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ORIGINAL ARTICLE
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Can intrauterine infusion of granulocyte colony-stimulating factor or platelet-rich plasma increase implantation rate in women undergoing in vitro fertilization with normal endometrium: boon or bane?


1 Department of Obstetrics and Gynaecology, AFMS, New Delhi, India
2 Department of Medical Informatics, AFMS, New Delhi, India
3 Department of Community Medicine, AFMS, New Delhi, India
4 Third year MCOMS, Pokhara, Nepal

Date of Submission28-May-2022
Date of Decision04-Aug-2022
Date of Acceptance10-Aug-2022
Date of Web Publication20-Oct-2022

Correspondence Address:
Seema Rai,
Department of Obstetrics and Gynaecology, AFMS, New Delhi
India
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/jmms.jmms_89_22

  Abstract 


Objective: The objective of this study was to investigate if intrauterine infusion of granulocyte colony-stimulating factor (G-CSF) or platelet-rich plasma (PRP) can improve endometrial parameters and the pregnancy rate in patients undergoing fresh embryo transfer with normal endometrial thickness. Materials and Methods: We conducted a retrospective observational study, at one of the assisted reproductive technique centers in central India. Data were collected from the institutional record system which was established as per the standard protocols. Data on a total of 225 patients who had an endometrial thickness >7 mm without any adjunct and underwent fresh embryo transfer in their first in vitro fertilization cycle during the past 2 years were collected and were categorized into three groups: Group A who had received intrauterine infusion of G-CSF (n = 75), Group B who received intrauterine infusion of PRP (n = 75), and Group C who (control group) received no intervention (n = 75). Results: The implantation rate was significantly higher in Group A (37.24%) than in Groups B and C (21.91% and 21.37%, respectively). No significant difference was found in the clinical pregnancy rate among Groups A, B, and C (50.68%, 40.54%, and 37.84%, respectively). Endometrial thickness increased significantly in Group A by approximately 2.3 mm. No significant change in other endometrial parameters in the three groups. The number of biochemical pregnancies was higher in the G-CSF group compared to the other two groups. Conclusion: Intrauterine infusion of G-CSF in women receiving fresh embryo transfer with normal endometrial thickness can increase endometrial thickness and the implantation rate without affecting other endometrial parameters and the cumulative pregnancy rate.

Keywords: Granulocyte colony-stimulating factor, platelet-rich plasma, endometrial receptivity, in vitro fertilizationW



How to cite this URL:
Rai S, Yasaswi K, Raj R, Hiremath RN, Singh S. Can intrauterine infusion of granulocyte colony-stimulating factor or platelet-rich plasma increase implantation rate in women undergoing in vitro fertilization with normal endometrium: boon or bane?. J Mar Med Soc [Epub ahead of print] [cited 2022 Dec 7]. Available from: https://www.marinemedicalsociety.in/preprintarticle.asp?id=359151




  Introduction Top


Assisted reproductive techniques (ARTs) have come a long way since the birth of the first baby through in vitro fertilization (IVF), Louise Brown, 1978. ART has witnessed dramatic advancements, and its success rate has increased from 10% to approximately 40%. This great leap in the success rate is due to an enhanced understanding of reproductive physiology and improved laboratory techniques. Tremendous research has led to an improvement and advancement in the stimulation regimen, culture media, and culture conditions. With the advent of intracytoplasmic sperm injection (ICSI), embryologists have been able to attain a fertilization rate of more than 80% and an increased yield of superior-quality (Grade 1) embryos. Moreover, ultrasound-guided embryo transfer has made feasible the transfer of embryos precisely at the maximum implantation point, thus augmenting further the success rate of IVF.

The key elements required for successful IVF are superior-quality embryos, receptive endometrium, and an experienced embryologist capable of performing precise embryo transfer.[1],[2] Because IVF is an expensive, time-consuming, and emotionally and financially draining procedure, attempts are made to ensure its success.[3],[4] Despite advancements in the field of ART, the IVF success rate has plateaued at 40%–45% and the take-home baby rate at approximately 32% in the best of centers. Implantation remains a rate-limiting step in ART despite superior-quality embryos and an ideal embryo transfer technique. Successful implantation relies on the presence of a receptive endometrium.[5] Other parameters affecting the success of the procedure are age, antral follicle count, anti-Müllerian hormone level, and the number of embryos transferred in the patient. Achieving the parameters apt for ensuring the success of implantation is a complex task that is associated with controversies and is poorly defined.

The endometrium is the innermost glandular layer of the uterus and acts as fertile soil for the implanting embryo. Implantation is a very delicate and well-orchestrated dialog between the implanting embryo and receptive endometrium mediated by a cascade of cytokines, chemokines, and growth factors.[6]

At the mid-luteal phase of the menstrual cycle, the endometrium becomes receptive and suitable for adherence of the blastocyst to the trophoblast; however, this implantation window is extremely short. Successful implantation and pregnancy is the ultimate test for endometrial receptivity. Various factors presumed to affect endometrial receptivity at the gross level include endometrial thickness, endometrial pattern, subendometrial and endometrial blood flow, and uterine artery blood flow. However, the interactions occurring at the immunological level also influence embryo implantation and sustenance and continuation of pregnancy. Different empirical methods in vogue employed to increase endometrial thickness and receptivity include extended estrogen administration and the administration of vaginal sildenafil, Vitamin E, pentoxifylline, aspirin, and low-molecular-weight heparin.[7],[8] However, none of the therapies have been found effective in increasing the clinical pregnancy rate, particularly in the presence of endometrium of requisite thickness (n ≥7 mm). Two novel adjuncts employed to increase the implantation rate are granulocyte colony-stimulating factor (G-CSF) and platelet-rich plasma (PRP).

This study was conducted to investigate the effects of G-CSF and PRP infusions on the pregnancy rate in patients with an endometrial thickness of ≥7 mm without using any adjuncts to reach optimum endometrial thickness. Furthermore, we determined whether G-CSF and PRP could be utilized as novel tools by embryologists to increase implantation and cumulative pregnancy rates in patients with normal endometrial thickness.


  Materials and Methods Top


We conducted a retrospective observational study, at one of the ARTs centers in central India. Data were collected from the institutional record system which was established as per the standard protocols. In this study, we collected data of patients who underwent their first IVF treatment with fresh embryo transfer during the past 2 years. We found a total of 225 patients who fitted into the inclusion criteria and categorized into three groups: Group A who had received intrauterine infusion of G-CSF (n = 75), Group B who received intrauterine infusion of PRP (n = 75), and Group C who (control group) received no intervention (n = 75) [Figure 1]. The primary objective of the study was to study the implantation rate, clinical pregnancy rate after intrauterine infusion of G-CSF and PRP, in women undergoing fresh embryo transfer cycles with optimal endometrial thickness and pattern as assessed by transvaginal ultrasound. Secondary objectives were to study the effect of intrauterine infusion of G-CSF and PRP on the endometrial thickness and vascularity and uterine artery indices. This study was approved by the institutional ethical committee.
Figure 1: Flow diagram depicting inclusion and analysis in the study. GCSF: Granulocyte colony-stimulating factor, PRP: Platelet-rich plasma

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Inclusion criteria: Patients (age: <40 years) who received fresh embryo transfer cycles were included in the study. Additional inclusion criteria were normoresponsiveness and endometrial thickness of >7 mm. Exclusion criteria: Patients were excluded on the basis of the following criteria: The presence of endometrial polyps or any uterine anomaly, such as fibroid; received a frozen embryo transfer cycle; having an endometrial thickness of <7 mm; and the presence of Asherman's syndrome; contraindication to G-CSF/PRP administration; received any other intervention to augment the endometrial thickness.

The first group (Group A) had received intracavitary infusion of 300 mcg/mL G-CSF (rHu G-CSF Neukine INTAS) through a soft embryo transfer catheter immediately after ovum pick-up (OPU) and vaginal examination for any bleeding under anesthesia. The second group had (Group B) received intracavitary infusion of 0.5 mL PRP (prepared on the day of OPU from autologous blood) through a soft embryo transfer catheter. The control group (Group C) had received no intervention. For fertilization, either insemination or ICSI were performed, as required. Ultrasound-guided embryo transfer was performed on day 3 at the 6–8 cell stage. Injection of micronized progesterone (100 mg/2 mL daily) and tablet dydrogesterone (10 mg BD) were used for luteal support. Data collected were entered into an Excel sheet, cleaned and analysed using SPSS Statistics for Windows, version 17. 0 (SPSS Inc., Chicago, Ill., USA).


  Results Top


[Table 1] presents the demographic characteristics of the patients and study outcomes. The age of the patients varied between 22 and 40 years; the average age was 30.52 ± 4.25, 29.65 ± 4.25, and 29.40 ± 7.07 years in Groups A, B, and C, respectively, and the difference was not statistically significant. The duration of infertility reported in Groups A, B, and C was also comparable (5.12 ± 2.83, 5.26 ± 1.41, and 5.57 ± 2.12, respectively). The total gonadotropin dose used and the estradiol level at the time of human chorionic gonadotropin trigger were higher in Group B, and the difference was statistically significant.
Table 1: Baseline characteristics of patients and study outcome

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The overall endometrial thickness increased significantly in Group A. Endometrial blood flow after G-CSF perfusion also improved in Group A compared with the PRP and control groups. The implantation rate in Group A was significantly higher than that in Groups B and C (37.24%, 21.92%, and 21.38%, respectively). However, no difference was observed in the clinical pregnancy rate between the three groups (P > 0.05) [Table 2].The number of biochemical pregnancies was higher in the G-CSF group, although no statistical significance was observed [Figure 2], [Figure 3], [Figure 4], [Figure 5], [Figure 6], [Figure 7].
Figure 2: Pre GCSF instillation: (a) endometrial thickness, 8.7 mm. (b) Color Doppler revealing poor subendometrial vascularity with only 1–2 small vessels in zone 1 and no vessels in zone 2. (c) Left uterine (UT) artery: RI, 0.92; PI, 3.44; and PSV, 31.6 cm/s; (d) Right UT artery: RI, 0.99; PI, 3.67; and PSV, 25.4 cm/s. PSV: Peak systolic velocity, RI: Resistive index, PI: Pulsatility index

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Figure 3: Post GCSF instillation: (a) endometrial thickness, 12.1 mm. (b) Color Doppler revealing improved endometrial flow and multiple vessels reaching zone 2. (c) Left UT artery: RI, 0.82; PI, 1.86; and PSV, 32.1 cm/s. (d) Right UT artery: RI, 0.81; PI, 2.04; and PSV, 25.2 cm/s. PSV: Peak systolic velocity, GCSF: Granulocyte colony-stimulating factor, RI: Resistive index, PI: Pulsatility index

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Figure 4: Pre PRP instillation: (a) endometrial thickness, 8 mm. (b) Color Doppler revealing no appreciable vessels in zones 1 and 2. (c) UT artery: RI, 0.84; PI, 2.22, and PSV; 35.7 cm/s. PSV: Peak systolic velocity, PRP: Platelet-rich plasma, RI: Resistive index, PI: Pulsatility index

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Figure 5: Post PRP instillation: (a) endometrial thickness, 9 mm. (b) Color Doppler revealing superior subendometrial vascularity with few vessels reaching zone 2. (c) UT artery: RI, 0.82; PI, 1.87; PSV, 37.5 cm/s. PSV: Peak systolic velocity, PRP: Platelet-rich plasma, RI: Resistive index, PI: Pulsatility index

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Figure 6: Control on the day of OPU: (a) endometrial thickness, 8 mm. (b) Color Doppler shows sub-endometrial flow. (c) UT artery: RI, 0.88; PI, 2.63; and PSV, 44.3 cm/s. PSV: Peak systolic velocity, OPU: Ovum pick-up, RI: Resistive index, PI: Pulsatility index

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Figure 7: Control on the day of ET: (a) endometrial thickness, 10 mm. (b) Color Doppler revels good endometrial and subendometrial flow. (c) UT artery: RI, 0.76; PI, 1.84; and PSV, 52.2 cm/s. PSV: Peak systolic velocity, RI: Resistive index, PI: Pulsatility index

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Table 2: Association of various outcome parameters with various groups of patients

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  Discussion Top


Infusion of G-CSF and PRP has resulted in higher implantation and pregnancy rates in patients with a thin endometrium (<7 mm). Based on this observation, we aimed to assess the effects of infusion of G-CSF and PRP in patients with endometrial thickness of >7 mm.

Until recently, the endometrium, a vital anatomical structure required for the initiation and continuation of pregnancy, had not received considerable attention.[9] Endometrial thickness and endometrial blood supply are measurable prognostic markers for endometrial receptivity that are essential for implantation. Ultrasound, being a noninvasive and convenient tool, in addition to assessing the anatomical details and folliculometry, can determine endometrial receptivity by evaluating the endometrial thickness, endometrial pattern, and endometrial blood flow.[10] Extended estrogen administration, low-dose aspirin, vaginal sildenafil, and antioxidants are some of the adjuncts that have been used empirically and investigated widely to enhance endometrial receptivity;[11] however, none of the therapies have been proven successful. A thin endometrium is a dependable indicator of poor endometrial receptivity, and embryos are usually frozen if the endometrial thickness is < 7 mm;[12] <1% of the patients undergoing IVF treatment exhibit a thin endometrium.[13]

G-CSF, as a cytokine, may prove beneficial in increasing cumulative pregnancy rates.[14] G-CSF infusion improves implantation and pregnancy rates in patients with a thin endometrium, which may be attributed to the decreased inherent capacity of the thin endometrium to produce G-CSF. The use of G-CSF, investigated for the first time by Gleicher et al. in four patients with a thin, nonresponsive endometrium has demonstrated great success.[15] The hypothesis underlying a lower implantation rate in patients with a thin endometrium is postulated as the direct exposure of an implanting embryo to a higher oxygen concentration near the basal layer in the presence of an extremely thin upper stratum functionalis.[16] G-CSF has emerged as a promising adjunct in enhancing the success rate of IVF by increasing synchrony between the endometrium and embryo, and this growth factor has been investigated by several researchers for its utility in a thin, nonresponsive endometrium, and in recurrent implantation failures with varying results.[15],[17] G-CSF has been investigated systematically as well as locally by different researchers; however, the results of G-CSF infusion studies are inconsistent. Some studies have demonstrated high implantation and pregnancy rates with subcutaneous administration of G-CSF,[18],[19] which may be attributed to the favorable effects of G-CSF on oocyte quality and embryonic development. G-CSF, secreted by the endometrium, is also produced by granulosa cells during ovulation, promoting follicular growth and steroidogenesis when administered subcutaneously.[20] Some studies have demonstrated increased clinical pregnancy rates after G-CSF administration without any effect on the endometrium, implicating the influence of G-CSF at different implantation stages and levels, and making it an appropriate adjunct for increasing the pregnancy rate.[17],[21] At the immunological level, G-CSF decreases interferon-γ production and cytotoxicity of decidual mononuclear cells and magnetic cell sorting (MACS) CD56 cells.[22] G-CSF infusion in the general population with adequate endometrium thickness has not been found encouraging in terms of pregnancy rates.[23] However, some researchers have found that endometrial thickness and endometrial vascularity increase with G-CSF infusion.[1]

PRP has been extensively used in ophthalmology, orthopedics, dermatology, and other medical fields due to its positive effect on tissue repair and regeneration.[5] PRP is obtained from fractionation of the autologous blood with the concentration of platelets 4–5 times more than normal.[24] PRP being harvested from autologous blood may be a safe, effective, and affordable adjunct for improving the implantation rate.[15] Due to its influence on the repair and regeneration of tissues, researchers have extended its use in reproductive medicine. A few studies have claimed its benefit on ovarian rejuvenation, thin endometrium, and recurrent implantation failures.[24],[25],[26] On separation, platelets get activated and secrete >300 active cytokines and a high concentration of growth factors, such as vascular endothelial growth factor, transforming growth factor, platelet-derived growth factor, and epidermal growth factor.[27] These cytokines and growth factors are considered to exert a positive paracrine influence on implantation by moderating the proliferation of cell adhesion and immune response and promoting angiogenesis. A few studies have investigated the effect of PRP on a thin or refractory endometrium,[24],[28] and all have demonstrated encouraging results.

Our study is probably the first to compare the effects of PRP and G-CSF on normal thickness endometrium (>7 mm). In the present study, we observed the beneficial effect of G-CSF infusion on endometrial proliferation and the implantation rate but not on the pregnancy rate. However, the number of biochemical pregnancies is also increased with G-CSF infusion, which tends to give an illusion of a positive pregnancy outcome and a false hope to the patient. PRP infusion was also found to increase endometrial proliferation but without improving the implantation and pregnancy rates. Overall, our findings suggest that intrauterine infusion of the endometrial cavity with G-CSF before embryo transfer can increase endometrial proliferation and the implantation rate. Intrauterine perfusion of G-CSF on the day of OPU can be a novel strategy for embryologists aiming to further increase the implantation rates.


  Conclusion Top


Intrauterine infusion of G-CSF in women receiving fresh embryo transfer with normal endometrial thickness can increase endometrial thickness and the implantation rate without affecting other endometrial parameters and the cumulative pregnancy rate. PRP infusion was also found to increase endometrial proliferation but without improving the implantation and pregnancy rates. However, more extensive studies are required to validate the results of our study.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.



 
  References Top

1.
Jain S, Mahey R, Malhotra N, Kalaivani M, Sangeeta P, Bhatt A, et al. Effect of intrauterine perfusion of granulocyte colony-stimulating factor on endometrial parameters and in vitro fertilization outcome in women undergoing in vitro fertilization/intracytoplasmic sperm injection cycles: A randomized controlled trial. J Hum Reprod Sci 2018;11:254-60.  Back to cited text no. 1
[PUBMED]  [Full text]  
2.
Gardner DK, Weissman A, Howles CM, Shoham Z, editors. Textbook of Assisted Reproductive Techniques – Clinical Perspectives. 4th ed. Boca Raton: Taylor and Francis Group CRC Press; 2012.  Back to cited text no. 2
    
3.
Li J, Mo S, Chen Y. The effect of G-CSF on infertile women undergoing IVF treatment: A meta-analysis. Syst Biol Reprod Med 2017;63:239-47.  Back to cited text no. 3
    
4.
Zeyneloglu HB, Onalan G. Remedies for recurrent implantation failure. Semin Reprod Med 2014;32:297-305.  Back to cited text no. 4
    
5.
Farimani M, Poorolajal J, Rabiee S, Bahmanzadeh M. Successful pregnancy and live birth after intrauterine administration of autologous platelet-rich plasma in a woman with recurrent implantation failure: A case report. Int J Reprod Biomed 2017;15:803-6.  Back to cited text no. 5
    
6.
Robert CA, Abbas MK, Zaidi AR, Thiha S, Malik BH. Mediator in the embryo-endometrium cross-talk: Granulocyte colony-stimulating factor in infertility. Cureus 2019;11:e5390.  Back to cited text no. 6
    
7.
Zhao J, Xu B, Xie S, Zhang Q, Li YP. Whether G-CSF administration has beneficial effect on the outcome after assisted reproductive technology? A systematic review and meta-analysis. Reprod Biol Endocrinol 2016;14:62.  Back to cited text no. 7
    
8.
Gargett CE, Healy DL. Generating receptive endometrium in Asherman's syndrome. J Hum Reprod Sci 2011;4:49-52.  Back to cited text no. 8
[PUBMED]  [Full text]  
9.
Eftekhar M, Sayadi M, Arabjahvani F. Transvaginal perfusion of G-CSF for infertile women with thin endometrium in frozen ET program: A non-randomized clinical trial. Iran J Reprod Med 2014;12:661-6.  Back to cited text no. 9
    
10.
Wang L, Qiao J, Li R, Zhen X, Liu Z. Role of endometrial blood flow assessment with color Doppler energy in predicting pregnancy outcome of IVF-ET cycles. Reprod Biol Endocrinol 2010;8:122.  Back to cited text no. 10
    
11.
Cakmak H, Taylor HS. Implantation failure: Molecular mechanisms and clinical treatment. Hum Reprod Update 2011;17:242-53.  Back to cited text no. 11
    
12.
Singh N, Bahadur A, Mittal S, Malhotra N, Bhatt A. Predictive value of endometrial thickness, pattern and sub-endometrial blood flows on the day of hCG by 2D doppler in in-vitro fertilization cycles: A prospective clinical study from a tertiary care unit. J Hum Reprod Sci 2011;4:29-33.  Back to cited text no. 12
[PUBMED]  [Full text]  
13.
Singh M, Chaudhry P, Asselin E. Bridging endometrial receptivity and implantation: Network of hormones, cytokines, and growth factors. J Endocrinol 2011;210:5-14.  Back to cited text no. 13
    
14.
Crawford G, Ray A, Gudi A, Shah A, Homburg R. The role of seminal plasma for improved outcomes during in vitro fertilization treatment: Review of the literature and meta-analysis. Hum Reprod Update 2015;21:275-84.  Back to cited text no. 14
    
15.
Gleicher N, Vidali A, Barad DH. Successful treatment of unresponsive thin endometrium. Fertil Steril 2011;95:7.e13-7.  Back to cited text no. 15
    
16.
Casper RF. It's time to pay attention to the endometrium. Fertil Steril 2011;96:519-21.  Back to cited text no. 16
    
17.
Gleicher N, Kim A, Michaeli T, Lee HJ, Shohat-Tal A, Lazzaroni E, et al. A pilot cohort study of granulocyte colony-stimulating factor in the treatment of unresponsive thin endometrium resistant to standard therapies. Hum Reprod 2013;28:172-7.  Back to cited text no. 17
    
18.
Santjohanser C, Knieper C, Franz C, Hirv K, Meri O, Schleyer M, et al. Granulocyte-colony stimulating factor as treatment option in patients with recurrent miscarriage. Arch Immunol Ther Exp (Warsz) 2013;61:159-64.  Back to cited text no. 18
    
19.
Aleyasin A, Abediasl Z, Nazari A, Sheikh M. Granulocyte colony-stimulating factor in repeated IVF failure, a randomized trial. Reproduction 2016;151:637-42.  Back to cited text no. 19
    
20.
Würfel W. Treatment with granulocyte colony-stimulating factor in patients with repetitive implantation failures and/or recurrent spontaneous abortions. J Reprod Immunol 2015;108:123-35.  Back to cited text no. 20
    
21.
Lédée N, Gridelet V, Ravet S, Jouan C, Gaspard O, Wenders F, et al. Impact of follicular G-CSF quantification on subsequent embryo transfer decisions: A proof of concept study. Hum Reprod 2013;28:406-13.  Back to cited text no. 21
    
22.
Morelli S, Mandal M, Goldsmith LT, Kashani BN, Ponzio NM. The maternal immune system during pregnancy and its influence on fetal development. Res Rep Biol 2015;6:171-89.  Back to cited text no. 22
    
23.
Eftekhar M, Hosseinisadat R, Baradaran R, Naghshineh E. Effect of granulocyte colony stimulating factor (G-CSF) on IVF outcomes in infertile women: An RCT. Int J Reprod Biomed 2016;14:341-6.  Back to cited text no. 23
    
24.
Chang Y, Li J, Chen Y, Wei L, Yang X, Shi Y, et al. Autologous platelet-rich plasma promotes endometrial growth and improves pregnancy outcome during in vitro fertilization. Int J Clin Exp Med 2015;8:1286-90.  Back to cited text no. 24
    
25.
Zwiep T, Humphrey R, Fortin D, Inculet RI, Malthaner RA. Autologous platelet rich plasma and concentrated platelet poor plasma are safe in patients requiring lobectomies but do not reduce the duration of air leak: A randomised controlled trial. Ann Surg Int 2016;2:ASI-2-011.  Back to cited text no. 25
    
26.
Toliopoulos IK, Papageorgiou S. Ovarian rejuvenation therapy with PRP (plasma rich in platelets) an innovative solution for women's infertility. J Transl Sci 2018;5:1-2.  Back to cited text no. 26
    
27.
Golebiewska EM, Poole AW. Platelet secretion: From haemostasis to wound healing and beyond. Blood Rev 2015;29:153-62.  Back to cited text no. 27
    
28.
Zadehmodarres S, Salehpour S, Saharkhiz N, Nazari L. Treatment of thin endometrium with autologous platelet-rich plasma: A pilot study. JBRA Assist Reprod 2017;21:54-6.  Back to cited text no. 28
    


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