Intrauterine instillation of autologous platelet-rich plasma in infertile females with thin endometrium undergoing intrauterine insemination: an open-label randomized controlled trial

Abstract

BACKGROUND

Thin endometrium in infertile female patients has always been a clinical challenge for the treating reproductive physicians.

OBJECTIVE

This study aimed to evaluate the effect of intrauterine instillation of autologous platelet-rich plasma on endometrial thickness and vascularity in infertile female patients with thin endometrium undergoing ovarian stimulation with intrauterine insemination.

STUDY DESIGN

This prospective randomized control study included 120 women undergoing ovarian stimulation with intrauterine insemination, aged between 21 and 37 years, with persistent thin endometrium (<7 mm) on ≥1 cycle in previous ovarian stimulation cycles, even after conventional treatment with estradiol valerate. The women were randomly assigned to study group A and control group B. Baseline endometrial thickness and endometrial vascularity were noted. Intrauterine instillation of autologous platelet-rich plasma was done on the day of trigger in group A, whereas estradiol valerate was given in group B. Another evaluation of endometrial parameters was done on the day of intrauterine insemination. The cycle was repeated for 3 cycles or until the pregnancy was achieved, whichever occurred earlier. Parameters were noted for both groups. Primary outcomes were the change in endometrial thickness and endometrial vascularity. Secondary outcomes were positive pregnancy rate and clinical pregnancy rate.

RESULTS

In group A, mean pre–platelet-rich plasma endometrial thickness was 4.66±0.79 mm, which significantly increased to 7.47±0.85 mm after platelet-rich plasma instillation (P<.05) after 3 cycles. This increase was significantly greater than that observed in group B. There was significant increase in endometrial vascularity in group A compared with group B (P<.05). The positive pregnancy rate and clinical pregnancy rate in group A were 23.73% and 18.64%, respectively, and significantly higher than those in group B.

CONCLUSION

Autologous platelet-rich plasma is a promising, easily procurable, and novel option for management of infertile females with thin endometrium not responding to conventional treatment. Using it in intrauterine insemination cycles can reduce costs and psychological burden of this subgroup of women by reducing the need for resorting to advanced options such as in vitro fertilization and surrogacy.

AJOG Global Reports at a Glance.

Why was this study conducted?

Persistent thin endometrium lining unresponsive to standard therapies in assisted reproductive technology programs poses a significant challenge, leading to repeated cycle cancelation and psychological and financial strain. There is no robust evidence about the use of any adjuvant for this clinical entity. This study explores the usefulness of autologous platelet-rich plasma (APRP) in cycles with ovarian stimulation followed by intrauterine insemination in women with thin endometrium. The existing literature supports its use in frozen embryo transfer cycles.

Key findings

APRP is a promising, easily procurable, novel, and economic option for treatment of infertile patients with thin endometrium. Its use in ovarian stimulation cycles can reduce costs and psychological burden in this subgroup by avoiding the need for resorting to advanced options such as in vitro fertilization (IVF) and surrogacy.

What does this add to what is known?

Although APRP has been used in IVF and frozen embryo transfer cycles, there is no literature demonstrating the efficacy of intrauterine instillation of APRP in infertile women with thin endometrium during ovarian stimulation cycles. Thus, we have explored the potential of APRP as a low-cost promising option in this subgroup of women.

Introduction

The endometrium undergoes dynamic physiological and morphologic changes during the menstrual cycle to prepare for successful implantation. Appropriate endometrial thickness (ET) and vascularity (EV) play an essential role as determinants of successful pregnancy in infertility programs.^1,2 Persistent thin endometrium lining, particularly when resistant to standard therapies, poses a significant clinical challenge.

Thin or suboptimal ET is ET below the threshold required for embryo implantation. It has been associated with poor pregnancy rates, spontaneous abortion, ectopic pregnancy, and abnormal placentation.^3,4 Quantitatively, 7 mm is the often-stated cutoff for thin or suboptimal endometrium, which is associated with poor outcomes in artificial reproductive technology cycles.^1,4,5

Ovarian stimulation with intrauterine insemination (OS-IUI) is an easy, cost-effective, and noninvasive first-line management option for couples diagnosed with unexplained or mild male subfertility. Although the literature does not support intrauterine insemination (IUI) cycle cancelation for thin endometrium, appropriate ET and endometrial blood flow are determinants of success in IUI cycles.^1,6, 7, 8

Whereas the incidence of thin endometrium is between 1% and 2.5% in in vitro fertilization (IVF) cycles, it reaches 38% to 66% in ovarian stimulation (OS) cycles.¹ Thin ET often leads to repeated cycle cancelations (in both IUI and IVF), thereby causing psychological and financial strain, and the woman is ultimately left with no other option but surrogacy.

There is insufficient evidence to recommend adjuvants such as luteal estradiol, low-dose aspirin, sildenafil citrate, pentoxifylline, vitamin E, human chorionic gonadotropin (hCG), and intrauterine granulocyte colony-stimulating factor (G-CSF) instillation for restoring optimal ET in women with thin endometrium.¹ Platelet-rich plasma (PRP) is also a novel treatment option. PRP is blood plasma prepared from fresh whole blood, which is enriched with platelets. It contains several growth factors, such as VEGF, FGF, EGF, PDGF, TGF, and other cytokines that stimulate proliferation and growth.⁹ These secreted proteins have paracrine effects on different cells, promoting cellular migration, proliferation, angiogenesis, and subsequently tissue regeneration. It is used in IVF and frozen embryo transfer (FET) cycles, although the evidence supporting such practice is still not robust.¹ The autologous PRP (APRP) instilled in the endometrial cavity results in endometrial regeneration, thus allowing successful implantation of fertilized ovum.

Although PRP has been used widely in IVF and FET cycles, there is no literature demonstrating the efficacy of intrauterine instillation of APRP in infertile women with thin endometrium during OS followed by IUI. Thus, this study aimed to assess the effect of intrauterine instillation of APRP on the endometrium in women with thin endometrium.

Materials and Methods

This open-label randomized controlled study was conducted over a period of 6 months after approval from the Institutional Ethics Committee on 120 infertile women planned for OS-IUI-cycle at IVF-Centre, Department of Obstetrics and Gynaecology, Safdarjung Hospital, Vardhman Mahavir Medical College, New Delhi. The primary objective was to study the effect of intrauterine APRP instillation on ET and EV in infertile women with thin endometrium undergoing OS-IUI. The secondary objective was to assess the positive and clinical pregnancy rates.

Sample size calculation

Chang et al¹⁰ observed that ET in the PRP group was 7.65±0.22 mm, whereas in the control group it was 6.52±0.31 mm. The clinical pregnancy rates were 44.12% and 20% in the PRP and control groups, respectively. Taking these values as the reference, the minimum required sample size with 80% power of study and 5% level of significance was 55 patients in each group. Assuming 10% error, 60 women were enrolled in each group. Thus, the total sample size was 120 (60 per group).

The formula used was: n≥((pc × (1-pc)+pe × (1-pe)) × (Zα+Zβ)²)/(pc-pe)², with “pc” being the clinical pregnancy rate in the PRP group, “pe” the clinical pregnancy rate in the control group, Zα the value of Z at 2-sided alpha error of 5%, and Zβ the value of Z at power of 80%. The formula for comparing the means of the 2 groups was:

$$\begin{array}{c}N\ge 2{\left(standarddeviation\right)}^2\times {\left(Z_{\alpha }+Z_{\beta }\right)}^2\hfill \\ {\left(meandifference\right)}^2\hfill \end{array}$$

Zα is the value of Z at 2-sided alpha error of 5%, Zβ is the value of Z at power of 80%, and mean difference is the difference in mean values between the 2 groups.

After providing informed consent, those meeting the eligibility criteria were randomized and assigned to study group A and control group B. The inclusion criteria were infertile female patients aged between 21 and 37 years with persistent thin ET <7 mm on ≥1 cycle in previous OS cycles (stimulated with letrozole [2.5 mg × 5 days] followed by sequential human menopausal gonadotropin [HMG]), hemoglobin ≥11 g/dL, and platelets ≥150,000/μL. The exclusion criteria included history suggestive of platelet dysfunction, such as easy bruisability, frequent nosebleeds, heavy menstrual bleeding, bleeding gums, or excessive bleeding during dental procedures; blood-borne diseases such as hepatitis B, hepatitis C, syphilis, and HIV; nonsteroidal antiinflammatory drug intake in 10 days before the procedure; being on anticoagulant therapy; severe male factor infertility; tubal factor infertility; World Health Organization (WHO) group 1 to 3 ovulatory dysfunction; Müllerian anomalies; Asherman syndrome; moderate or severe endometriosis; active vaginal or cervical infection; pelvic inflammatory disease; or any significant comorbidity or psychiatric disorder that interferes with providing consent, study participation, follow-up, or study interpretation.

Statistical analysis

Categorical variables were presented as numbers and percentages. The quantitative data with normal distribution were presented as mean±standard deviation. The comparison of quantitative variables was done using the independent t test. Repeated-measures analysis of variance (ANOVA) was used for comparison of ET across follow-up. The comparison of qualitative data was analyzed using the chi-square test. If any cell had an expected value of <5, the Fisher exact test was used. Data entry was done in a Microsoft Excel spreadsheet. IBM SPSS Statistics, Version 25.0 (IBM Corp., Armonk, NY) was used for final analysis. P value <.05 was considered statistically significant.

The couple underwent basic infertility workup. After establishing tubal patency and ruling out other factors such as advanced female age >37 years, severe male factor infertility, tubal factor infertility, moderate to severe endometriosis, poor ovarian reserve, and correction of hormonal factors such as hypothyroidism and hyperprolactinemia, patients were planned for IUI. Baseline hysteroscopy was done to rule out Asherman syndrome or any other intrauterine pathology. Enrollment into the study group was done by computer-generated random sequence until a sample size of 120 was reached.

After baseline scanning on days 2 or 3 (to rule out ovarian cysts and evaluate antral follicle count and baseline ET), OS was performed with an ovulogen (2.5–5-mg letrozole × 5 days from day 2 to 6 with sequential gonadotropins [HMG]). Further cycle monitoring was done from day 7 onward. Those having thin endometrium (<7 mm) even with follicle size ≥16 mm and suboptimal EV, defined as <5 vascular signals reaching the central zone (zone 3 and 4 per Applebaum grading of the endometrium measured by power Doppler by the same observer), were prescribed estradiol valerate 6 mg/d during the cycle.¹¹

Thin endometrium was defined as ET <7 mm with follicle size <16 mm on ultrasound monitoring, not responding to estradiol valerate in ≥1 cycle.

Those enrolled in the study group were given ovulation induction using letrozole (2.5–5 mg once daily from day 3–7) followed by sequential gonadotropin treatment (HMG injection, 75 IU on alternate days from day 8 onward), and received estradiol valerate 6 mg/d from day 8 onward. Serial transvaginal ultrasound (TVS) examinations were performed using a transvaginal probe of 5 to 9 MHz of the ultrasound machine DC-N3 (Mindray, Shenzhen, China) (starting from day 8 and repeated as required).After noting the baseline ET and EV on the day of ovulation trigger, intrauterine instillation of 1 mL of APRP preparation was done on the same day; 36 to 40 hours later, IUI was done (for successive 3 cycles or until conception, whichever was earlier). Post-APRP-instillation TVS with power Doppler was done by the same observer after 36 to 40 hours (day of IUI) to assess ET and EV. Luteal support using vaginal micronized progesterone was given to all women undergoing IUI. Urine pregnancy test was done after 2 weeks. Those with negative test results underwent similar second and third cycles. Change in ET and EV was assessed again in subsequent cycles on the trigger day and 36 to 40 hours later.

“Positive pregnancy” was defined as a positive urine pregnancy test (urinary beta-hCG test), and “clinical pregnancy” as demonstration of cardiac activity on ultrasound. The ET and EV were assessed in the subsequent cycles before and after PRP instillation.

The study's endpoint was achieving positive pregnancy or completion of 3 cycles of intrauterine instillation of APRP (whichever occurred earlier).

For ET assessment, the TVS was done with empty bladder. ET was measured in the median longitudinal plane of the uterus, and defined as the maximum distance from one basal endometrial interface across the endometrial canal to the opposite endometrial-myometrial interface. For EV measurement, vascular signals were identified using power Doppler with machine presets for the endometrium. Accordingly, zones were categorized as: Zone 1—when blood flow reaches the hypoechoic endometrial-myometrial interface; Zone 2—when blood flow reaches the outer hyperechoic area of the endometrium; Zone 3—when blood flow reaches the intervening hypoechoic area; and Zone 4—when blood flow reaches the central echogenic line.¹¹ According to the blood supply distribution, the vascularity was categorized as: (1) excellent: at least 5 signals in Zones 3 and 4; (2) moderate: 2 to 4 signals in Zones 3 and 4; and (3) poor: <1 signal in Zones 3 and 4.

The primary outcomes were change in ET and EV. The secondary outcomes were positive pregnancy rate and clinical pregnancy rate.

The preparation of APRP is shown in Figure 1: 13.5 mL of venous blood was collected from the median cubital vein and added directly to the 1.5-mL anticoagulant (citrate-phosphate-dextrose-adenosine [CPDA]) (Figure 1, A). Sterile CPDA was obtained from blood bags from a blood bank (Figure 2). Each blood bag contained 49 mL of CPDA. To reach a sample size of 60 (3 cycles of PRP), 6 bags were used. The blood was centrifuged for 15 minutes at 1500 rpm using the REMI R-8C centrifuge machine (Remi Elektrotechnik Ltd, Mumbai, India). After the first spin, at least 50% of the volume should be plasma on top (yellow arrow in Figure 1, B). Then, PRP was collected without taking the buffy coat (blue arrow in Figure 1, B) on the red blood cell layer (red arrow in Figure 1, B). Plasma was transferred to another tube and centrifuged at 3000 rpm for 15 minutes to obtain platelet pellet (blue arrow in Figure 1, C); 10% plasma was left behind above the pellet, and the rest was discarded using a sterile pipette (Figure 1, D). The platelet pellet was resuspended in the remaining plasma using a sterile pipette, and a syringe was loaded and used immediately (Figure 1, E). The final preparation produced through this procedure has 4 to 5 times the concentration of platelets compared with that of whole blood, thus providing appropriate quantities of growth factors. Intrauterine instillation of this prepared APRP (0.5–1 mL) was done using IUI catheter under ultrasound guidance with all aseptic precautions. Pyrogen-free disposable materials were used. The preparation protocol was standardized to achieve platelet concentration 4 to 5 times that of baseline (Figure 3).

Figure 1.

Preparation of Platelet rich plasma.

A, Blood with the anticoagulant. B, After the first spin, 3 layers are formed: plasma (yellow arrow), buffy coat (blue arrow), and red blood cell layer (red arrow). C, After the second spin, platelet pellet (blue arrow) is obtained. D, Final platelet-rich plasma volume after discarding plasma. E, Final loading of the intrauterine insemination cannula.

Pandey. Role of autologous platelet-rich plasma for thin endometrium in ovarian stimulation cycle. Am J Obstet Gynecol Glob Rep 2023.

Figure 2.

Blood bag from which sterile anticoagulant CPDA was procured

CPDA, citrate-phosphate-dextrose-adenosine.

Pandey. Role of autologous platelet-rich plasma for thin endometrium in ovarian stimulation cycle. Am J Obstet Gynecol Glob Rep 2023.

Figure 3.

Baseline and postpreparation platelet concentration

HCT, Hematocrit; HGB, Hemoglobin; MCH, Mean Corpuscular Hemoglobin; MCHC, Mean Corpuscular Hemoglobin concentration; MCV, Mean Corpuscular Volume; MPV, Mean Platelet Volume; PCT, Plateletcrit; PDW, Platelet Distribution Width; P-LCC, Platelet Large Cell Count; P-LCR, Platelet Large Cell Ratio; PLT, Platelet; RBC, Red blood cells; RDW-CV, Red Cell distribution width-coefficient of variation; RDW-SD, Red Cell distribution width-standard deviation; WBC, White blood cells.

Pandey. Role of autologous platelet-rich plasma for thin endometrium in ovarian stimulation cycle. Am J Obstet Gynecol Glob Rep 2023.

Results

A total of 349 infertile women were screened for eligibility, of whom 229 were excluded, and 120 were randomized and allocated to the study and control groups of 60 each on the basis of computer-generated random number sequence. One women from the study group and 2 from the control group were lost to follow-up. Hence, final analysis was done on a total of 117 women (59 in the study group and 58 in the control group) (Figure 4).

Figure 4.

Consort diagram of the study

Pandey. Role of autologous platelet rich plasma for thin endometrium in ovarian stimulation cycle. Am J Obstet Gynecol Glob Rep 2023.

The 2 groups were similar in terms of clinical characteristics such as age, body mass index, duration of infertility, type of infertility, history of extragenital tuberculosis, and baseline ET.

There was significant increase in ET in both groups after 3 cycles. Intragroup statistical analysis in both groups showed that there was significant rise in ET from baseline to the end of successive cycles. The pregnancy test was positive in 23.7% (14/59) of women. The difference in clinical pregnancy rate between the study group (18.64%; 10/59) and the control group (6.9%; 4/58) was statistically significant (Table 1).

Table 1.

Basic clinicodemographic parameters, change in endometrial thickness, positive pregnancy rate, and clinical pregnancy rate in the study and control groups

		Study group (n=59)		Control group (n=58)
S.No	Parameters	N	%	N	%	P value
1.	Mean age (y)	29.2±1.89	—	29.11±1.89	—	.797a
2.	Body mass index	21.1±1.6	—	20.9±1.8	—	.676a
3.	Duration of infertility (y)	4.28±1.54	—	4.17±1.27		.675a
4.	Type of infertility
	Primary	53	89.8	51	87.9	.744b
	Secondary	6	10.2	07	12.1
5.	History of extragenital/genital tuberculosis
	Yes	25c	42.4	23	39.7	.765b
	No	34	57.6	35	60.3
6.	Endometrial thickness (mm)
	Baseline	4.66±0.79	—	4.99±0.77	—	.124a
	After 1 cycle	6.51±0.68		5.89±0.305		<.0001a
	After 2 cycles	6.68±0.78		5.87±0.77		<.0001a
	After 3 cycles	7.47±0.85		5.89±0.68		<.0001a
	Repeated-measures ANOVA	Baseline vs first P<.0001 Baseline vs second P<.0001 Baseline vs third P<.0001 first vs second P<.0001 first vs third P<.0001 second vs third P<.0001		Baseline vs first P<.0001 Baseline vs second P<.0001 Baseline vs third P<.0001 first vs second P=1 first vs third P=1 second vs third P=1
7.	Positive pregnancy rate	14/59d	23.73	4/58	6.89	.019e
8.	Clinical pregnancy rate	10/59f	18.64	4/58	6.89	.043e

P<.05 was considered as statistically significant.

^aIndependent t test

^bChi-square test

^cOut of 25 women with history of tuberculosis,21 had hypomenorrhea. Out of these 21(35.6%) women who had hypomenorrhea at the start of study,17(81%) noticed improvement in the menstrual pattern

^dOut of 14 with positive UPT, 10 had ultrasound documentation of cardiac activity (ie, clinical pregnancy), whereas 4 had positive UPT but negative clinical pregnancy; of these 4, 1 had biochemical pregnancy and 3 had missed abortion, and all 4 conceived in the second PRP cycle of IUI

^eFisher exact test

^fTen women with clinical pregnancy conceived in the third PRP cycle of IUI.

Pandey. Role of autologous platelet-rich plasma for thin endometrium in ovarian stimulation cycle. Am J Obstet Gynecol Glob Rep 2023.

The mean ET in the study group was 4.66±0.79 mm before PRP instillation, which increased significantly to 6.50±0.68 mm, 6.68±0.78 mm, and 7.47±0.85 mm after the first, second, and third cycle of intrauterine PRP instillation, respectively (P<.05) (Table 1; Figure 5).

Figure 5.

Mean ET before and after PRP instillation after each cycle

ET, endometrial thickness; PRP, platelet-rich plasma.

Pandey. Role of autologous platelet-rich plasma for thin endometrium in ovarian stimulation cycle. Am J Obstet Gynecol Glob Rep 2023.

EV improved significantly in the study group in comparison with the control group after 3 cycles (P<.001). There was significant improvement in vascularity after 3 cycles of APRP instillation (Figure 2). Initially, 77.9% of women had poor vascularity and 22.3% had moderate vascularity. However, at the end of the third cycle, excellent and moderate vascularity was achieved in 10.2% and 69.49% of patients, respectively, whereas 20.3% continued to have poor vascularity. This was a statistically significant change from the initial rates (P<.05) (Table 2; Figure 6).

Table 2.

Endometrial vascularity before and after platelet-rich plasma instillation in both groups

	Study group (n=59)			Control group (n=58)			P value
Vascularity	Poor	Moderate	Excellent	Poor	Moderate	Excellent
Baseline EV (n)	46	13	0	44	14	0	.787a
%	77.97	22.03	0	75.9	24.14	0
After first cycle	39	20	0	44	14	0	.245a
%	66.1	33.89	0	75.9	24.14	0
After second cycle	25	31	3	37	20	1	.345b
%	42.3	52.54	5.1	63.8	34.5	1.72
After third cycle	12	41	6	31	26	1	.0003b
%	20.33	69.49	10.2	53.45	44.83	1.72

EV, endometrial vascularity.

^aFisher Exact Test

^bChi Square Test.

Pandey. Role of autologous platelet-rich plasma for thin endometrium in ovarian stimulation cycle. Am J Obstet Gynecol Glob Rep 2023.

Figure 6.

Change in EV before and after PRP instillation in study group

EV, endometrial vascularity; PRP, platelet-rich plasma.

Pandey. Role of autologous platelet-rich plasma for thin endometrium in ovarian stimulation cycle. Am J Obstet Gynecol Glob Rep 2023.

Discussion

The PRP used in this study is the portion of the plasma fraction of autologous blood with approximately 5 times higher platelet concentration compared with circulating blood.¹² It has potential to create a local milieu rich in cytokines and growth factors helping in tissue regeneration. This has been successfully applied in various tissues.¹³ Four types of platelet concentrates have been described. These are: (1) leukocyte-poor or pure PRP (p-PRP), which was used in this study, (2) leukocyte PRP, (3) pure platelet-rich fibrin, and (4) leukocyte platelet-rich fibrin.¹⁴ The leukocyte content in platelet preparations increases inflammation and decreases tissue regeneration.^14,15 Thus, p-PRP was used in this study.

An anticoagulant is needed when preparing APRP, and it is chosen on the basis of its capacity to preserve platelets’ function, integrity, and morphology. Thus, EDTA was not used as an anticoagulant because it may damage the platelet membrane.¹³ Anticoagulants with citrate and dextrose of sodium citrate have been recommended for this purpose.¹³ Alternatively, use of CPDA has also been suggested, which is similar to ACD-A (Citra Labs, LLC, Braintree, MA) but has fewer supportive ingredients, making it 10% less effective in maintaining platelet viability.¹⁶,¹⁷

ACD-A is commercially available in PRP preparation kits, which are costly. Conversely, CPDA was a practical and cost-effective option for our circumstances because it was easily available and procurable from the blood bank of our hospital (Figure 2). Moreover, it is advisable for laboratories to standardize the preparation protocol while taking into account cost effectiveness and ease of adoption. In this study, the protocol was standardized to achieve platelet concentration 4 to 5 times the baseline level (Figure 3).

Although it is known that PRP acts via raised concentration of growth factors, the exact molecular mechanisms of PRP therapy in endometrial proliferation are not fully understood. A recent study by Aghajanova et al¹⁸ evaluated an in vitro model of activated PRP for endometrial regeneration. Activated PRP promoted migration of all the cells studied, namely human primary endometrial epithelial cells, endometrial stromal fibroblasts, endometrial mesenchymal stem cells (MSC), and bone marrow–derived MSC. These data provide an initial ex vivo proof of principle for use of APRP to promote endometrial regeneration in Asherman syndrome and thin endometrial lining, thus demonstrating the potential of APRP application in thin endometrium management,¹⁸ and prompting the inception of this study. Being autologous and prepared in aseptic conditions with use of pyrogen-free consumables, the safety of this preparation is beyond doubt.

Chang et al¹⁹ were the first to demonstrate the efficacy of intrauterine-instilled PRP in women with thin endometrium. Five women with thin endometrium (on the day of hCG administration) received intrauterine APRP on day 10 of FET cycle, which was repeated if ET did not improve after 72 hours. On achieving ET of at least 7 mm, embryo transfer was done. Successful endometrial growth and positive pregnancy were achieved in all 5 women.¹⁹ Subsequently, several studies demonstrated the effectiveness of intrauterine PRP instillation for thin endometrium and improved pregnancy outcomes in IVF cycles (Table 3).20, 21, 22, 23, 24, 25, 26 The live birth rates in these studies were above 25%. Velasco et al²⁷ also cited the use of APRP as a promising method for endometrial growth in women with refractory endometrium.

Table 3.

Comparison of results between this study and the published studies

Author	Year	Sample size Inclusion criteria	HRT+FET	IUI cycle	Post-PRP ET	Positive pregnancy rate	Clinical pregnancy rate	Missed abortion	EV
Chang et al19	2015	5 ET <7 mm on day of trigger in previous cycle despite HRT	yes	—	>7 mm in 100% of cases	100%	80%	20%	—
Colombo et al25	2017	8 3 canceled FET cycle because of ET <6 mm	—	—	>6.5 mm in 88%	85.7%	57%	14.3%	—
Zadehmodarres et al24	2017	10 ET <7 mm; 4 women with intrauterine adhesion	yes	—	>7 mm in 100%	50%	40%	—	—
Tandulwadkar et al22	2017	68 ET <7 mm/<5 vascular signals	yes	—	7.22 mm (mean)	60.9%	45.3%	7.35%	Significant improvement
Molina et al23	2018	19 h/o resistant endometrium with at least 1 failed IVF cycle	yes	—	>9 mm in 100%	73.7%	26.3%	2.6%	—
Eftekhar et al26	2018	40 ET <7 mm	yes	—	8.64±0.64 mm (mean)	12.7%	30%	—	—
Kim et al21	2019	20
Present study	2022	59 ET <7 mm at follicle size ≥16 mm despite estrogen in previous IUI cycle	—	yes	7.47±0.85 mm (mean)	23.7%	18.6%	8%	Significant improvement

ET, endometrial thickness; EV, endometrial vascularity; FET, frozen embryo transfer; h/o, history of; HRT, hormone replacement therapy; IUI, intrauterine insemination; IVF, in vitro fertilization, PRP, platelet-rich plasma.

Pandey. Role of autologous platelet-rich plasma for thin endometrium in ovarian stimulation cycle. Am J Obstet Gynecol Glob Rep 2023.

This study demonstrated the effect of APRP in OS-IUI cycles. Previous studies have proven that optimum ET on the day of IUI has positive impact on outcome.²⁸

In contrast to our results, Kim et al²¹ in their pilot study on women with refractory endometrium concluded that intrauterine PRP had no significant effect on ET. The increase in ET was only 0.6 mm, and there was no correlation between this increase and pregnancy outcome. The disparity may be because of the small sample size.

In addition to ET, EV is another crucial parameter that determines the implantation potential and hence positive pregnancy rates. The reason for low implantation potential in low EV is the high radial artery resistance leading to poor endometrial growth because of reduced VEGF.²⁹ Moreover, absence of the functional layer of the endometrium in women with thin endometrium exposes the embryo to high oxygen concentration because of proximity to spiral arterioles. Hence, thin endometrium is detrimental for embryo implantation and growth. This may explain the failed clinical pregnancies in 4 women in the study group. Nagori et al² demonstrated favorable conception rates with low abortion rates in women with Zone-3 and 4 EV, as opposed to Zone-1 and 2 (per Applebaum grading). Similar observations were made by Sardana et al,³⁰ who concluded that presence of EV significantly improves the outcome in FET cycles. Tandulwadkar et al²² were the first to demonstrate improvement in EV after intrauterine APRP instillation. EV improved significantly in this study as well. Moreover, it was evident in this study that with successive PRP cycles, the ET and EV increased, which can explain the conception and clinical pregnancy rates in the study group. All 10 women with positive clinical pregnancy conceived in the third PRP cycle. This indicates that PRP was helpful in the development of the functional layer of the endometrium. Amable et al⁹ demonstrated that levels of 12 proteins, that is, 6 growth factors (PDGF-AA, PDGF-AB, PDGF-BB, TGF-β1, TGF-β2, and EGF), 3 antiinflammatory cytokines (IL-4, IL-13, and IFN-α), and 3 proinflammatory cytokines (IL-8, IL-17, and TNF-α) were increased in activated platelets. This can explain the tissue regeneration and resultant increased vascularity.

The pregnancy rate in the control group remained 6.89%. This was similar to the result of Liu et al,³¹ who found that the pregnancy rate in gonadotropin-stimulated IUI cycles was 8.89% in women with ET <7 mm. Gonadotropins do not attenuate the endometrium, as observed with other common oral ovulogens such as clomiphene. Thus, letrozole with sequential gonadotropins was used for OS in IUI cycles in this study. Moreover, even in the control group there was statistically significant increase in ET in successive cycles (Table 1). This can be attributed to the use of gonadotropins in the stimulation protocol. However, the final ET remained thin with poor EV. The pregnancy rate was significantly higher in the study group (23.7%) when intrauterine instillation of APRP was done, indicating the positive trophic effect on thin endometrium in terms of both thickness and vascularity.

There was also improvement in the menstrual pattern among the enrolled women. Of the 21 (35.6%) women who had hypomenorrhea at the start of the study, 17 (81%) noticed improvement in the menstrual pattern (Table 2). Taking into account these observations, the use of PRP can be extended to women with menstrual abnormality, especially in low grades of the Asherman syndrome (after adhesiolysis) and possibly even genital tuberculosis.

Strengths and limitations

This was a randomized control study that evaluated both the ET and EV after intrauterine PRP infusion in women with thin endometrium during OS-IUI cycles. Moreover, CPDA—an easily available, low-cost anticoagulant—was used in this study and found to have positive effects on endometrium parameters. In addition to improving ET, EV, and the rate of achieving pregnancy, APRP therapy also improved menstrual patterns in many women. The study observed the effect of APRP therapy (pre and post) in the same sample and also compared it with the control group.

Further studies with larger sample sizes for more robust evidence can be conducted. There was lack of live birth outcome data and long-term follow-up in this study.

Conclusion

PRP can increase ET and improve EV by providing a local milieu rich in growth factors. For women with thin endometrium, APRP is a treatment modality that is safe, easy to prepare, free of adverse effects, and cost-effective, with no risk of immunologic reactions because it is prepared from autologous blood. This is a promising option that can be practiced in OS-IUI cycles to improve the endometrium, thereby reducing the financial and psychological burden in these women. Further large-scale randomized controlled trials can be planned, which can support our observations and enable reproductive clinicians to optimize success rates of OS-IUI cycles in women with thin endometrium.