Ovario-protective effect of intra-ovarian injection of platelet rich plasma (PRP) after laparoscopic cystectomy of ovarian endometrioma: a randomized control trial

We designed an open label phase II parallel randomized clinical trial to evaluate the effect of intra-ovarian injection of PRP on post-operative ovarian reserve of patients with ovarian endometrioma. This single-center study was conducted at Hazrat Rasoul Hospital, a university-affiliated tertiary referral center in Tehran, Iran. The study was conducted by the Helsinki declaration and was approved by the Iran University of Medical Sciences’ Committee for Ethics in Medical Research by the code: IR.IUMS.REC.1400.521. The study was prospectively registered in the Iranian Registry of Clinical Trials under the number IRCT20191123045476N3 on December 1, 2021 ( https://irct.behdasht.gov.ir/trial/58688 ). The study reporting adheres to Consolidated Standards of Reporting Trials (CONSORT) guidelines. Women aged 20 to 35 years with unilateral or bilateral endometriomas measuring 4–10 cm, experiencing pelvic pain and unresponsive to six months of medical treatment, who were scheduled for laparoscopic cystectomy, were included in the study. All patients underwent transvaginal ultrasound examination for ovarian cysts assessment according to the Ovarian-Adnexal Reporting and Data System (O-RADS) classification and evaluation antral follicular count (AFC) before surgery.AMH levels were measured using the ELISA (Enzyme-Linked Immunosorbent Assay) method for assessing ovarian reserve before intervention. Exclusion criteria included Anti-Müllerian Hormone (AMH) levels > 1.1 ng/mL, use of hormone therapy within the past three months, a Body Mass Index (BMI) > 30 or < 18, autoimmune diseases, malignancies, sexually transmitted infections, endocrine disorders, tubal factor infertility, and recent use of anticoagulants. Participants were randomly assigned to two study groups using simple randomization generated through the SealedEnvelope.com randomization tool. The allocation sequence was concealed within sealed opaque envelopes, which were opened one day before the surgery. It’s important to note that the study was conducted in an open-label fashion. This study compared two groups: a post-surgical Platelet-Rich Plasma (PRP) group and a control group that received no intraoperative product. The PRP group was administered 2–4 cc of PRP immediately following laparoscopic cystectomy. All cystectomies were performed using the stripping technique by a single expert surgeon at an advanced laparoscopic clinic in a university-affiliated hospital (Hazrat-e-Rasoul Hospital, Tehran, Iran). The procedure commenced with salpingo-oophorectomy, followed by aspiration of cyst contents. The cyst wall was then carefully separated from the ovarian cortex using traction and counter-traction techniques. Hemostasis was achieved by applying bipolar coagulation at 40 watts and, when necessary, sutures. The use of bipolar electrocautery was minimized to reduce thermal damage. Ovarian suturing was performed with 3 − 0 Vicryl, using the fewest possible sutures to approximate the ovarian cortex, particularly in cases involving large cysts or inadequate hemostasis following cystectomy. Preceding the surgery, 20 cc of the patient’s blood was gathered into a tube utilizing the PRP kit (ROOYAGEN). The tube underwent vigorous shaking at a speed of 3600 rounds per minute (RPM) for a duration of 6 min. Following centrifugation, the PRP supernatant was extracted using a No. 16 needle, yielding approximately 2–4 cc of PRP. This PRP was injected subcortically to the ovary which underwent cystectomy by spinal needle 20 gaug. The main measure of effectiveness was the alteration in Anti-Mullerian Hormone (AMH) observed six months after the intervention. All samples were processed and analyzed in a single central laboratory using the same ELISA kit batch to ensure consistency across measurements and laboratory personnel were blinded to group allocation. The antral follicular count (AFC) six months post-surgery served as the secondary outcome. Safety outcomes encompassed the incidence of post-interventional complications, including allergic reactions, fever, pain, bleeding or hematoma, and post-surgical abscess at the injection site within the initial 48 h and six months post-intervention. The sample size was calculated based on the data of Sadullayev et al. study [ 8 ], using Pass 2021 software. Assuming a post intervention mean serum AMH level of 0.95 with a standard deviation of 0.35 for PRP group and a mean of 0.65 with a standard deviation of 0.25 for the other group, along with type I and type II error rates of 0.05 and 0.2 respectively, and a 10% dropout rate, a total sample size of 38 was calculated. The analysis of data was conducted using SPSS version 23 software. Quantitative variables were compared between the two study groups using the Student t-test and the U Mann-Whitney test (for non-parametric variables). Before the comparison, the normal distribution of variables was assessed using the Kolmogorov-Smirnov test. The comparison of categorical variables involved the use of the Chi-square and Fisher exact tests. To assess the impact of the intervention on ovarian reserve, we initially compared the serum Anti-Müllerian Hormone (AMH) level and Antral Follicular Count (AFC) six months post-intervention. Subsequently, to adjust for baseline variations, we compared the mean differences before and after the intervention in each patient. Additionally, we employed the ANCOVA test to compare post-intervention outcomes between the two study groups, adjusting for the baseline quantities. The modified intention-to-treat approach was used for the analysis of the outcome.

From March to November 2022, a total of 142 patients underwent evaluation to determine their eligibility for the study. Among them, 38 patients were ultimately enrolled in two study groups: Platelet-Rich Plasma (PRP) group ( n  = 20) and a group with no intervention ( n  = 18) (Fig.  1 ). One case in the PRP group was pregnant with natural conception in the 4th month of follow-up and one case in the control group was declined in the 6th month post interventional assessment, and the analysis of the outcomes was performed on 36 patients. The mean (SD) age of the patients was 36.72 (6.02), with approximately 64% having a prior diagnosis of infertility. Fig. 1 Participant’s flow diagram Participant’s flow diagram The baseline characteristics of the patients revealed no significant imbalances between the two study groups, suggesting that the randomization process was effective (see Table  1 ). Ovarian endometrioma was observed in nearly all patients in both study groups, with 56% and 65% of the control and intervention groups having bilateral endometriomas. The percentages for adenomyosis were (50% vs. 40%), for cul-de-sac involvement (72.2% vs. 70%), and for rectal endometriosis (22.2% vs. 35.5%) (See Table  2 ). Table 1 Baseline characteristics of patients in two study groups Control ( n  = 18) PRP ( n  = 20) P value Age a 35.75(4.43)26–41 36.5(4.51)27–41 0.62 Gravid b 0(1.75)0–3 0(2)0–4 0.86 Live birth b 0(1)0–2 0(1)0–2 0.64 Abortion b 0(0.75)0–2 0(1)0–3 0.67 Dysmenorrhea b 7.19(2.14) 4–10 7.05(2.5) 3–10 0.96 AFC b 3(2)2–5 3(1)1–7 0.46 AMH b 0.75(0.18) 0.5–1.1 0.78(0.16) 0.5–1.1 0.62 Married c 15(83.3%) 18(90%) 0.54 History of infertility c 9(50%) 14(70%) 0.21 a Data are presented as Mean (SD) Range and were compared using student t-test b Data are presented as Median (IQR) Range and were compared using U Mann Whitney test c Data are presented as frequency (percentage) and were compared using chi square test Baseline characteristics of patients in two study groups a Data are presented as Mean (SD) Range and were compared using student t-test b Data are presented as Median (IQR) Range and were compared using U Mann Whitney test c Data are presented as frequency (percentage) and were compared using chi square test Table 2 Distribution of pelvic endometriosis lesions and symptoms in patients of two study groups Control ( n  = 18) PRP ( n  = 20) P value Ovary Unilateral 7(38.9%) 6(30%) 0.564 Bilateral 11(61.1%) 14(70%) Endometrioma Size (mm) Right 45.37(26.4) 42.5(18.99) 0.80 Left 32.37(20.97) 38.75(26.33) 0.48 Hydrosalpynx a 8(44.4%) 4(20) 0.164 Rectum involvement a 4(22.2%) 7(35%) 0.386 Cul de sac involvement a 13(72.2%) 14(70%) 0.88 Adenomyosis a 9(50%) 8(40%) 0.536 Myoma a 7(38.9%) 7(35%) 0.80 Dysmenorrhea VAS score b 6.45(2.66) 5.67(2.77) 0.5 Dyspareunia VAS score 7.19(2.14) 7.02(2.5) 0.96 a Data are presented as frequency (percentage) and were compared using chi square test b Data are presented as Mean (SD) Range and were compared using student t-test Distribution of pelvic endometriosis lesions and symptoms in patients of two study groups a Data are presented as frequency (percentage) and were compared using chi square test b Data are presented as Mean (SD) Range and were compared using student t-test We actively monitored all patients for various solicited adverse events, including allergic and autoimmune reactions, fever, pain, bleeding or hematoma, and post-surgical abscess at the injection site within the initial 48 h and six months post-intervention. No adverse events were reported during the first 48 h after the injection. In addition, no intervention-related serious adverse event was reported in all patients during the 6 months of follow-up. Following cystectomy, the control group exhibited a considerable decrease in serum Anti-Müllerian Hormone (AMH) levels, with a mean difference of -0.24 and a 95% confidence interval of (-0.5- 0.12). In contrast, the intervention group showed a negligible change in AMH levels, with a mean difference of 0.05 and 95% CI of (-0.17–0.26). Similar results were observed in the results of ANCOVA analysis after adjustment for baseline serum AMH level. Adjusted mean difference (95% CI) was − 0.308(-0.626-0.01) between two study groups P  = 0.057. (See Table  3 ) Table 3 Comparison of baseline and post intervention AMH and AFC and their changes during the study period in two groups Control ( n  = 16) PRP ( n  = 19) P value AMH Baseline a 0.78(0.16) 0.75(0.18) 0.62 post intervention a 0.50(0.39( 0.83(0.35) 0.045 Difference a -0.24(0.38) 0.05(0.35) 0.068 AFC Baseline b 3(1–7) 3(2–5) 0.46 post intervention b 2(0–10) 2(0–6) 0.49 Difference b 0(-5-7) -1(-4-3) 0.42 a Data are presented as Mean (SD) Range and were compared using student t-test b Data are presented as Median (SD) Range and were compared using U Mann Whitney test Comparison of baseline and post intervention AMH and AFC and their changes during the study period in two groups a Data are presented as Mean (SD) Range and were compared using student t-test b Data are presented as Median (SD) Range and were compared using U Mann Whitney test While the antral follicular count (AFC) seemed to decrease in the control group (mean and 95% CI of 0.9, -0.54 -2.36), this decline was not statistically significant. Additionally, the change in AFC for the intervention group was negligible. The results of ANCOVA analysis showed Adjusted mean differences (95% CI) for AFC were − 0.733(-2.76-1.3) between two study groups, P  = 0.462.

Our findings suggest that intra-ovarian PRP injection may help maintain AMH levels during the first six months after cystectomy. However, as the observed difference did not reach statistical significance, further studies with larger sample sizes are necessary to establish the efficacy of this intervention.

The recent study introduces a novel regenerative approach for protecting ovarian function against tissue damage and inflammation following surgical procedures in women with endometriosis. By administering intra-ovarian prophylactic platelet-rich plasma (PRP) after surgery, we observed that patients’ anti-Müllerian hormone (AMH) levels remained consistent relative to preoperative values. However, this intervention did not affect the antral follicle count (AFC) changes experienced by these patients. The evaluation of the safety of intra-ovarian PRP injections reported no significant adverse effects. Due to the well-documented reduction in serum anti-Müllerian hormone (AMH) levels following endometrioma surgery, increasing attention has been directed toward medical approaches aimed at preserving ovarian reserve in women with endometriotic cysts [ 8 ]. Despite these concerns, surgery remains the last resort in selected patients—particularly those experiencing severe pain, poor response or contraindications to medical therapy, or when malignancy is suspected [ 9 ]. While laparoscopic cystectomy can effectively reduce symptoms and recurrence, it is well established that the procedure carries a risk of diminishing ovarian reserve, especially in cases of bilateral or large (> 4 cm) endometriomas, as reflected in postoperative declines in AMH [ 10 , 11 ]. According to the ESHRE guidelines, individualized management should be prioritized for women wishing to preserve fertility. Conservative medical treatment (e.g., progestins, GnRH analogues, or hormonal contraceptives) is typically preferred as first-line therapy, with surgery reserved for patients who fail medical therapy or present with infertility, persistent symptoms, or suspected malignancy. In addition, preoperative fertility preservation counseling—such as oocyte or embryo cryopreservation—is recommended for women under 35 years of age with low ovarian reserve or bilateral endometriomas [ 2 ]. Our study explores PRP injection as a potential method to mitigate surgery-induced ovarian damage, but clinicians must first carefully assess whether cystectomy itself is warranted. During recent decades PRP has frequently been used in gynecologic disorders, especially in that need tissue or cell regeneration [ 5 ]. The PRP was applied in the treatment of endometrial proliferation defects e.g. thin endometrium [ 6 , 12 , 13 ] and Asherman syndrome [ 14 ]. Additionally, intra-ovarian injection of PRP was frequently applied to recover ovarian reserve in patients with poor ovarian reserves, ovarian insufficiency and poor ovarian responders to ovarian hyper stimulation [ 7 , 15 , 16 ]. Secondary studies on the effect of intra-ovarian injection of PRP looks controversial. Although meta-analyses concluded the positive effect of PRP on pregnancy of patients with poor ovarian reserve [ 17 , 18 ], some studies proposed concerns in order to methods of PRP products preparation which might lead to different clinical results [ 5 , 19 ]. Experiences regarding the application of PRP as an ovario-protective product after ovarian cystectomy in endometriosis patients are limited. A recent study compared the effects of PRP combined with sclerotherapy to PRP alone and a control group following laparoscopic cystectomy in patients with endometriosis. The study found that both intervention groups preserved ovarian reserve, with better outcomes observed in the combination therapy group. Levels of serum Anti-Mullerian Hormone (AMH) and ovarian antral follicle count improved in both intervention groups. While their findings aligned with our study concerning AMH levels, we did not observe any differences in the decline of post-operative Antral Follicle Count (AFC) between the two study groups [ 8 ]. Although the ANCOVA analysis demonstrated a between-group adjusted mean difference in AMH levels favoring the PRP group, the result did not reach conventional statistical significance ( P  = 0.057). These findings suggest a potential protective effect of PRP on ovarian reserve, but due to the borderline significance and small sample size, the evidence should be interpreted cautiously. Larger studies are warranted to confirm these preliminary observations. Both groups experienced minimal declines in AFC, resulting in no notable differences between them. Conversely, the reversed response to PRP observed in AMH and AFC outcomes in the current study had been noted earlier by Sadat et al., who employed PRP to treat women with poor ovarian reserves [ 15 ], although it’s crucial to consider distinct mechanisms underlying reduced ovarian reserve in endometriosis patients undergoing cystectomy. The decline in ovarian reserve in patients who have undergone cystectomy for endometrioma appears to result from two separate mechanisms. Firstly, post-surgical ovarian reserve decline may stem from the removal of ovarian cortex tissue during laparoscopic cystectomy [ 20 ]; thus, PRP administration could potentially stimulate remaining germ cells or germ stem cells to compensate for ovarian reserve loss and maintain AMH levels steady [ 19 ]. Another established mechanism of reducing ovarian reserve after laparoscopic cystectomy is post-surgical inflammation. Previous studies have indicated that inflammation, oxidative stress, fibrosis, and DNA damage contribute to diminished ovarian reserve in both operated and unoperated ovarian endometriomas [ 1 , 21 ]. Research suggests that PRP possesses anti-inflammatory properties within ovaries and alleviates follicular atresia via antiangiogenic factors [ 22 ]. In addition, PRP promotes tissue healing without eliciting inflammatory reactions utilizing anti-inflammatory molecules [ 15 ]. No short-term safety concerns were observed with intra-ovarian autologous PRP injection, and the intervention was generally well-tolerated without adverse effects in patients, aligning with previous reports [ 15 , 23 ]. However, a long-term safety assessment of the intervention was not conducted, highlighting the necessity for large-sample clinical trials with extended follow-up periods. It is essential to prioritize safety considerations when exploring the use of autologous PRP injections to ensure patient well-being and minimize any unforeseen complications. Given the limited number of participants and the lack of prior confirmatory trials in this context, this study should be considered exploratory. Its aim was to generate preliminary evidence and inform future, more definitive research.

Pelvic endometriosis predominantly impacts the ovaries, leading to a condition known as ovarian endometrioma (OMA). OMA commonly imposes damage on the cortical ovarian tissue, leading to a decline in ovarian reserve and serum AMH level as its predictor [ 1 ]. In cases where severe pelvic pain and infertility are prominent symptoms, a decisive surgical intervention like ovarian cystectomy is often recommended to minimize the risk of OMA recurrence [ 2 ]. Unfortunately, these surgical procedures can reduce the ovarian reserve too [ 3 , 4 ]. Consequently, the pursuit of scientific interventions that mitigate the adverse effects of OMA and its invasive treatments is crucial. Platelet-rich plasma (PRP) has become a widely utilized biological product in regenerative medicine over the past few decades. In the field of gynecology, PRP therapy is commonly employed to address the urgent requirement for the recovery of damaged or insufficient tissues [ 5 ]. A meta-analysis has reported the positive impact of PRP on the endometrial receptivity of women with a thin endometrium following assisted reproduction techniques (ART) [ 6 ]. Research is currently underway to explore the therapeutic potential of intra-ovarian injection of Platelet Rich Plasma (PRP) for ovarian insufficiency or poor ovarian response, although its confirmed benefits are yet to be established [ 7 ]. Given the potential promise of PRP as a regenerative technique in gynecological treatment, we have designed a clinical trial aimed at assessing the impact of intra-ovarian PRP injection on the post-operative reduction of ovarian reserve following laparoscopic cystectomy of ovarian endometrioma. It’s worth noting that due to the absence of a realistic animal model for ovarian endometrioma, there is currently no preclinical support for the ongoing clinical trial.