A pilot study on effect of autologous platelet-rich plasma on refractory thin endometrium in frozen embryo transfer cycle

INTRODUCTION

Prediction of pregnancy outcome following in vitro fertilization(IVF) remains elusive. The endometrium plays a major role during assisted reproductive technology (ART) treatment. Implantation involves a synchronised process of attachment and invasion of the embryo into the endometrium. It starts with the apposition of the maternal and embryonic epithelium, followed by adhesion, penetration and invasion of placental cells deep into the endometrium.^[1] The tissue of the endometrium contains various receptors of growth factors, cytokines, and lipids which are vital for endometrial and embryonic development which is necessary for successful implantation.^[2] In frozen embryo transfer (FET) cycles, implantation mainly depends on endometrium receptivity and embryo quality and technique of transfer. On transvaginal ultrasonography endometrium thickness, blood flow and its pattern can be measured. A thin endometrium is encountered in 2.4% of ART cycles.^[3] Though pregnancies have been reported at 4 and 5 mm it is apparent that an endometrial thickness <7 mm is associated with a trend toward lower probability of pregnancy.^[4] There are around 0.6–0.8% cycles of IVF in which embryo transfer is cancelled because of thin endometrium, that is less than 7 mm.^[5] Angiogenesis is an important parameter for endometrial growth. Vascular epidermal growth factor (VGEF) is a significant mediator for hormonal dependent endometrium growth. The decreased expression of VGEF receptor due to increased resistance in the uterine arteries results in poor endometrial growth. Hormonal manipulation like an extended dose of estrogen or improving endometrial perfusion by low dose aspirin, pentoxifylline and vitamin E, sildenafil, and new modalities like Granulocyte colony-stimulating factor (G-CSF) are used for endometrial expansion.^[6] However, these methods cannot improve all cases with thin endometrium.

Platelet- rich Plasma (PRP) application is a successful catalyst in the healing process for wide varieties of conditions. PRP therapy began gaining popularity in the mid 90s.^[7] Methods of harvesting and using PRP are becoming widely practised in the fields of sports medicine, dentistry, dermatology and cosmetology and aesthetic medicine. Main function is haemostasis by attaching to any breach in vessel where platelets aggregate, activate and release granules. Alpha granules contain clotting mediators such as factor V, factor VIII, fibrinogen, fibronectin, platelet-derived growth factor (PDGF) and chemotactic factors.^[8] Activated platelets release granules by exocytosis which contain more than sixty different biologically active substances that are involved in processes of tissue regeneration, angiogenesis, epithelialization, and proliferation. PRP infusion is newer modality of treatment in thin endometrium. It is prepared from autologous blood plasma that is concentrated with platelets.

The aim of our study was to evaluate the effect of intrauterine infusion of platelet-rich plasma in case of thin endometrium and its impact on endometrial thickness and clinical pregnancy rate.

MATERIAL AND METHODS

Study design: Prospective clinical intervention

Study setting: Reproductive Biology & IVF Centre, Maulana Azad Medical College (MAMC), New Delhi

Study duration: 6 months (1^st October 2018 to 1^st April 2019)

STATISTICAL METHODS

Statistical testing was conducted with the statistical package for the social science system version SPSS 17.0. The comparison of continuous variables between the groups was performed using Student’s t test. Nominal categorical data between the groups were compared using Chi-squared test. For all statistical tests, a P-value less than 0.05 will be taken to indicate a significant difference.

RESULTS

In the study, 24 patients were enrolled who fulfilled the inclusion and exclusion criteria. Optimal endometrial thickness could not be achieved in four patients and hence were excluded.

The average age of the patients was 39.04 years. The mean duration of infertility in these women was 19.02 years. Out of which, 14 women had primary infertility while 10 had secondary infertility. Various causes of infertility among these women included tubal (n=4), endometritis (n = 4), unexplained (n = 6), DOR (n = 8) and male factor (n = 2) [Table 1].

Table 1 Causes of infertility

Patients were given estradiol valerate (tablet Estrabet, Abott India) from day 2/3 of menses. The mean duration of estrogen given was 18.37 days. The mean of pre-treatment endometrial thickness was 4.68±0.96 mm while after treatment with PRP it was 6.65±0.52 mm (P < 0.05) [Figure 1]. The mean of pre-treatment flow zone is 1.96± 0.75 while that of post-treatment is 3.63±0.49 (P < 0.001) [Figure 2]. Biochemical pregnancy was seen in 55%, while clinical pregnancy rate was 45% [Table 2].

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Figure 1

Post and pre PRP flow zone of the endometrium.

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Figure 2

Endometrial thickness pre and post PRP treatment.

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Table 2 Biochemical and clinical pregnancy rate

DISCUSSION

Optimal endometrium thickness is one of the critical factors for successful embryo implantation. It is accepted that an endometrial thickness of less than 7 mm would be considered as refractory and that would entail to altered pregnancy and live birth. Refractory endometrium can be idiopathic, congenital, surgical (uterine curettage); caused by inflammatory processes, infections and radiation therapy, among other causes.^[9] Consequently, endometrium preparation has been considered a crucial step for embryo transfer. PRP is an autologous blood plasma that has been enriched with platelets. It stimulates proliferation and regeneration with a large amount of growth factors and cytokines, including PDGF, TGF, VGEF, EGF, fibroblast growth factor (FGF), insulin-like growth factor I, II (IGF I, II), interleukin 8 (IL-8) and connective tissue growth factor (CTGF).^[10] PRP has a role in improving implantation rate and clinical pregnancy rate with thin endometrium patients.^[11] Tandulwadkar et al. assessed not only the endometrial thickness, but also the endometrial vascularity after PRP infusion. They included 68 patients in their study, 64 achieved optimal thickness for FET cycle. Similar to our study, of the 64 patients that underwent FET, endometrial vascularity increased in all of them. Average mean lining thickness before PRP infusion was 5mm, and 7.22 mm after the infusion. They also stated that it decreases the cycle cancellation and psychological and financial stress of infertile women.^[12] The main advantage of autologous PRP is that it is safe, with no side effects, cheap, easily accessible treatment of refractory endometrium.

In 2015, Chang et al.^[13] studied the intrauterine infusion of PRP for thin endometrium. They instilled PRP prepared from patient’s own blood in 5 women with <7mm endometrial thickness. Post treatment the ET was increased >7mm. All these patients became pregnant after successful embryo transfer, only 1 had missed abortion. Chang Y et al.^[14] did a prospective cohort study and included 42 in study group and 30 patients in control. After PRP infusion, the average endometrium thickness on the day of progesterone administration in PRP group was 7.65 + 0.22mm which was significantly thicker than the control group (6.52 +0.31). They concluded that PRP plays an active role in promoting endometrium proliferation, improving embryo implantation rate and clinical pregnancy rate. These results agree with our study. In the same way, 10 patients were evaluated by Zadehmodarres et al who had history of cancelled cycles due to inadequate endometrial growth (<7mm). They also instilled PRP twice, and found that the endometrial thickness increased to more than 7 mm. Five patients became pregnant (50%) after the embryo transfer. They concluded that PRP was effective for endometrial growth in infertile women with a thin endometrium.^[11]

Garcia-Velasco et al performed an extensive review on the management of refractory endometrium. The review included studies with estradiol valerate, as well as various drugs including acetylsalicylic acid (aspirin), sildenafil, vitamin E, GnRH agonists, HCG, l-arginine, pentoxifylline among others; as well as autologous preparations as growth factors, mainly G-CSF, stem cells, PRP and bone marrow. They concluded, that despite the vast basket of resources available, it is not easy to provide an evidence-based approach on how to improve refractory endometrium.^[15]

CONCLUSION

Thin refractory endometrium still remains one of the challenges. The use of the PRP preparation and its effect on the endometrium seems to offer benefits to refractory endometrium, providing an increase in endometrial receptivity and a consequent increase in implantation rates. Being an autologous resource, it is harmless to the patient, easy to obtain and of very low cost. It is recommended to be included in the different protocols for endometrial preparation in assisted reproduction techniques. Further studies are recommended in terms of the population size as well as the approach of comparative studies between drugs and autologous preparations.