables, and Friedman test was applied for non-normally distributed variables, comparing variables within patients for trend. ***: Due to inclusion of glucose in the equations of HOMA-IR and the annotated non-invasive indices, models 2 and 4 were not applied in the respective indices. Model 1: adjustment for age; linear regression analysis. Model 2: adjustment for age and log(HOMA-IR); linear regression analysis. Model 3: adjustment for age; repeated measures ANCOVA. Model 4: adjustment for age and log(HOMA-IR); repeated measures ANCOVA. Abbreviations : ALT, Alanine Aminotransferase; ANCOVA, analysis of covariance; AST, Aspartate Aminotransferase; BARD, ΒΜΙ – AST/ALT Ratio Diabetes mellitus; BMI, Body Mass Index; DSI, Dallas Steatosis Index; FSI, Framingham Steatosis Index; HSI, Hepatic Steatosis Index; MetS-IR, Metabolic Score for Insulin Resistance; NAFLD, Nonalcoholic Fatty Liver Disease; TG/HDL, Triglycerides/High Density Lipoprotein; TyG, Triglyceride-Glucose index. Within the intervention group, TC, LDL-C, HDL-C, TC, AST/ALT ratio, insulin, and HOMA-IR increased overall during the study period (Table 1 ). In pairwise comparisons (after Bonferroni correction), TC and LDL-C concentrations were increased at 6 months (p = 0.005 and p = 0.008, respectively) and at the endpoint (p = 0.007 and p = 0.005, respectively) compared to baseline. HDL-C concentrations increased at 6 months compared to the baseline (p = 0.005). AST/ALT concentrations were increased at the endpoint compared to those at baseline (p = 0.001). Insulin and HOMA-IR did not marginally provide statistically significant changes in the pairwise comparisons (p = 0.057 and p = 0.053, respectively, between baseline and endpoint), despite an overall significant trend towards an increase over time. Importantly, significant improvement at the endpoint compared to month 6 was not observed for any of the variables of interest (Table 1 ). No significant changes were observed in other variables of interest (Table 1 ). Regarding non-invasive indices of hepatic steatosis or fibrosis (Table 2 ), MetS-IR showed an overall significant change (p = 0.04); in pairwise comparisons (after Bonferroni correction), no significant difference in MetS-IR was observed. In the repeated-measures ANCOVA, after adjusting for age (Model 3), the overall significant trend remained robust for MetS-IR, providing a marginally significant trend. The error bars for all non-invasive indices for the control group (baseline) and intervention group (baseline, month 6, and endpoint) are depicted in Fig. 1 . 4. Discussion In this study, non-invasive indices of hepatic steatosis and fibrosis were evaluated during a 6-month of goserelin treatment and its discontinuation in women with biopsy-proven endometriosis. All non-invasive indices of hepatic steatosis, apart from MetS-IR, as well as BARD, a non-invasive index of hepatic fibrosis, remained unchanged after goserelin treatment. MetS-IR showed an overall trend towards a decrease at month 6, but this was not statistically significant in pairwise comparisons vs. baseline or endpoint, after appropriate correction. These findings do not support our initial hypothesis that estrogen deficiency caused by goserelin, a GnRH analog, could worsen MASLD, as observed in natural or surgical menopause [ 7 ]. The study design does not allow us to draw secure conclusions regarding the apparently null effect of goserelin on the non-invasive indices of MASLD. We may assume that a 6-month period of estrogen deficiency may not be adequate to affect the respective, non-invasive indices. Of course, a yet unknown direct beneficial effect of goserelin, which may counterbalance its indirect effect through estrogen deficiency on hepatic steatosis, cannot be excluded. This study is the first to evaluate hepatic steatosis and fibrosis non-invasive indices in women with endometriosis treated with goserelin post-surgically. To date, no studies have evaluated the impact of goserelin on MASLD, while previous studies evaluating the effects of GnRH analogs on MASLD in other conditions are few, and methodologically highly heterogeneous. In women, there is only a case report of a girl with central precocious puberty, whose histologically confirmed MASH worsened after leuprorelin treatment, while partial improvement was observed after treatment [ 21 ]. Three studies in men with prostate cancer receiving GnRH agonists or antagonists, that is, androgen deprivation therapy, reported significant progression of hepatic steatosis [ 22 – 24 ]. First, ultrasonographic worsening of hepatic steatosis was shown after a 32-month leuprorelin treatment in a man with prostate cancer; however, this was a much longer duration than that of our study [ 23 ]. Subsequently, in a registry-based study, men who received androgen deprivation therapy for localized prostate cancer were shown to have higher rates of MASLD during follow-up [ 22 ]. In another study, hepatic steatosis, evaluated using computed tomography, worsened in men with prostate cancer receiving androgen deprivation therapy for 6 months [ 24 ]. Different medications, diseases, and sexes render the comparison of these studies with our study hardly feasible. It is important to note that, based on the values of all non-invasive indices of hepatic steatosis (Table 2 ), the rates of hepatic steatosis were low in both patients and controls, possibly due to the young age and the lack of concomitant diseases and medications of the participants, compared to the general population or other epidemiological studies in patients with endometriosis [ 25 , 26 ]. As a secondary observation, the overall lipid profile was adversely affected by iatrogenic temporary menopause (Table 1 ), as previously described, and remained above the baseline concentrations 6 months after menstruation restoration [ 12 ]. The same applies to insulin resistance, at least as evaluated with HOMA-IR. A relevant observation is that these changes were not reversed during menstruation restoration (month 6 to the endpoint), implying that even a 6-month treatment with GnRH analogs may infer longer-term adverse metabolic changes [ 12 ]. The strengths of our study include its originality, as there are no known completed or ongoing studies assessing the association between GnRH-induced estrogen deficiency and MASLD in patients with endometriosis. Moreover, the sample consisted of young premenopausal women without comorbidities or concomitant medications that might have affected MASLD. Furthermore, the study consisted of two periods: the first (6 months) characterized by estrogen deficiency and the second (6 months) by estrogen replenishment, during which the effect of goserelin is expected to be fully washed off. The current study has some limitations. First, the study was not specifically designed to show the effect of goserelin on non-invasive indices of hepatic steatosis and fibrosis, but was a post-hoc analysis of a study design with other aims [ 10 ]. Second, the control group was evaluated only at baseline. Third, the controls were chosen from a different pool; thus, there was a risk of selection bias. Ideally, we should have recruited women with endometriosis who were not subjected to any treatment for six months, which was difficult to perform in the setting of a post-hoc analysis. Fourth, the sample size maybe small and no a priori power analysis had been performed, largely owing to the fact that this was a post-hoc analysis, so we selected all available data of the mother study [ 10 ]. Furthermore, the time frame may be relatively short, since 6 months of goserelin-induced estrogen deficiency may not be sufficient for the development of hepatic steatosis and, much more, fibrosis. Moreover, the lack of paired liver biopsies or an appropriate imaging technique to quantify hepatic steatosis or fibrosis at baseline, month 6, and the endpoint is another limitation, largely owing to the retrospective retrieval of data from the intervention group. Due to the same reason, the lack of certain parameters, such as platelet count, did not allow us to calculate other non-invasive indices, especially of fibrosis (e.g., fibrosis-4 index, NAFLD fibrosis score); however, as mentioned above, we consider the 6-month period of estrogen deficiency a short period for the development of hepatic fibrosis. In conclusion, this study showed that the non-invasive indices of hepatic steatosis and BARD, a non-invasive index of fibrosis, were not significantly affected by the estrogen deficiency caused by 6-month treatment with goserelin in premenopausal women with endometriosis. Although further clinical studies based on histological confirmation or appropriate imaging of hepatic steatosis or fibrosis are needed to draw more secure results, the results of this study may imply that goserelin may be considered in patients with endometriosis and MASLD, because it is not expected to worsen MASLD when administered for a short period. Declarations Ethical approval The study was approved by the ethics committee of the 424 General Military Hospital (16329/04-09-2018). All procedures performed in the study were in accordance with the ethical standards of the institutional research committees and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards. Informed consent Informed consent was retrospectively obtained from all participants for the use of their data for the purpose of the study. Author contributions Conception and design of the work: Dimitrios A. Anastasilakis, Athanasios D. Anastasilakis, Stergios A. Polyzos Acquisition of data: Dimitrios A. Anastasilakis, Athanasios D. Anastasilakis, Athina Gkiomisi, Stergios Papachatzopoulos Analysis of data: Dimitrios A. Anastasilakis, Anastasios Semertzidis, Stergios A. Polyzos Interpretation of data: All authors Drafting the article: Dimitrios A. 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Anastasilakis, S. Papachatzopoulos, P. Makras, A. Gkiomisi, P. Nikolakopoulos, S.A. Polyzos, C. Ntenti, I. Ballaouri, S. Gerou, O. Tsachouridou, A. Papatheodorou, K. Aliazis, S. Fermanoglou, I. Bisbinas, M.P. Yavropoulou, The effect of pharmacological cessation and restoration of menstrual cycle on bone metabolism in premenopausal women with endometriosis. Bone. 158 , 116354 (2022) L. Gong, S. Zhang, Y. Han, Q. Long, S. Zou, Y. Cao, Initiation of GnRH agonist treatment on 3–5 days postoperatively in endometriosis patients: a randomized controlled trial. J. Clin. Pharmacol. 55 , 848–853 (2015) A.D. Anastasilakis, S.A. Polyzos, P.A. Vorkas, A. Gkiomisi, M.P. Yavropoulou, M. Rauner, P. Nikolakopoulos, S. Papachatzopoulos, P. Makras, S. Gerou, L.C. Hofbauer, A. Palermo, E. Tsourdi, Lipid Profile after Pharmacologic Discontinuation and Restoration of Menstruation in Women with Endometriosis: A 12-Month Observational Prospective Study. J. Clin. Med. 12 , (2023) J.H. Lee, D. Kim, H.J. 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Gotoda, A Case of Nonalcoholic Steatohepatitis in Central Precocious Puberty Aggravated by Gonadotropin-releasing Hormone Analog. JPGN Rep. 1 , e014 (2020) P. Gild, A.P. Cole, A. Krasnova, B.A. Dickerman, N. von Landenberg, M. Sun, L.A. Mucci, S.R. Lipsitz, F.K. Chun, P.L. Nguyen, A.S. Kibel, T.K. Choueiri, S. Basaria, Q.D. Trinh, Liver Disease in Men Undergoing Androgen Deprivation Therapy for Prostate Cancer. J. Urol. 200 , 573–581 (2018) C. Gabbi, F. Carubbi, L. Losi, P. Loria, M. Costantini, M. Bertolotti, N. Carulli, Nonalcoholic fatty liver disease induced by leuprorelin acetate. J. Clin. Gastroenterol. 42 , 107–110 (2008) H. Sawazaki, Y. Kitamura, K. Yagi, Y. Arai, Impact of Androgen Deprivation Therapy on Non-Alcoholic Fatty Liver Disease in Patients with Prostate Cancer: A CT Evaluation. Urol. Int. 104 , 425–430 (2020) H. Huang, Z. Liu, J. Ruan, Z. Fang, C. Xu, Laparoscopically confirmed endometriosis and the risk of incident NAFLD: a prospective cohort study. Reprod. Biol. Endocrinol. 23 , 55 (2025) E.S. Makri, E. Makri, S.A. Polyzos, Combination Therapies for Nonalcoholic Fatty Liver Disease. J. Pers. Med. 12 , (2022) Additional Declarations No competing interests reported. Cite Share Download PDF Status: Published Journal Publication published 29 Dec, 2025 Read the published version in Endocrine → Version 1 posted Editorial decision: Revision requested 11 Oct, 2025 Reviews received at journal 09 Oct, 2025 Reviewers agreed at journal 21 Sep, 2025 Reviewers invited by journal 16 Sep, 2025 Editor assigned by journal 10 Sep, 2025 Submission checks completed at journal 10 Sep, 2025 First submitted to journal 08 Sep, 2025 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-7566446","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":518254251,"identity":"6c98c0d7-e35e-427a-b99a-b276fb35aa30","order_by":0,"name":"Dimitrios A. 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(B) TyG; (C) TYG-BMI; (D) TG/HDL-C; (E) MetS-IR; (F) DSI; (G) FSI; (H) BARD.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAbbreviations:\u003c/strong\u003e ALT, Alanine Aminotransferase; AST, Aspartate Aminotransferase; BARD, ΒΜΙ – AST/ALT Ratio Diabetes mellitus; BMI, Body Mass Index; DSI, Dallas Steatosis Index; FSI, Framingham Steatosis Index; HSI, Hepatic Steatosis Index; MetS-IR, Metabolic Score for Insulin Resistance; TG/HDL-C, Triglycerides/High Density Lipoprotein; TyG, Triglyceride-Glucose index.\u003c/p\u003e","description":"","filename":"Figure1.jpg","url":"https://assets-eu.researchsquare.com/files/rs-7566446/v1/bc35c7a9821ba28c8bd853ee.jpg"},{"id":99545280,"identity":"f03331af-5579-4268-9c5c-62f8e69c9313","added_by":"auto","created_at":"2026-01-05 16:05:10","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1126215,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7566446/v1/c1383dfa-71c1-4d95-b333-228b7727d89e.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Effect of goserelin acetate on non-invasive indices of hepatic steatosis in women with endometriosis","fulltext":[{"header":"1. Introduction","content":"\u003cp\u003eMetabolic dysfunction-associated steatotic liver disease (MASLD) [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e], previously known as non-alcoholic fatty liver disease (NAFLD), is a highly prevalent disease (25\u0026ndash;30% of the general adult population) [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e]. However, the first medication, a selective thyroid hormone receptor-β agonist called resmetirom, was approved by the FDA specifically for its treatment in March 2024 [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e]. Resmetirom, which has been approved for advanced disease [i.e., metabolic dysfunction-associated steatohepatitis (MASH)], previously known as nonalcoholic steatohepatitis, with fibrosis stage (F)2\u0026ndash;3, represents a breakthrough for a disease in which only pioglitazone and vitamin E were previously recommended as \u003cem\u003eoff-label\u003c/em\u003e treatments [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eThe pathogenesis of MASLD seems to be multifactorial, and the prevailing concept is the \u0026ldquo;multiple hit\u0026rdquo; hypothesis, in which insulin resistance (IR), hormones, adipokines and cytokines constitute common contributors (\u0026ldquo;hits\u0026rdquo;) to a multifactorial disease [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e]. Among the multiple contributors, estrogens affect hepatic lipogenesis in women directly through estrogen receptors (ERs) and indirectly through changes in the lipid profile [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e]. A recent meta-analysis showed that menopause, whose hallmark is the lack of estrogen, is associated with a higher prevalence of MASLD [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e]. However, there is a lack of interventional studies showing a causal association between estrogen and MASLD. In light of the above, a potential model to study the effect of the presence or deficiency of estrogen on hepatic steatosis and fibrosis could be essential to strengthen this hypothesis.\u003c/p\u003e\u003cp\u003eEndometriosis is defined as the presence of extrauterine endometrial-like tissue that can be affected by the menstrual cycle and can potentially lead to estrogen-driven inflammation [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e]. The diagnosis of endometriosis may be based on the patient\u0026rsquo;s history (chronic pelvic pain, dyspareunia, and/or dysmenorrhea) and ultrasound imaging (\u0026ldquo;chocolate\u0026rdquo; cysts); however, the gold standard for the diagnosis of endometriosis is histological confirmation after laparoscopic surgery and biopsy of the lesion [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e, \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. Following histological confirmation and surgical excision of the endometrial tissues, adjunctive hormone therapy may be administered, including contraceptives and gonadotropin-releasing hormone (GnRH)-agonists [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. More specifically, GnRH agonists, such as goserelin acetate, lead to pharmaceutical menopause by suppressing the hypothalamus-pituitary-ovaries axis, thus causing a type of pharmacologic, acute, albeit reversible, ovarian cessation, which lasts as long as the use of GnRH agonists. The adverse effects of GnRH are generally reversible after discontinuation of treatment [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e, \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e]. Therefore, the treatment of endometriosis with GnRH-agonists can be considered a pharmacological model of the menopause.\u003c/p\u003e\u003cp\u003eConsidering the above, the primary aim of this \u003cem\u003epost-hoc\u003c/em\u003e analysis of a previous prospective study was to evaluate changes in non-invasive indices of hepatic steatosis in premenopausal women with endometriosis before and after treatment with goserelin, reflecting estrogen deficiency and subsequent estrogen replenishment.\u003c/p\u003e"},{"header":"2. Materials and Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e\u003ch2\u003e2.1 Patients\u003c/h2\u003e\u003cp\u003eThis was a \u003cem\u003epost-hoc\u003c/em\u003e analysis of a previous single-center, prospective, non-randomized, interventional study (ClinicalTrials.gov identifier: NCT04203212) [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]. The medical records of patients with endometriosis included in the study who were followed up on an outpatient basis at the Gynecology Clinic of the 424 General Military Hospital, Thessaloniki, Greece, were retrieved and re-evaluated. The inclusion criteria for the group of patients with endometriosis (intervention group) were as follows: 1) women of reproductive age; 2) diagnosis of endometriosis, histologically verified by a blinded pathologist; 3) 6-month treatment with goserelin acetate; and 4) available appropriate data and serum samples before the initiation of goserelin acetate (baseline), at 6 months after treatment initiation (estrogen deficiency period), and at 12 months (i.e., 6 months after goserelin acetate discontinuation and menstrual restoration; estrogen sufficiency period, endpoint).\u003c/p\u003e\u003cp\u003eSpecifically, for the needs of this \u003cem\u003epost-hoc\u003c/em\u003e analysis, we recruited a new control group in September 2023, consisting of apparently healthy individuals undergoing check-ups at the 424 General Military Hospital in Thessaloniki, Greece. The inclusion criteria for the participants in the control group were as follows: 1) women of reproductive age who were age- and BMI-matched with the women in the intervention group; 2) normal menstruation; 3) no history, symptoms, or signs of endometriosis; and 4) no previous treatment with GnRH analogs.\u003c/p\u003e\u003cp\u003eThe exclusion criteria for both groups were as follows: 1) alcohol use\u0026thinsp;\u0026gt;\u0026thinsp;20 g/day; 2) liver cirrhosis; 3) other liver diseases, such as viral hepatitis, autoimmune hepatitis, drug-induced liver injury, hemochromatosis, Wilson disease, α1 antitrypsin deficiency, or mixed liver diseases; 4) type I diabetes mellitus; 5) uncontrolled hypothyroidism or hyperthyroidism; 6) end-stage renal disease; 7) thrombophilia; 8) any malignancy; 9) pregnancy; 10) any acute infection or acute illness; 11) any drug addiction; and 12) any medication that could positively or negatively affect MASLD, received up to 12 months prior to the initiation of goserelin, including glucagon-like peptide-1 receptor agonists, dipeptidyl peptidase-4 inhibitors, pioglitazone, insulin, vitamin E, ursodeoxycholic acid, orlistat, interferon, tamoxifen, amiodaron, metronidazole, antiviral, antitubercular drugs, and antiepileptic drugs.\u003c/p\u003e\u003cp\u003eAll enrolled patients (intervention group) received monthly subcutaneous injections of goserelin acetate 3.6 mg (Zoladex\u0026reg;) for 6 months as per the standard of care protocol [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e], which was in use at that time. The menstrual cycle was expected to be restored 30\u0026ndash;40 days after goserelin acetate discontinuation. The patients were then monitored for another 6 months after menstrual restoration ( from the first day of the restored menstrual cycle). The women in the control group received no treatment, and their data were only available at baseline.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec4\" class=\"Section2\"\u003e\u003ch2\u003e2.2. Methods\u003c/h2\u003e\u003cp\u003eAt baseline, data on medical history were retrieved, including age, concomitant medications, smoking status, and alcohol consumption. Special focus was given to the history of diabetes mellitus, prediabetes, and the use of contraceptives. At the three time points of the study (i.e., baseline, 6 months after treatment, and endpoint), data on blood pressure, weight, and height were also retrieved, and the body mass index (BMI) was calculated. Morning fasting blood samples were obtained from all the included patients at baseline, 6 months, and endpoint, and from women in the control group at baseline. Several data from the intervention group have already been reported in the original article [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e] and a previous analysis focusing on changes in lipidomics [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e]. For the needs of this \u003cem\u003epost-hoc\u003c/em\u003e analysis, we measured specific biochemical parameters using previously unthawed serum samples from women in the intervention group, as well as samples obtained from women in the control group at baseline. The following biochemical parameters were included: glucose, insulin, total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C), triglycerides (TG), albumin, aspartate aminotransferase (AST), alanine aminotransferase (ALT), and γ-glutamyl transferase (γ-GT) concentrations. Laboratory tests were performed at the same laboratory as the 424 General Military Hospital, Thessaloniki, Greece, using standard methods. The Homeostasis Model Assessment - Insulin Resistance (HOMA-IR) was calculated using the equation: glucose (mg/dL) \u0026times; insulin (mU/L)/405.\u003c/p\u003e\u003cp\u003eHepatic steatosis was evaluated using the following non-invasive indices: Hepatic Steatosis Index (HSI) [\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e], Triglyceride/Glucose Index (TyG) [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e], TyG-BMI [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e], TG-HDL [\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e], Metabolic Score for Insulin Resistance (MetS-IR) [\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e], Dallas Steatosis Index (DSI) [\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e], and Framingham Steatosis Index (FSI) [\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e]. Hepatic fibrosis was a secondary endpoint of this study, which was evaluated using the non-invasive index of the BMI \u0026ndash; AST/ALT Ratio Diabetes mellitus (BARD) [\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e].\u003c/p\u003e\u003cp\u003e The study was approved by the Ethics Committee of the 424 General Military Hospital. All procedures were performed in accordance with the Declaration of Helsinki and International Conference on Harmonization for Good Clinical Practice. Written informed consent was obtained from all patients included in this study.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec5\" class=\"Section2\"\u003e\u003ch2\u003e2.3. Statistical analysis\u003c/h2\u003e\u003cp\u003eThe Shapiro\u0026ndash;Wilk test was used to evaluate the normality of distribution. Continuous variables are presented as mean\u0026thinsp;\u0026plusmn;\u0026thinsp;standard deviation (SD), and categorical variables are presented as frequencies and/or percentages (%). Repeated measures analysis of variance (ANOVA) for normally distributed variables or the Friedman test for non-normally distributed variables were used to compare non-invasive indices over time. Bonferroni correction was used for multiple pairwise comparisons in the case of significant trends in the repeated measures ANOVA or Friedman test. An independent sample t-test for normally distributed variables or Mann-Whitney U test for non-normally distributed variables were used to compare continuous variables between the control and intervention groups at baseline. The chi-square or Fisher\u0026rsquo;s exact test was used to compare categorical variables. Repeated measures analysis of covariance (ANCOVA) was applied to adjust the results of repeated measures ANOVA for potential confounders, such as age and log(HOMA-IR). Linear regression analysis was also applied at baseline to adjust for differences between groups for the same potential confounders (e.g., age and log(HOMA-IR). For the needs of these adjustments, HOMA-IR was logarithmically transformed because it was not normally distributed. The level of statistical significance (two-tailed) was set at p\u0026thinsp;\u0026lt;\u0026thinsp;0.05 for all tests. Statistical analyses were performed using R project, version 4.3.1 (The R Foundation, Vienna, Austria).\u003c/p\u003e\u003c/div\u003e"},{"header":"3. Results","content":"\u003cp\u003eTwenty-one premenopausal women with endometriosis (intervention group) and 27 apparently healthy premenopausal women (control group) were included in the analysis. The demographic and biochemical data of the variables of interest are presented in Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e. Menstrual cessation was induced in all patients during the 6-month treatment with goserelin acetate. Thirty to 40 days after the last injection of goserelin acetate (median 33 days), the menstrual cycle was restored in all patients. At baseline, hepatic steatosis was presumed in 6/21 (29%) patients in the intervention group and in 7/27 (26%) participants in the control group when the diagnosis was based on a positive result of \u0026ge;\u0026thinsp;3 non-invasive indices (p\u0026thinsp;=\u0026thinsp;0.838). Hepatic fibrosis was presumed in 3/21 (14%) patients in the intervention group and in 4/27 (15%) participants in the control group (p\u0026thinsp;=\u0026thinsp;0.959). Presumed hepatic steatosis (6/21 patients at month 6 and 5/21 patients at the endpoint) and fibrosis (4/21 patients at month 6 and 5/21 patients at the endpoint) rates did not significantly change in the intervention group during the follow-up period.\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003eBaseline and endpoint data of variables of interest.\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"7\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eControls (n\u0026thinsp;=\u0026thinsp;27)\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colspan=\"3\" nameend=\"c5\" namest=\"c3\"\u003e\u003cp\u003ePatients\u003c/p\u003e\u003cp\u003e(n\u0026thinsp;=\u0026thinsp;21)\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e\u003cp\u003eP-value (between group comparison at baseline)*\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c7\"\u003e\u003cp\u003eTrend for p-value (within intervention group comparison)**\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eBaseline\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u003cp\u003eBaseline\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e\u003cp\u003eMonth 6\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e\u003cp\u003eEndpoint\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c7\"\u003e\u0026nbsp;\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003eAge (years)\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e33.9\u0026thinsp;\u0026plusmn;\u0026thinsp;8.8\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e34.9\u0026thinsp;\u0026plusmn;\u0026thinsp;7.8\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e-\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e-\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e0.75\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e-\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003eSmoking (N)\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e9\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e6\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e-\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e-\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e0.74\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e-\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003eBMI (kg/m\u003c/b\u003e\u003csup\u003e\u003cb\u003e2\u003c/b\u003e\u003c/sup\u003e\u003cb\u003e)\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e23.2\u0026thinsp;\u0026plusmn;\u0026thinsp;3.9\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e24.0\u0026thinsp;\u0026plusmn;\u0026thinsp;4.8\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e24.1\u0026thinsp;\u0026plusmn;\u0026thinsp;5.3\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e24.8\u0026thinsp;\u0026plusmn;\u0026thinsp;4.9\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e0.61\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e0.059\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003eTC (mg/dL)\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e171.9\u0026thinsp;\u0026plusmn;\u0026thinsp;28.0\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e166.1\u0026thinsp;\u0026plusmn;\u0026thinsp;21.8\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e194.5\u0026thinsp;\u0026plusmn;\u0026thinsp;31.4\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e188.1\u0026thinsp;\u0026plusmn;\u0026thinsp;23.9\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e0.42\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e\u0026lt;\u0026thinsp;0.001\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003eTG (mg/dL)\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e74.4\u0026thinsp;\u0026plusmn;\u0026thinsp;40.3\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e82.9\u0026thinsp;\u0026plusmn;\u0026thinsp;27.8\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e83.7\u0026thinsp;\u0026plusmn;\u0026thinsp;52.9\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e71.3\u0026thinsp;\u0026plusmn;\u0026thinsp;25.2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e0.16\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e0.54\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003eHDL-C (mg/dL)\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e55.0\u0026thinsp;\u0026plusmn;\u0026thinsp;9.0\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e46.8\u0026thinsp;\u0026plusmn;\u0026thinsp;10.3\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e55.9\u0026thinsp;\u0026plusmn;\u0026thinsp;9.9\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e52.6\u0026thinsp;\u0026plusmn;\u0026thinsp;9.1\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e0.004\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e0.008\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003eLDL-C (mg/dL)\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e102.0\u0026thinsp;\u0026plusmn;\u0026thinsp;28.9\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e103.1\u0026thinsp;\u0026plusmn;\u0026thinsp;16.5\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e122.0\u0026thinsp;\u0026plusmn;\u0026thinsp;27.3\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e121.3\u0026thinsp;\u0026plusmn;\u0026thinsp;22.5\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e0.87\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e0.002\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003eγ-GT (U/L)\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e16.7\u0026thinsp;\u0026plusmn;\u0026thinsp;8.0\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e21.7\u0026thinsp;\u0026plusmn;\u0026thinsp;19.1\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e17.5\u0026thinsp;\u0026plusmn;\u0026thinsp;10.7\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e16.0\u0026thinsp;\u0026plusmn;\u0026thinsp;6.5\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e0.83\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e0.76\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003eAST (U/L)\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e18.1\u0026thinsp;\u0026plusmn;\u0026thinsp;3.1\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e16.9\u0026thinsp;\u0026plusmn;\u0026thinsp;25.2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e10.7\u0026thinsp;\u0026plusmn;\u0026thinsp;2.3\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e10.2\u0026thinsp;\u0026plusmn;\u0026thinsp;2.6\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e\u0026lt;\u0026thinsp;0.001\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e0.09\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003eALT (U/L)\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e14.1\u0026thinsp;\u0026plusmn;\u0026thinsp;5.6\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e12.1\u0026thinsp;\u0026plusmn;\u0026thinsp;17.0\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e5.0\u0026thinsp;\u0026plusmn;\u0026thinsp;2.2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e3.9\u0026thinsp;\u0026plusmn;\u0026thinsp;1.4\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e0.002\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e0.10\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003eAST/ALT\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e1.41\u0026thinsp;\u0026plusmn;\u0026thinsp;0.45\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e1.88\u0026thinsp;\u0026plusmn;\u0026thinsp;0.75\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e2.47\u0026thinsp;\u0026plusmn;\u0026thinsp;1.14\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e2.84\u0026thinsp;\u0026plusmn;\u0026thinsp;0.99\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e0.01\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e0.005\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003eGlucose (mg/dL)\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e91.7\u0026thinsp;\u0026plusmn;\u0026thinsp;13.5\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e84.2\u0026thinsp;\u0026plusmn;\u0026thinsp;9.5\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e82.0\u0026thinsp;\u0026plusmn;\u0026thinsp;7.4\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e84.3\u0026thinsp;\u0026plusmn;\u0026thinsp;7.0\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e0.009\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e0.57\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003eInsulin (mU/L)\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e7.50\u0026thinsp;\u0026plusmn;\u0026thinsp;5.02\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e5.91\u0026thinsp;\u0026plusmn;\u0026thinsp;3.79\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e6.39\u0026thinsp;\u0026plusmn;\u0026thinsp;3.09\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e7.95\u0026thinsp;\u0026plusmn;\u0026thinsp;4.35\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e0.12\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e0.015\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003eHOMA-IR\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e1.67\u0026thinsp;\u0026plusmn;\u0026thinsp;1.00\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e1.23\u0026thinsp;\u0026plusmn;\u0026thinsp;0.80\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e1.31\u0026thinsp;\u0026plusmn;\u0026thinsp;0.72\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e1.68\u0026thinsp;\u0026plusmn;\u0026thinsp;1.01\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e0.03\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e0.01\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colspan=\"7\" nameend=\"c7\" namest=\"c1\"\u003e\u003cp\u003eData are presented as mean\u0026thinsp;\u0026plusmn;\u0026thinsp;standard deviation.\u003c/p\u003e\u003cp\u003e*: Independent sample t-test was applied for normally distributed variables, and Mann Whitney U test was applied for non-normally distributed variables, comparing patients with controls at baseline.\u003c/p\u003e\u003cp\u003e**: Repeated measures analysis of variance (ANOVA) was applied for normally distributed variables, and Friedman test was applied for non-normally distributed variables, comparing variables within patients for trend.\u003c/p\u003e\u003cp\u003e\u003csup\u003ea\u003c/sup\u003e: p\u0026thinsp;\u0026lt;\u0026thinsp;0.05 compared to baseline (pairwise comparisons with Bonferroni correction).\u003c/p\u003e\u003cp\u003e\u003cb\u003eAbbreviations\u003c/b\u003e: ALT, Alanine Aminotransferase; AST, Aspartate Aminotransferase; BMI, Body Mass Index; γ-GT, γ-Glutamyl Transferase; HDL-C, High Density Lipoprotein-Cholesterol; LDL-C, Low Density Lipoprotein-Cholesterol; TG, Triglycerides; TC, Total Cholesterol.\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e\u003cp\u003eThe comparisons between the control and intervention groups at baseline are presented in Tables\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e and \u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e. HDL-C, AST, ALT, glucose, and HOMA-IR concentrations were lower, whereas the AST/ALT ratio was higher in the intervention group than in the control group. No significant differences were observed in any of the other variables between the control and intervention groups at baseline, including all non-invasive indices (Tables\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e and \u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e). In the linear regression analysis, after adjustment for age (model 1) or age and log(HOMA-IR) (model 2), the comparisons between the controls and patients at baseline remained non-significant for all non-invasive indices (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e). Model 2 was not applied to TyG, TyG-BMI, and MetS-IR to avoid overadjustment because these indices, as well as HOMA-IR, include glucose in their equations.\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003eBaseline and endpoint of non-invasive indices of hepatic steatosis and fibrosis.\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"11\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c8\" colnum=\"8\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c9\" colnum=\"9\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c10\" colnum=\"10\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c11\" colnum=\"11\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eControls (n\u0026thinsp;=\u0026thinsp;27)\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colspan=\"3\" nameend=\"c5\" namest=\"c3\"\u003e\u003cp\u003ePatients (n\u0026thinsp;=\u0026thinsp;21)\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colspan=\"3\" nameend=\"c8\" namest=\"c6\"\u003e\u003cp\u003eP-value\u003c/p\u003e\u003cp\u003e(between group comparison at baseline)\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colspan=\"3\" nameend=\"c11\" namest=\"c9\"\u003e\u003cp\u003eTrend for p-value\u003c/p\u003e\u003cp\u003e(within intervention group comparison)\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e\u003cp\u003eUnadjusted*\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c7\"\u003e\u003cp\u003emodel 1\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c8\"\u003e\u003cp\u003emodel 2\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c9\"\u003e\u003cp\u003eUnadjusted**\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c10\"\u003e\u003cp\u003emodel 3\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c11\"\u003e\u003cp\u003emodel 4\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eBaseline\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u003cp\u003eBaseline\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e\u003cp\u003eMonth 6\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e\u003cp\u003eEndpoint\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c7\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c8\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c9\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c10\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c11\"\u003e\u0026nbsp;\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003eHSI\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e31.4\u0026thinsp;\u0026plusmn;\u0026thinsp;4.3\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e31.3\u0026thinsp;\u0026plusmn;\u0026thinsp;5.8\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e29.8\u0026thinsp;\u0026plusmn;\u0026thinsp;6.0\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e30.1\u0026thinsp;\u0026plusmn;\u0026thinsp;5.1\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e0.53\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e0.83\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e0.46\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e0.07\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e0.07\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003e0.06\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003eTyG\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e8.00\u0026thinsp;\u0026plusmn;\u0026thinsp;0.53\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e8.10\u0026thinsp;\u0026plusmn;\u0026thinsp;0.37\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e7.97\u0026thinsp;\u0026plusmn;\u0026thinsp;0.61\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e7.95\u0026thinsp;\u0026plusmn;\u0026thinsp;0.37\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e0.50\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e0.56\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e***\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e0.17\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e0.15\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003e***\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003eTyG-BMI\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e185.8\u0026thinsp;\u0026plusmn;\u0026thinsp;34.6\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e195.0\u0026thinsp;\u0026plusmn;\u0026thinsp;43.8\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e193.3\u0026thinsp;\u0026plusmn;\u0026thinsp;51.2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e199.9\u0026thinsp;\u0026plusmn;\u0026thinsp;44.2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e0.48\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e0.49\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e***\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e0.71\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e0.71\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003e***\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003eTG/HDL-C\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e0.65\u0026thinsp;\u0026plusmn;\u0026thinsp;0.52\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e0.82\u0026thinsp;\u0026plusmn;\u0026thinsp;0.36\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e0.70\u0026thinsp;\u0026plusmn;\u0026thinsp;0.52\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e0.62\u0026thinsp;\u0026plusmn;\u0026thinsp;0.27\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e0.18\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e0.23\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e0.10\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e0.08\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e0.06\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003e0.07\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003eMetS-IR\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e32.3\u0026thinsp;\u0026plusmn;\u0026thinsp;6.4\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e34.8\u0026thinsp;\u0026plusmn;\u0026thinsp;7.7\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e33.3\u0026thinsp;\u0026plusmn;\u0026thinsp;9.0\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e35.0\u0026thinsp;\u0026plusmn;\u0026thinsp;8.1\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e0.17\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e0.25\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e***\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e0.04\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e0.04\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003e***\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003eDSI\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e-3.01\u0026thinsp;\u0026plusmn;\u0026thinsp;0.87\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e-2.80\u0026thinsp;\u0026plusmn;\u0026thinsp;1.08\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e-3.14\u0026thinsp;\u0026plusmn;\u0026thinsp;1.13\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e-3.10\u0026thinsp;\u0026plusmn;\u0026thinsp;0.96\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e0.68\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e0.51\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e0.63\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e0.12\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e0.10\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003e0.09\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003eDSI-NAFLD risk (%)\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e6.5\u0026thinsp;\u0026plusmn;\u0026thinsp;6.8\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e8.9\u0026thinsp;\u0026plusmn;\u0026thinsp;10.6\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e7.2\u0026thinsp;\u0026plusmn;\u0026thinsp;9.1\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e6.2\u0026thinsp;\u0026plusmn;\u0026thinsp;6.2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e0.66\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e0.39\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e0.07\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e0.37\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e0.36\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003e0.33\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003eFSI\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e-3.22\u0026thinsp;\u0026plusmn;\u0026thinsp;0.79\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e-2.96\u0026thinsp;\u0026plusmn;\u0026thinsp;0.94\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e-2.99\u0026thinsp;\u0026plusmn;\u0026thinsp;1.15\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e-2.90\u0026thinsp;\u0026plusmn;\u0026thinsp;0.97\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e0.23\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e0.24\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e0.04\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e0.93\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e0.92\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003e0.93\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003eBARD\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e2.15\u0026thinsp;\u0026plusmn;\u0026thinsp;0.36\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e2.05\u0026thinsp;\u0026plusmn;\u0026thinsp;0.59\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e2.19\u0026thinsp;\u0026plusmn;\u0026thinsp;0.40\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e2.26\u0026thinsp;\u0026plusmn;\u0026thinsp;0.45\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e0.69\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e0.33\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e0.54\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e0.08\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e0.05\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003e0.07\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colspan=\"11\" nameend=\"c11\" namest=\"c1\"\u003e\u003cp\u003eData are presented as mean\u0026thinsp;\u0026plusmn;\u0026thinsp;standard deviation.\u003c/p\u003e\u003cp\u003e*: Student\u0026rsquo;s t-test was applied for normally distributed variables, and Mann Whitney U test was applied for non-normally distributed variables, comparing patients and controls at baseline.\u003c/p\u003e\u003cp\u003e**: Repeated measures ANOVA was applied for normally distributed variables, and Friedman test was applied for non-normally distributed variables, comparing variables within patients for trend.\u003c/p\u003e\u003cp\u003e***: Due to inclusion of glucose in the equations of HOMA-IR and the annotated non-invasive indices, models 2 and 4 were not applied in the respective indices.\u003c/p\u003e\u003cp\u003eModel 1: adjustment for age; linear regression analysis.\u003c/p\u003e\u003cp\u003eModel 2: adjustment for age and log(HOMA-IR); linear regression analysis.\u003c/p\u003e\u003cp\u003eModel 3: adjustment for age; repeated measures ANCOVA.\u003c/p\u003e\u003cp\u003eModel 4: adjustment for age and log(HOMA-IR); repeated measures ANCOVA.\u003c/p\u003e\u003cp\u003e\u003cb\u003eAbbreviations\u003c/b\u003e: ALT, Alanine Aminotransferase; ANCOVA, analysis of covariance; AST, Aspartate Aminotransferase; BARD, ΒΜΙ \u0026ndash; AST/ALT Ratio Diabetes mellitus; BMI, Body Mass Index; DSI, Dallas Steatosis Index; FSI, Framingham Steatosis Index; HSI, Hepatic Steatosis Index; MetS-IR, Metabolic Score for Insulin Resistance; NAFLD, Nonalcoholic Fatty Liver Disease; TG/HDL, Triglycerides/High Density Lipoprotein; TyG, Triglyceride-Glucose index.\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e\u003cp\u003eWithin the intervention group, TC, LDL-C, HDL-C, TC, AST/ALT ratio, insulin, and HOMA-IR increased overall during the study period (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). In pairwise comparisons (after Bonferroni correction), TC and LDL-C concentrations were increased at 6 months (p\u0026thinsp;=\u0026thinsp;0.005 and p\u0026thinsp;=\u0026thinsp;0.008, respectively) and at the endpoint (p\u0026thinsp;=\u0026thinsp;0.007 and p\u0026thinsp;=\u0026thinsp;0.005, respectively) compared to baseline. HDL-C concentrations increased at 6 months compared to the baseline (p\u0026thinsp;=\u0026thinsp;0.005). AST/ALT concentrations were increased at the endpoint compared to those at baseline (p\u0026thinsp;=\u0026thinsp;0.001). Insulin and HOMA-IR did not marginally provide statistically significant changes in the pairwise comparisons (p\u0026thinsp;=\u0026thinsp;0.057 and p\u0026thinsp;=\u0026thinsp;0.053, respectively, between baseline and endpoint), despite an overall significant trend towards an increase over time. Importantly, significant improvement at the endpoint compared to month 6 was not observed for any of the variables of interest (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). No significant changes were observed in other variables of interest (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). Regarding non-invasive indices of hepatic steatosis or fibrosis (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e), MetS-IR showed an overall significant change (p\u0026thinsp;=\u0026thinsp;0.04); in pairwise comparisons (after Bonferroni correction), no significant difference in MetS-IR was observed. In the repeated-measures ANCOVA, after adjusting for age (Model 3), the overall significant trend remained robust for MetS-IR, providing a marginally significant trend. The error bars for all non-invasive indices for the control group (baseline) and intervention group (baseline, month 6, and endpoint) are depicted in Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e.\u003c/p\u003e\u003cp\u003e\u003c/p\u003e"},{"header":"4. Discussion","content":"\u003cp\u003eIn this study, non-invasive indices of hepatic steatosis and fibrosis were evaluated during a 6-month of goserelin treatment and its discontinuation in women with biopsy-proven endometriosis. All non-invasive indices of hepatic steatosis, apart from MetS-IR, as well as BARD, a non-invasive index of hepatic fibrosis, remained unchanged after goserelin treatment. MetS-IR showed an overall trend towards a decrease at month 6, but this was not statistically significant in pairwise comparisons vs. baseline or endpoint, after appropriate correction.\u003c/p\u003e\u003cp\u003eThese findings do not support our initial hypothesis that estrogen deficiency caused by goserelin, a GnRH analog, could worsen MASLD, as observed in natural or surgical menopause [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e]. The study design does not allow us to draw secure conclusions regarding the apparently null effect of goserelin on the non-invasive indices of MASLD. We may assume that a 6-month period of estrogen deficiency may not be adequate to affect the respective, non-invasive indices. Of course, a yet unknown direct beneficial effect of goserelin, which may counterbalance its indirect effect through estrogen deficiency on hepatic steatosis, cannot be excluded.\u003c/p\u003e\u003cp\u003eThis study is the first to evaluate hepatic steatosis and fibrosis non-invasive indices in women with endometriosis treated with goserelin post-surgically. To date, no studies have evaluated the impact of goserelin on MASLD, while previous studies evaluating the effects of GnRH analogs on MASLD in other conditions are few, and methodologically highly heterogeneous. In women, there is only a case report of a girl with central precocious puberty, whose histologically confirmed MASH worsened after leuprorelin treatment, while partial improvement was observed after treatment [\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e]. Three studies in men with prostate cancer receiving GnRH agonists or antagonists, that is, androgen deprivation therapy, reported significant progression of hepatic steatosis [\u003cspan additionalcitationids=\"CR23\" citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e]. First, ultrasonographic worsening of hepatic steatosis was shown after a 32-month leuprorelin treatment in a man with prostate cancer; however, this was a much longer duration than that of our study [\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e]. Subsequently, in a registry-based study, men who received androgen deprivation therapy for localized prostate cancer were shown to have higher rates of MASLD during follow-up [\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e]. In another study, hepatic steatosis, evaluated using computed tomography, worsened in men with prostate cancer receiving androgen deprivation therapy for 6 months [\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e]. Different medications, diseases, and sexes render the comparison of these studies with our study hardly feasible.\u003c/p\u003e\u003cp\u003eIt is important to note that, based on the values of all non-invasive indices of hepatic steatosis (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e), the rates of hepatic steatosis were low in both patients and controls, possibly due to the young age and the lack of concomitant diseases and medications of the participants, compared to the general population or other epidemiological studies in patients with endometriosis [\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e, \u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eAs a secondary observation, the overall lipid profile was adversely affected by iatrogenic temporary menopause (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e), as previously described, and remained above the baseline concentrations 6 months after menstruation restoration [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e]. The same applies to insulin resistance, at least as evaluated with HOMA-IR. A relevant observation is that these changes were not reversed during menstruation restoration (month 6 to the endpoint), implying that even a 6-month treatment with GnRH analogs may infer longer-term adverse metabolic changes [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eThe strengths of our study include its originality, as there are no known completed or ongoing studies assessing the association between GnRH-induced estrogen deficiency and MASLD in patients with endometriosis. Moreover, the sample consisted of young premenopausal women without comorbidities or concomitant medications that might have affected MASLD. Furthermore, the study consisted of two periods: the first (6 months) characterized by estrogen deficiency and the second (6 months) by estrogen replenishment, during which the effect of goserelin is expected to be fully washed off. The current study has some limitations. First, the study was not specifically designed to show the effect of goserelin on non-invasive indices of hepatic steatosis and fibrosis, but was a \u003cem\u003epost-hoc\u003c/em\u003e analysis of a study design with other aims [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]. Second, the control group was evaluated only at baseline. Third, the controls were chosen from a different pool; thus, there was a risk of selection bias. Ideally, we should have recruited women with endometriosis who were not subjected to any treatment for six months, which was difficult to perform in the setting of a \u003cem\u003epost-hoc\u003c/em\u003e analysis. Fourth, the sample size maybe small and no a \u003cem\u003epriori\u003c/em\u003e power analysis had been performed, largely owing to the fact that this was a post-hoc analysis, so we selected all available data of the mother study [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]. Furthermore, the time frame may be relatively short, since 6 months of goserelin-induced estrogen deficiency may not be sufficient for the development of hepatic steatosis and, much more, fibrosis. Moreover, the lack of paired liver biopsies or an appropriate imaging technique to quantify hepatic steatosis or fibrosis at baseline, month 6, and the endpoint is another limitation, largely owing to the retrospective retrieval of data from the intervention group. Due to the same reason, the lack of certain parameters, such as platelet count, did not allow us to calculate other non-invasive indices, especially of fibrosis (e.g., fibrosis-4 index, NAFLD fibrosis score); however, as mentioned above, we consider the 6-month period of estrogen deficiency a short period for the development of hepatic fibrosis.\u003c/p\u003e\u003cp\u003eIn conclusion, this study showed that the non-invasive indices of hepatic steatosis and BARD, a non-invasive index of fibrosis, were not significantly affected by the estrogen deficiency caused by 6-month treatment with goserelin in premenopausal women with endometriosis. Although further clinical studies based on histological confirmation or appropriate imaging of hepatic steatosis or fibrosis are needed to draw more secure results, the results of this study may imply that goserelin may be considered in patients with endometriosis and MASLD, because it is not expected to worsen MASLD when administered for a short period.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eEthical approval\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe study was approved by the ethics committee of the 424 General Military Hospital (16329/04-09-2018). All procedures performed in the study were in accordance with the ethical standards of the institutional research committees and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eInformed consent\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eInformed consent was retrospectively obtained from all participants for the use of their data for the purpose of the study.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eConception and design of the work: Dimitrios A. Anastasilakis, Athanasios D. Anastasilakis, Stergios A. Polyzos\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eAcquisition of data: Dimitrios A. Anastasilakis, Athanasios D. Anastasilakis, Athina Gkiomisi, Stergios Papachatzopoulos\u003c/p\u003e\n\u003cp\u003eAnalysis of data: Dimitrios A. Anastasilakis, Anastasios Semertzidis, Stergios A. Polyzos\u003c/p\u003e\n\u003cp\u003eInterpretation of data: All authors\u003c/p\u003e\n\u003cp\u003eDrafting the article: Dimitrios A. Anastasilakis\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eRevising the article critically for important intellectual content: All authors\u003c/p\u003e\n\u003cp\u003eFinal approval of the submitted version: All authors\u003c/p\u003e\n\u003cp\u003eAgree to be accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved: All authors\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAvailability of data and material\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eData will be available upon reasonable request by the corresponding author.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cbr\u003e \u003c/strong\u003e\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eM.E. Rinella, J.V. Lazarus, V. Ratziu, S.M. 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Pharmacol. \u003cb\u003e55\u003c/b\u003e, 848\u0026ndash;853 (2015)\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eA.D. Anastasilakis, S.A. Polyzos, P.A. Vorkas, A. Gkiomisi, M.P. Yavropoulou, M. Rauner, P. Nikolakopoulos, S. Papachatzopoulos, P. Makras, S. Gerou, L.C. Hofbauer, A. Palermo, E. Tsourdi, Lipid Profile after Pharmacologic Discontinuation and Restoration of Menstruation in Women with Endometriosis: A 12-Month Observational Prospective Study. J. Clin. Med. \u003cb\u003e12\u003c/b\u003e, (2023)\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eJ.H. Lee, D. Kim, H.J. Kim, C.H. Lee, J.I. Yang, W. Kim, Y.J. Kim, J.H. Yoon, S.H. Cho, M.W. Sung, H.S. Lee, Hepatic steatosis index: a simple screening tool reflecting nonalcoholic fatty liver disease. Dig. Liver Dis. \u003cb\u003e42\u003c/b\u003e, 503\u0026ndash;508 (2010)\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eS. Zhang, T. Du, J. Zhang, H. Lu, X. Lin, J. Xie, Y. Yang, X. 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Gogia, B.A. Neuschwander-Tetri, Development and validation of a simple NAFLD clinical scoring system for identifying patients without advanced disease. Gut. \u003cb\u003e57\u003c/b\u003e, 1441\u0026ndash;1447 (2008)\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eY. Sasaki, H. Kajino, H. Gotoda, A Case of Nonalcoholic Steatohepatitis in Central Precocious Puberty Aggravated by Gonadotropin-releasing Hormone Analog. JPGN Rep. \u003cb\u003e1\u003c/b\u003e, e014 (2020)\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eP. Gild, A.P. Cole, A. Krasnova, B.A. Dickerman, N. von Landenberg, M. Sun, L.A. Mucci, S.R. Lipsitz, F.K. Chun, P.L. Nguyen, A.S. Kibel, T.K. Choueiri, S. Basaria, Q.D. Trinh, Liver Disease in Men Undergoing Androgen Deprivation Therapy for Prostate Cancer. J. Urol. \u003cb\u003e200\u003c/b\u003e, 573\u0026ndash;581 (2018)\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eC. Gabbi, F. Carubbi, L. Losi, P. Loria, M. Costantini, M. Bertolotti, N. Carulli, Nonalcoholic fatty liver disease induced by leuprorelin acetate. J. Clin. Gastroenterol. \u003cb\u003e42\u003c/b\u003e, 107\u0026ndash;110 (2008)\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eH. Sawazaki, Y. Kitamura, K. Yagi, Y. Arai, Impact of Androgen Deprivation Therapy on Non-Alcoholic Fatty Liver Disease in Patients with Prostate Cancer: A CT Evaluation. Urol. Int. \u003cb\u003e104\u003c/b\u003e, 425\u0026ndash;430 (2020)\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eH. Huang, Z. Liu, J. Ruan, Z. Fang, C. Xu, Laparoscopically confirmed endometriosis and the risk of incident NAFLD: a prospective cohort study. Reprod. Biol. Endocrinol. \u003cb\u003e23\u003c/b\u003e, 55 (2025)\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eE.S. Makri, E. Makri, S.A. Polyzos, Combination Therapies for Nonalcoholic Fatty Liver Disease. J. Pers. Med. \u003cb\u003e12\u003c/b\u003e, (2022)\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"endocrine","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"endo","sideBox":"Learn more about [Endocrine](https://www.springer.com/journal/12020)","snPcode":"12020","submissionUrl":"https://submission.nature.com/new-submission/12020/3","title":"Endocrine","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false},"keywords":"menopause, fibrosis, goserelin, metabolic dysfunction-associated steatotic liver disease, metabolic dysfunction-associated steatohepatitis, nonalcoholic fatty liver disease, non-invasive indices, steatosis","lastPublishedDoi":"10.21203/rs.3.rs-7566446/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-7566446/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003ch2\u003ePurpose\u003c/h2\u003e\u003cp\u003eMenopause has been associated with metabolic dysfunction-associated steatotic liver disease (MASLD). This study primarily aimed to evaluate changes in non-invasive indices of hepatic steatosis in premenopausal women with endometriosis after treatment with goserelin resulting in pharmacologic menopause.\u003c/p\u003e\u003ch2\u003eMethods\u003c/h2\u003e\u003cp\u003eThis was a \u003cem\u003epost-hoc\u003c/em\u003e study of an interventional study. Non-invasive indices of hepatic steatosis and, secondarily, fibrosis were calculated in premenopausal women with endometriosis (intervention group, n\u0026thinsp;=\u0026thinsp;21) before (baseline), 6 months after goserelin treatment (estrogen-deficient period), and 6 months after goserelin discontinuation (estrogen replenishment period). Matched premenopausal women were recruited for baseline comparisons (control group, n\u0026thinsp;=\u0026thinsp;27). Steatosis was evaluated using the Hepatic Steatosis Index (HSI), Triglyceride/Glucose Index (TyG), TyG-Body Mass Index (BMI), Triglyceride/High Density Lipoprotein (TG/HDL), Metabolic Score for Insulin Resistance (MetS-IR), Dallas Steatosis Index (DSI), and Framingham Steatosis Index (FSI), and fibrosis with BMI \u0026ndash; Aspartate Aminotransferase (AST)/Alanine Aminotransferase (ALT) Ratio Diabetes Mellitus (BARD).\u003c/p\u003e\u003ch2\u003eResults\u003c/h2\u003e\u003cp\u003eThe non-invasive indices of hepatic steatosis and BARD were similar between patients and controls at baseline and did not significantly change after goserelin treatment, except for MetS-IR, which showed an overall significant change (p\u0026thinsp;=\u0026thinsp;0.04); however, in pairwise comparisons after Bonferroni correction, no significant difference in MetS-IR was observed between baseline (34.8\u0026thinsp;\u0026plusmn;\u0026thinsp;7.7), month 6 (33.3\u0026thinsp;\u0026plusmn;\u0026thinsp;9.0), and endpoint (35.0\u0026thinsp;\u0026plusmn;\u0026thinsp;8.1) values. As expected, the lipid profiles and insulin resistance worsened during the study.\u003c/p\u003e\u003ch2\u003eConclusion\u003c/h2\u003e\u003cp\u003eNon-invasive hepatic steatosis and BARD indices were not significantly affected by estrogen deficiency caused by 6-month goserelin treatment in premenopausal women with endometriosis.\u003c/p\u003e","manuscriptTitle":"Effect of goserelin acetate on non-invasive indices of hepatic steatosis in women with endometriosis","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-09-26 06:24:56","doi":"10.21203/rs.3.rs-7566446/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2025-10-11T07:56:46+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-10-09T16:32:02+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"274740201777217296462754154253498239611","date":"2025-09-21T12:43:38+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2025-09-16T09:42:46+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2025-09-10T04:17:16+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2025-09-10T04:15:35+00:00","index":"","fulltext":""},{"type":"submitted","content":"Endocrine","date":"2025-09-08T16:48:14+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"endocrine","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"endo","sideBox":"Learn more about [Endocrine](https://www.springer.com/journal/12020)","snPcode":"12020","submissionUrl":"https://submission.nature.com/new-submission/12020/3","title":"Endocrine","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"3e8a91b6-6e1d-4bed-b4e3-fa31014980c0","owner":[],"postedDate":"September 26th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2026-01-05T16:00:39+00:00","versionOfRecord":{"articleIdentity":"rs-7566446","link":"https://doi.org/10.1007/s12020-025-04511-5","journal":{"identity":"endocrine","isVorOnly":false,"title":"Endocrine"},"publishedOn":"2025-12-29 15:57:53","publishedOnDateReadable":"December 29th, 2025"},"versionCreatedAt":"2025-09-26 06:24:56","video":"","vorDoi":"10.1007/s12020-025-04511-5","vorDoiUrl":"https://doi.org/10.1007/s12020-025-04511-5","workflowStages":[]},"version":"v1","identity":"rs-7566446","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-7566446","identity":"rs-7566446","version":["v1"]},"buildId":"0SHbDDIpRTBOrFPTvp6pu","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}