Variations in serum ferritin levels in women with polycystic ovarian syndrome undergoing assisted reproduction.

Dhanya Vikhnan : Investigation, Validation, Writing-Original draft, Data curation; Vanishree V. Madhvacharya : Investigation, Validation, Writing-Original draft, Formal analysis; Anjali Mundkur : Data acquisition; Vidyashree G. Poojari : Data acquisition; R Vani Lakshmi: Validation, Formal analysis, Data curation; Pratap K. Narayan : Methodology, Supervision; Satish K. Adiga: Methodology, Data acquisition; Supervision; Prashanth K. Adiga : Methodology, Writing-Review & Editing, Supervision; Guruprasad Kalthur : Conceptualization, Methodology, Writing-Review & Editing, Supervision.

The authors gratefully acknowledge financial support from the 10.13039/501100001411 Indian Council of Medical Research (ICMR, No. EM/Dev/SG/85/1642/2023).

The study was approved by Kasturba Medical College and Kasturba Hospital, Institutional Ethics Committee (IEC2: 558/2022 and IEC1: 385/2022).

The mean age of patients was significantly lower in the PCOS group ( p < 0.05) compared with the non-PCOS group. Type of infertility, duration of infertility, and the distribution of male factor infertility were comparable in both groups. The prevalence of menstrual irregularities ( p < 0.0001) and BMI ( p < 0.01) were significantly higher in the PCOS group compared with the non-PCOS group ( Table 2 ). The distribution of gonadotropin types did not differ significantly between the PCOS and non-PCOS groups ( p = 0.114), indicating that there was no statistically significant association between the two variables ( Table 1 ). Table 2 Clinico-demographic data of the study participants. Table 2 Parameters Non-PCOS (N = 50) PCOS (N = 40) p-value b Mean ± SD Median (Q1, Q3) Mean ± SD Median (Q1, Q3) Age (years) 32.56 ± 3.79 33 (5) 30.82 ± 3.70 31 (5) 0.032 BMI (kg/m 2 ) 22.93 ± 3.88 23 (6) 25.40 ± 4.56 25 (7) 0.007 Duration of infertility (years) 5.63 ± 3.01 5 (5) 5.69 ± 2.99 5 (5) 0.948 (ns) Type of infertility a Primary 42 (84) 38 (95) 0.175 (ns) Secondary 8 (16) 2 (5) Menstrual history a Regular 47 (94) 12 (30) <0.001 Irregular 3 (6) 28 (70) Male infertility a Present 19 (38) 14 (35) 0.769 (ns) Absent 31 (62) 26 (65) a Categorical variables are expressed using counts and percentages. b ns indicates non-significant differences. Clinico-demographic data of the study participants. Categorical variables are expressed using counts and percentages. ns indicates non-significant differences. Baseline serum E2 levels did not differ significantly between the PCOS and non-PCOS groups. However, AMH ( p < 0.0001) and LH ( p < 0.001) were significantly elevated, whereas FSH levels were reduced ( p < 0.01) in the PCOS group, resulting in a markedly increased LH to FSH ratio ( p < 0.0001) ( Figure 2 A–E). Furthermore, the total gonadotropin dose administered was significantly lower in PCOS women ( p < 0.0001) compared with non-PCOS women ( Figure 2 F). On day 5 of ovarian stimulation, PCOS patients exhibited a significant rise in serum E2 levels ( p < 0.0001) , which continued to be significantly elevated even on day 9 of stimulation in the PCOS group ( p < 0.0001) ( Figure 2 G). LH levels were significantly higher in PCOS patients on day 5 ( p < 0.05), but no significant difference was observed between the two groups on day 9 ( Figure 2 H). Figure 2 Demographic data and baseline hormone profile on day 2 of the menstrual cycle in PCOS and non-PCOS women undergoing ART treatment. Reproductive hormone levels in serum: (A) AMH, (B) E2, (C) LH, (D) FSH, and (E) LH to FSH ratio. (F) Total gonadotropin dose administered. (G) Serum E2 levels on day 2, 5, and 9 of the menstrual cycle. (H) Serum LH levels on day 2, 5, and 9 of the menstrual cycle. Red dots represent the non-PCOS group and blue dots represent the PCOS group. Data represent mean ± standard error of mean. For AMH, E2, LH, and gonadotropin dose data, sample size in non-PCOS = 50 and PCOS = 40. For FSH and LH/FSH ratio, sample size in non-PCOS = 50 and PCOS = 26. Figure 2 Demographic data and baseline hormone profile on day 2 of the menstrual cycle in PCOS and non-PCOS women undergoing ART treatment. Reproductive hormone levels in serum: (A) AMH, (B) E2, (C) LH, (D) FSH, and (E) LH to FSH ratio. (F) Total gonadotropin dose administered. (G) Serum E2 levels on day 2, 5, and 9 of the menstrual cycle. (H) Serum LH levels on day 2, 5, and 9 of the menstrual cycle. Red dots represent the non-PCOS group and blue dots represent the PCOS group. Data represent mean ± standard error of mean. For AMH, E2, LH, and gonadotropin dose data, sample size in non-PCOS = 50 and PCOS = 40. For FSH and LH/FSH ratio, sample size in non-PCOS = 50 and PCOS = 26. The median serum ferritin level in the entire study population (including PCOS and non-PCOS women) was determined as 34.1 ng/mL. The median serum ferritin level was 27.5 ng/mL in the PCOS group, and it was higher in the non-PCOS group but not significantly (37.3 ng/mL). High serum ferritin levels (above the median value) were observed in 42.2 % of PCOS patients and 57.8 % of non-PCOS patients ( Table 3 ). However, the difference in the ferritin levels was not significant between these two groups ( Figure 3 A). Furthermore, there were no significant differences in the hemoglobin concentration or hematocrit value between the PCOS and non-PCOS groups ( Figure 3 B,C). Table 3 Association between serum ferritin levels across PCOS and Non-PCOS groups. Table 3 Serum Ferritin Group Total Chi square test statistic p value Non-PCOS (N = 50) PCOS (N = 40) Low a 24 (53.3) 21 (46.7) 45 (100) 0.18 0.67 High a 26 (57.8) 19 (42.2) 45 (100) Total a 50 (55.6) 40 (44.4) 90 (100) a Subgrouping of serum ferritin levels into low and high categories based on the median ferritin values in the PCOS and non-PCOS groups (Median serum ferritin = 34.1 ng/mL). Categorical variables expressed using counts and percentages. Figure 3 Iron status markers assessed in PCOS and non-PCOS women undergoing ART treatment. (A) Serum ferritin assessed on day 2/day 3 of the menstrual cycle. (B) Hemoglobin level. (C) Hematocrit assessed on day 5/day 9 of ovarian stimulation. Data for serum ferritin and hemoglobin represent mean ± standard error of mean and hematocrit data represent percentages. Sample size in non-PCOS = 50 and PCOS = 40. Figure 3 Association between serum ferritin levels across PCOS and Non-PCOS groups. Subgrouping of serum ferritin levels into low and high categories based on the median ferritin values in the PCOS and non-PCOS groups (Median serum ferritin = 34.1 ng/mL). Categorical variables expressed using counts and percentages. Iron status markers assessed in PCOS and non-PCOS women undergoing ART treatment. (A) Serum ferritin assessed on day 2/day 3 of the menstrual cycle. (B) Hemoglobin level. (C) Hematocrit assessed on day 5/day 9 of ovarian stimulation. Data for serum ferritin and hemoglobin represent mean ± standard error of mean and hematocrit data represent percentages. Sample size in non-PCOS = 50 and PCOS = 40. AFC and the number of follicles aspirated from PCOS patients during oocyte retrieval were significantly higher ( p < 0.0001) compared with non-PCOS women. However, the follicular output rate (FORT; ratio of pre-ovulatory follicle count on the day of hCG administration relative to the baseline AFC) did not differ between the two groups ( Figure 4 A–C). The total number of oocytes retrieved was significantly higher in PCOS patients ( p < 0.0001) ( Figure 4 D). However, compared with non-PCOS patients, no significant differences were observed in the oocyte maturation rate, fertilization rate, and number of transferrable embryos (grade 1) on day 3 post-fertilization in PCOS patients ( Figure 4 E–G). Figure 4 Embryological parameters assessed in PCOS and non-PCOS women undergoing ART treatment: (A) AFC; (B) number of follicles tapped; (C) FORT; (D) total number of oocytes retrieved per woman; (E) percentage of germinal vesicle (GV), metaphase I (MI), and metaphase II (MII) oocytes; (F) fertilization rate; and (G) percentage of transferable embryos on day 3. Red dots represent non-PCOS group and blue dots represent PCOS group. Data for AFC, number of follicles, and oocytes represent mean ± standard error of mean. Data for maturation rate, fertilization rate, and transferable embryos represent percentages. For AFC, number of follicles tapped, FORT, and number and percentage of oocytes retrieved, sample size in non-PCOS = 50 and PCOS = 40. For transferable embryos, sample size in non-PCOS = 49 and PCOS = 39. Figure 4 Embryological parameters assessed in PCOS and non-PCOS women undergoing ART treatment: (A) AFC; (B) number of follicles tapped; (C) FORT; (D) total number of oocytes retrieved per woman; (E) percentage of germinal vesicle (GV), metaphase I (MI), and metaphase II (MII) oocytes; (F) fertilization rate; and (G) percentage of transferable embryos on day 3. Red dots represent non-PCOS group and blue dots represent PCOS group. Data for AFC, number of follicles, and oocytes represent mean ± standard error of mean. Data for maturation rate, fertilization rate, and transferable embryos represent percentages. For AFC, number of follicles tapped, FORT, and number and percentage of oocytes retrieved, sample size in non-PCOS = 50 and PCOS = 40. For transferable embryos, sample size in non-PCOS = 49 and PCOS = 39. Correlation analysis performed together for PCOS and non-PCOS groups found non-significant and very weak positive correlations between serum ferritin levels with BMI (r s = 0.082), and with serum reproductive hormonal parameters such as AMH (r s = 0.028), LH (r s = 0.055), and LH/FSH ratio (r s = 0.043) ( Figure 5 A–F). The gonadotropin dose required for ovarian stimulation was not correlated with the serum ferritin level in both the PCOS and non-PCOS groups (r s = 0.009) ( Figure 5 G). Furthermore, correlation analysis between serum ferritin and embryological parameters found very weak associations with AFC (r s = 0.086), number of oocytes retrieved (r s = 0.153), maturation rate (r s = −0.110), fertilization rate (r s = 0.027), and transferable embryos (r s = 0.067) ( Figure 5 H–L). No evidence of a linear relationship was found in the data. Similar patterns were observed for clinico-demographic data when analyses were performed separately for the PCOS and non-PCOS group. The maturation rate had a moderate negative correlation with serum ferritin in the PCOS group (r s = −0.313, p < 0.05), but no association was found in the non-PCOS group (r s = 0.020). Furthermore, the percentage of transferable embryos on day 3 had a weak negative correlation with serum ferritin in the non-PCOS group (r s = −0.235), whereas the opposite trend was found in the PCOS group, with a weak positive correlation (r s = 0.232). However, in both groups, the magnitudes of the correlation coefficients indicated weak associations with limited linear relevance ( Figure 6 A–N and 7 A–J). Figure 5 Correlations between serum ferritin levels and clinico-demographic, hormonal, and embryological parameters in PCOS and non-PCOS women. Correlations assessed in PCOS and non-PCOS women (combined analysis) between serum ferritin with: (A) BMI, (B) AMH, (C) E2, (D) FSH, (E) LH, (F) LH/FSH, (G) gonadotropin dose, (H) AFC, (I) number of oocytes retrieved per woman, (J) oocyte maturation rate, (K) fertilization rate, and (L) transferable embryos on day 3. For correlation analysis between serum ferritin with BMI, AMH, E2, LH, gonadotropin dose, AFC, number of oocytes retrieved per woman, oocyte maturation rate, and fertilization rate, sample size in non-PCOS = 50 and PCOS = 40. For correlation analysis between serum ferritin with FSH and LH/FSH ratio, sample size in non-PCOS = 50 and PCOS = 26. For correlation analysis between serum ferritin with transferable embryos on day 3, sample size in non-PCOS = 49 and PCOS = 39. Red dots represent non-PCOS group and blue dots represent PCOS group. Figure 5 Figure 6 Correlations between serum ferritin with clinico-demographic and hormonal data analyzed separately for PCOS and non-PCOS groups. Correlations between serum ferritin with: BMI in (A) non-PCOS, (B) PCOS; AMH in (C) non-PCOS, (D) PCOS; E2 in (E) non-PCOS, (F) PCOS ; FSH in (G) non-PCOS, (H) PCOS; LH in (I) non-PCOS, (J) PCOS; LH/FSH in (K) non-PCOS, (L) PCOS; gonadotropin dose in (M) non-PCOS, (N) PCOS, assessed on day 2 of the cycle. For correlation analysis between serum ferritin with BMI, AMH, E2, LH, and gonadotropin dose, sample size in non-PCOS = 50 and PCOS = 40. For correlation analysis between serum ferritin with FSH and LH/FSH ratio, sample size in non-PCOS = 50 and PCOS = 26. Figure 6 Figure 7 Correlations between serum ferritin with embryological parameters analyzed separately for PCOS and non-PCOS groups. Correlations between serum ferritin with: AFC in (A) non-PCOS, (B) PCOS; number of oocytes retrieved in (C) non-PCOS, (D) PCOS; oocyte maturation rate in (E) non-PCOS, (F) PCOS; fertilization rate in (G) non-PCOS, (H) PCOS; transferable embryos on day 3 in (I) non-PCOS, (J) PCOS. For correlation analysis between serum ferritin with AFC, number of oocytes retrieved per woman, oocyte maturation rate, and fertilization rate, sample size in non-PCOS = 50 and PCOS = 40. For correlation analysis between serum ferritin with transferable embryos on day 3, sample size in non-PCOS = 49 and PCOS = 39. Figure 7 Correlations between serum ferritin levels and clinico-demographic, hormonal, and embryological parameters in PCOS and non-PCOS women. Correlations assessed in PCOS and non-PCOS women (combined analysis) between serum ferritin with: (A) BMI, (B) AMH, (C) E2, (D) FSH, (E) LH, (F) LH/FSH, (G) gonadotropin dose, (H) AFC, (I) number of oocytes retrieved per woman, (J) oocyte maturation rate, (K) fertilization rate, and (L) transferable embryos on day 3. For correlation analysis between serum ferritin with BMI, AMH, E2, LH, gonadotropin dose, AFC, number of oocytes retrieved per woman, oocyte maturation rate, and fertilization rate, sample size in non-PCOS = 50 and PCOS = 40. For correlation analysis between serum ferritin with FSH and LH/FSH ratio, sample size in non-PCOS = 50 and PCOS = 26. For correlation analysis between serum ferritin with transferable embryos on day 3, sample size in non-PCOS = 49 and PCOS = 39. Red dots represent non-PCOS group and blue dots represent PCOS group. Correlations between serum ferritin with clinico-demographic and hormonal data analyzed separately for PCOS and non-PCOS groups. Correlations between serum ferritin with: BMI in (A) non-PCOS, (B) PCOS; AMH in (C) non-PCOS, (D) PCOS; E2 in (E) non-PCOS, (F) PCOS ; FSH in (G) non-PCOS, (H) PCOS; LH in (I) non-PCOS, (J) PCOS; LH/FSH in (K) non-PCOS, (L) PCOS; gonadotropin dose in (M) non-PCOS, (N) PCOS, assessed on day 2 of the cycle. For correlation analysis between serum ferritin with BMI, AMH, E2, LH, and gonadotropin dose, sample size in non-PCOS = 50 and PCOS = 40. For correlation analysis between serum ferritin with FSH and LH/FSH ratio, sample size in non-PCOS = 50 and PCOS = 26. Correlations between serum ferritin with embryological parameters analyzed separately for PCOS and non-PCOS groups. Correlations between serum ferritin with: AFC in (A) non-PCOS, (B) PCOS; number of oocytes retrieved in (C) non-PCOS, (D) PCOS; oocyte maturation rate in (E) non-PCOS, (F) PCOS; fertilization rate in (G) non-PCOS, (H) PCOS; transferable embryos on day 3 in (I) non-PCOS, (J) PCOS. For correlation analysis between serum ferritin with AFC, number of oocytes retrieved per woman, oocyte maturation rate, and fertilization rate, sample size in non-PCOS = 50 and PCOS = 40. For correlation analysis between serum ferritin with transferable embryos on day 3, sample size in non-PCOS = 49 and PCOS = 39.

This prospective study was conducted at the Department of Reproductive Medicine and Surgery, Kasturba Medical College, Manipal Academy of Higher Education (MAHE), Manipal, India, from April 2023 to June 2024. The study was approved by the Institutional Ethics Committee (IEC2: 558/2022 and IEC1: 385/2022), Kasturba Medical College & Kasturba Hospital, Manipal. Written informed consent was obtained from all participants. The study included women aged between 21 and 40 years (both inclusive) who were recruited for in vitro fertilization or intracytoplasmic sperm injection. All participants were assigned female at birth and self-identified as women. Diagnosis of PCOS was based on the modified Rotterdam criteria. 29 The study group of 90 participants consisted of 40 women with PCOS and 50 women without PCOS (normal ovarian reserve and unexplained infertility, with or without patent tube). Only those patients undergoing controlled ovarian stimulation with GnRH antagonist protocol were included in the study, whereas patients undergoing the agonist stimulation protocol and those with endometriosis, diminished ovarian reserve [anti-mullerian hormone (AMH) < 1.5 ng/mL], premature ovarian failure, and patients enrolled in the oocyte donation program were excluded. Patients who satisfied the inclusion criteria were categorized into PCOS and non-PCOS groups. Clinico-demographic data, including age, body mass index (BMI), type of infertility – primary (failure to achieve pregnancy in a person who has never conceived) or secondary (difficulty conceiving after at least one prior pregnancy) 30 – duration of infertility, menstrual history, and presence or absence of male factor infertility, were recorded prior to ovarian stimulation. Controlled ovarian stimulation was initiated on day 2 or day 3 of the menstrual cycle. 31 Prior to the onset of stimulation, serum ferritin, AMH, FSH, LH, and estradiol (E2) levels were calculated. Ferritin was assessed by the electrochemiluminescence immunoassay method using a specific immunoassay kit (04491785, Roche, Switzerland), with an inter-assay coefficient of variation of 4.9 %. The median ferritin value was calculated, and patients in each group were categorized into high and low ferritin subgroups. Recombinant FSH, highly purified human menopausal gonadotropin, or human menopausal gonadotropin were used for ovarian hyperstimulation ( Table 1 ), and the dosages were individualized based on ovarian reserve parameters, BMI, age, and socioeconomic status ( Figure 1 ). Table 1 Type of gonadotropin used for controlled ovarian stimulation. Table 1 Gonadotropin type Non-PCOS (N = 50) PCOS (N = 40) p-value rFSH a 32 (64) 33 (82.5) 0.114 HP- hMG b 13 (26) 4 (10) hMG c 5 (10) 3 (7.5) a rFSH- Recombinant follicle stimulating hormone. b HP-hMG- Highly purified human menopausal gonadotropin. c hMG- Human menopausal gonadotropin. Categorical variables expressed using counts and percentages. Figure 1 Experimental outline to understand the roles of serum ferritin in PCOS and non-PCOS women, and correlations with embryological outcomes. Figure created with BioRender.com . Figure 1 Type of gonadotropin used for controlled ovarian stimulation. rFSH- Recombinant follicle stimulating hormone. HP-hMG- Highly purified human menopausal gonadotropin. hMG- Human menopausal gonadotropin. Categorical variables expressed using counts and percentages. Experimental outline to understand the roles of serum ferritin in PCOS and non-PCOS women, and correlations with embryological outcomes. Figure created with BioRender.com . On day 5 of stimulation, serum E2 and LH were assessed, and follicular monitoring was initiated using transvaginal ultrasound (Voluson S8 BT22, GE Healthcare, USA; 8 MHz bandwidth probe). When serum E2 was ≥400 pg/mL and follicle size was either one leading follicle ≥14 mm or ≥6 follicles ≥11 mm, GnRH antagonist (Cetrorelix, Merck KGaA, Germany; or Ganirelix, Organon, The Netherlands) administration was initiated. From day 5 or 6 of stimulation, all patients received both gonadotropins and GnRH antagonist. Daily ultrasound monitoring was continued, until ≥3 follicles reached 18 mm. Final oocyte maturation was triggered, using either recombinant human chorionic gonadotropin (hCG, Ovitrelle, Merck KGaA, Germany) or GnRH agonist (Leuprolide, AbbVie, USA), depending on hormonal and ultrasound parameters. Oocyte retrieval was performed at 36 h after hCG trigger under intravenous anesthesia (Propofol) using a 17G single-lumen needle (K-MPIP-1035, Cook, USA) and a suction pump ( R29660 craft, Rocket Medical plc., UK) at a pressure of 120 mm Hg. Follicular fluids were scanned under a stereosome microscope and cumulus–oocyte complexes were identified. Cumulus–oocyte complexes were washed, incubated, and subjected to fertilization by in vitro fertilization or intracytoplasmic sperm injection. 31 Fertilization and embryo quality assessments were performed according to the Istanbul consensus on embryo assessment. 32 Correlation analysis was performed to assess the relationships between serum ferritin levels and various clinico-demographic variables and embryological outcomes. Based on the assumption of normality (assessed using the Shapiro–Wilks test) and homogeneity of variance (assessed using Levene's test), two-sample independent t -tests were conducted to compare the outcome characteristics across the two groups when both assumptions were satisfied. When the assumption of normality was violated, the Mann–Whitney U-test was employed. When the assumption of homogeneity of variance was not satisfied, Welch's t -test was used. All statistical analyses were conducted using jamovi (version 2.6) open-source statistical software (the jamovi project, 2025, https://www.jamovi.org ). Graphs were plotted using GraphPad Prism 8.0 (GraphPad Software Inc., USA). Data were expressed as the mean ± standard deviation (SD) or median with the interquartile range (Q1, Q3). The median serum ferritin levels were determined for the PCOS and non-PCOS groups. The statistical significance of categorical variables was assessed using the chi-square test. Correlations between two variables were evaluated using the Spearman's rank correlation coefficient (r s ). For continuous variables, statistical significance was assessed using the Mann–Whitney U test and independent samples t -test. The threshold for statistically significant difference was set at p < 0.05. AFC, Antral follicle count; AMH, Anti-müllerian hormone; ART, Assisted reproductive technology; BMI, Body mass index; E2, Estradiol; FORT, Follicular output rate; GnRH, Gonadotropin-releasing hormone; hCG, Human chorionic gonadotropin; LH, Luteinizing hormone; PCOS, Polycystic ovarian syndrome; r s , Spearman's rank correlation coefficient; SD, Standard deviation.

In the present study, serum ferritin levels were not elevated in women with PCOS and had no significant associations with embryological outcomes in either PCOS women or non-PCOS women undergoing ART treatments, suggesting that serum ferritin has limited utility as a predictive marker for embryological success. Nevertheless, iron status evaluation remains important due to its potential role in reproductive physiology. Future prospective studies are warranted with larger cohorts to evaluate both serum and follicular fluid iron, the total iron binding capacity, and inflammatory markers to better understand the relationship between iron metabolism in PCOS and reproductive outcomes.

According to the present study, serum ferritin levels were not significantly altered in PCOS patients and they had no significant effects on embryological outcomes in both PCOS and non-PCOS women. It is well established that women with PCOS have elevated AMH levels, increased LH/FSH ratios, and disrupted pulsatile gonadotropin release, contributing to menstrual irregularities. 4 , 7 , 29 Similar observations were obtained in the current study. Serum ferritin is a key indicator of systemic iron status and routinely used in clinical practice to reflect the total body iron stores. 33 The hormonal abnormalities observed in PCOS may alter iron metabolism. Yang et al. showed that E2 suppresses hepcidin, a key regulator of systemic iron homeostasis, thereby promoting cellular iron overload, which may lead to increased ferritin levels. 34 Studies of PCOS and transfusion-dependent β-thalassemia showed that serum AMH levels were negatively correlated with ferritin, suggesting that iron overload adversely affects ovarian reserve. 35 , 36 LH does not directly regulate iron metabolism but it contributes to hyperandrogenism and hyperprolactinemia in PCOS, and both are associated with elevated ferritin levels. 37 However, only very weak associations were observed between serum ferritin levels and serum reproductive hormones in the present study. Elevated ferritin has been implicated in several metabolic and inflammatory conditions, including metabolic syndrome, cardiovascular disease, chronic renal dysfunction, and autoimmune disease, as well as reproductive abnormalities such as unexplained infertility and recurrent pregnancy loss. 33 , 38 , 39 Previous studies reported elevated ferritin levels in PCOS women, particularly in obese or insulin-resistant patients. 21 , 23 , 24 , 25 , 26 Furthermore, Alam et al. observed high ferritin levels in women with higher BMI. 40 By contrast, our study failed to find a significant association between serum ferritin and BMI. The differences between our findings and those obtained in previous studies may be attributed to variations in lifestyle and diet between Indian and Western populations, as well as any medications taken by the subjects involved. Metformin therapy, 41 , 42 and chronic use of antacids and proton pump inhibitors have been reported to reduce serum ferritin levels. 43 Similarly, dietary supplements may contribute to altered serum ferritin levels. The World Health Organization and several studies recommend iron and folic acid supplementation during the pre-conception period to support maternal health and fetal development, which may increase the iron status in women. 44 , 45 , 46 However, calcium supplements can reduce divalent metal transporter 1-mediated iron absorption to potentially reduce iron levels and serum ferritin concentrations. 47 , 48 Diets rich in oxalates 49 and dark-green leafy vegetables are also known to reduce serum ferritin levels. 50 It is known that iron status can influence the embryo quality, 20 , 51 highlighting the importance of optimizing the iron status in women prior to ovarian stimulation. A major strength of the present study was the evaluation of serum ferritin levels and their correlations with embryological outcomes in both PCOS and non-PCOS women. However, serum ferritin had no associations with embryological parameters in either PCOS or non-PCOS women in the present study, suggesting its utility may be limited as a marker for predicting ART outcomes.

The present study had some limitations that should be acknowledged. The relatively small sample size may have reduced the statistical power of the findings. Information regarding important confounding factors, including metformin or insulin therapy in women with PCOS, dietary intake, lifestyle factors, and nutritional supplementation, which may have influenced serum ferritin levels and affected outcomes in both the PCOS and non-PCOS groups, was not recorded prior to sample collection. In addition, male factor infertility was not excluded from the study population, which may have confounded the observed embryological outcomes.

Polycystic ovarian syndrome (PCOS) is one of the most prevalent endocrine–metabolic disorders that affect women of reproductive age, where it is characterized by hyperandrogenism, irregular menstrual cycles, and cystic ovarian morphology. 1 Globally, PCOS affects approximately 6–13 % of women in the reproductive age group and up to 70 % of cases remaining undiagnosed. 2 In India, the prevalence of PCOS ranges from 3.7 % to 22.5 %. 3 Women with PCOS exhibit characteristic endocrine alterations, including elevated serum testosterone, defective gonadotropin-releasing hormone (GnRH) pulsatility, increased serum luteinizing hormone (LH) levels, decreased follicle stimulating hormone (FSH) levels, and an elevated LH to FSH ratio, which contribute to anovulation, reduced fertility, and a higher risk of miscarriage. 4 Furthermore, PCOS is closely linked to various metabolic abnormalities, including insulin resistance, type 2 diabetes mellitus, hypertension, and cardiovascular disease. 5 Chronic low-grade inflammation has been implicated in the pathophysiology of PCOS, contributing to anovulation, long-term metabolic complications, and infertility. 6 In addition, obesity is frequently observed in women with PCOS. 7 The prevalence of PCOS has been shown to increase by approximately 0.4 % for every 1 % rise in obesity, thereby significantly exacerbating the severity of the disease and complicating management strategies. 8 The metabolic patterns and associated physiological changes in PCOS are not fully understood, but recent studies indicate significant disruption of micronutrient homeostasis, 9 , 10 , 11 , 12 highlighting the role of micronutrients in the pathogenesis of PCOS. Iron is an essential micronutrient and a critical component of heme proteins, iron–sulfur clusters, and iron-centered proteins involved in functions such as oxygen transport and oxidative phosphorylation. Studies have demonstrated that ovarian iron overload compromises ovarian function by disrupting steroidogenesis and interferes with the ovarian microenvironment, 13 , 14 , 15 showing the importance of iron metabolism and its implications for female reproductive health. Iron homeostasis is tightly regulated, and deficiency can lead to anemia and pregnancy-related complications, whereas overload is associated with oxidative stress, organ dysfunction, 16 and an increased risk of cancer. 17 Furthermore, iron overload has been associated with female reproductive disorders such as endometriosis and ovarian aging, 18 , 19 mainly due to granulosa cell ferroptosis and lipid peroxidation. 18 , 20 Studies have indicated the altered expression of ferroptosis-related genes in serum 21 and proteins in granulosa cells of PCOS patients, 22 characterized by elevated nuclear receptor co-activator 4, and downregulated ferritin heavy chain 1 and glutathione peroxidase 4, suggesting the activation of ferroptosis. In rodent models, inhibition of ferroptosis has been shown to overcome anovulation, polycystic ovarian morphology, and poor oocyte quality. 22 Elevated levels of serum ferritin, an iron storage protein, have been observed in women with PCOS, and associated with hyperandrogenism, insulin resistance, and obesity, indicating systemic iron overload in PCOS. 23 , 24 , 25 , 26 Moreover, the level of hepcidin, a key regulator of iron homeostasis, was found to be decreased in PCOS patients, suggesting disrupted iron regulation. This imbalance was correlated with increased insulin resistance and hyperandrogenism, which are characteristic features of the PCOS condition. 27 , 28 Evidence remains inconsistent regarding serum ferritin levels across different PCOS phenotypes. Elevated ferritin levels have been reported in PCOS women with oligomenorrhea or amenorrhea, whereas those with hyperandrogenism have lower median ferritin concentrations compared with their normoandrogenic counterparts. 26 Further investigation of serum ferritin levels in women with PCOS is clinically relevant because PCOS is associated with hyperandrogenism, insulin resistance, type 2 diabetes mellitus, and obesity, and there are discrepancies in existing data regarding serum ferritin levels across different PCOS phenotypes. Furthermore, evidence from animal studies indicates that ovarian iron accumulation can impair ovarian function and reduce oocyte quality, suggesting a potential link between iron metabolism and reproductive outcomes. 20 These observations highlight the need to clarify whether serum ferritin levels are altered in women with PCOS and whether any alterations are associated with embryological outcomes. Considering these research gaps, the present study aimed to estimate and compare the serum ferritin level in PCOS and non-PCOS women undergoing assisted reproductive technology (ART). Furthermore, this study investigated whether serum ferritin levels are associated with the antral follicle count (AFC), ovarian response, oocyte maturity, fertilization potential, and preimplantation embryo development. In addition, this study explored the potential of using serum ferritin as a biomarker for predicting embryological outcomes in PCOS and non-PCOS women undergoing ART.

On behalf of all authors, the corresponding authors state that there are no conflicts of interest to declare.