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Endometrial cell-derived conditioned medium in combination with platelet-rich plasma promotes the development of mouse ovarian follicles

Published online by Cambridge University Press:  02 November 2022

Neda Taghizabet
Affiliation:
Histomorphometry and Stereology Research Center, Shiraz University of Medical Sciences, Shiraz, Iran Department of Anatomical sciences, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran
Fatemeh Rezaei-Tazangi
Affiliation:
Department of Anatomy, School of Medicine, Fasa University of Medical Sciences, Fasa, Iran
Mahboubeh Mousavi
Affiliation:
Department of Anatomy, Faculty of Medicine, Semnan University of Medical Sciences, Semnan, Iran
Farzaneh Dehghani
Affiliation:
Histomorphometry and Stereology Research Center, Shiraz University of Medical Sciences, Shiraz, Iran Department of Anatomical sciences, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran
Nehleh Zareifard
Affiliation:
Histomorphometry and Stereology Research Center, Shiraz University of Medical Sciences, Shiraz, Iran Department of Anatomical sciences, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran
Soha Shabani
Affiliation:
Faculty of Veterinary medicine, Azad University, Research Sciences Branch
Soghra Bahmanpour
Affiliation:
Department of Anatomical sciences, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran
Fereshteh Aliakbari
Affiliation:
Men’s Health and Reproductive Health Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran
Zahra Sadeghzadeh
Affiliation:
Men’s Health and Reproductive Health Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran
Hengameh Dortaj
Affiliation:
Department of Anatomy, ShahidSadoughi University of Medical Sciences, Yazd, Iran
Arezoo Chakerzehi
Affiliation:
Department of Biochemistry, ShahidSadoughi University of Medical Sciences, Yazd, Iran
Gholamreza Mohseni*
Affiliation:
Men’s Health and Reproductive Health Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran
*
Author for correspondence: Gholamreza Mohseni, Men’s Health and Reproductive Health Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran. Tel: +98 9181311734. E-mail: [email protected]
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Summary

Fertility preservation is one of the most important issues in assisted reproductive technology. Previous studies have shown that cytokines and growth factors can improve follicle growth. The endometrial stromal cells secrete various factors that are involved in maintaining the integrity of uterine and epithelial secretory function. The platelet-rich plasma contains a large assembly of platelets suspended in plasma that successfully improves the viability and growth of various cell lines. This work aimed to investigate the influences of conditioned medium (CM) and platelet-rich plasma (PRP) on the development of ovarian follicles in infertile mice due to cyclophosphamide (CYC) exposure. In this study, 65 healthy BALB/c female mice (∼28–30 g and 6-8 weeks old) in five groups were studied. Immunohistochemistry (IHC) was used to detect growth differentiation factor 9 (GDF9)-positive cells. The mRNA expression levels of SMAD1, SMAD2, and BMP15 was assessed using reverse transcription-polymerase chain reaction (RT-PCR) method. The expression levels of SMAD1, GDF9, BMP15, and SMAD2 in the CM+PRP group was significantly more than in the CM and PRP groups. In addition, live birth occurred in the CM+PRP group. Treatment with CM+PRP in infertile mice due to Cy exposure increased fertility and live-birth rate. In general, our study suggested that the CM and PRP combination could improve the growth of mice ovarian follicles in vivo.

Type
Research Article
Copyright
© The Author(s), 2022. Published by Cambridge University Press

Introduction

Infertility is a common and worrying issue for couples. Therefore, it is necessary to find methods to prevent infertility. Statistical studies have shown that 50% of infertility cases in couples are due to female factors (Turner, Reference Turner2003; Fathi et al., Reference Fathi, Valojerdi, Ebrahimi, Eivazkhani, Akbarpour and Tahaei2017). Some women develop premature ovarian failure (POF) so that, before the age of 40, the production of estrogen in the ovary stops and no ovulation takes place. The etiology of POF is unknown yet; however, it may be related to genetic disorders, autoimmune diseases, infections, enzyme deficiencies, metabolic syndromes and smoking. In some cases, it is due to medical interventions such as ovarian surgery or long-term use of GnRH or chemotherapy drugs (Van Der Voort et al., Reference Van Der Voort, van Der Weijer and Barentsen2003; Okeke et al., Reference Okeke, Anyaehie and Ezenyeaku2013). Cyclophosphamide (Cy) is described as an alkylating compound utilized for the therapy of various malignant and non-malignant disorders; however, it can have detrimental impacts, such as pulmonary fibrosis, gastrointestinal disorder, kidney infection, mutagenesis, and impaired fertility and could stimulated POF (Meirow et al., Reference Meirow, Assad, Dor and Rabinovici2004). The Cy also causes atrophy of the ovaries, shrinkage of the ovarian follicles, and reduction of the number of follicles in the stages of primary, secondary, and antral. This paves the way for ovarian infertility and infertility (Jiang et al., Reference Jiang, Zhao, Qi, Li, Zhang, Song, Yu and Gao2013). Utilizing platelet-rich plasma (PRP) in clinics has gained particular interest since the 1970s. The PRP is derived from autologous blood whose platelet concentration is significantly high and is rich in growth factors, for example, connective tissue growth factor, platelet-derived growth factor (PDGF), keratinocyte growth factor, interleukin 8 (IL-8), transforming growth factor beta (TGF-β), fibroblast growth factor (FGF), epidermal growth factor (EGF), and vascular endothelial growth factor (VEGF) (Sugiura et al., Reference Sugiura, Su, Li, Wigglesworth, Matzuk and Eppig2009; Lin et al., Reference Lin, Jia and Zhang2011; Nagashima et al., Reference Nagashima, Kim, Li, Lydon, DeMayo, Lyons and Matzuk2011; Dhillon et al., Reference Dhillon, Schwarz and Maloney2012). As many people with autoimmune diseases and cancer are at risk of losing their ovarian reserve or for whatever reason oocytes are unable to reach puberty in vivo, obtaining a protocol to improve the developmental process and ovarian reserve recovery is needed.

In this study, an attempt was made to provide an environment that improves the development process and restore the ability of damaged ovary. According to recent studies, conditioned medium (CM) or the optimal culture medium derived from cells includes a variety of enzymes, cytokines, growth factors, and hormones involved in regulating cell growth, differentiation, repair, and angiogenesis (Chen et al., Reference Chen, Zhuang, Chen and Huang2011; Srivastava et al., Reference Srivastava, Sengupta, Kriplani, Roy and Ghosh2013; Hosseini et al., Reference Hosseini, Shirazi, Naderi, Shams-Esfandabadi, Borjian Boroujeni, Sarvari, Sadeghnia, Behzadi and Akhondi2017). Given that PRP contains many of the factors required for folliculogenesis, it seems that PRP and CM can be effective in improving the developmental process and restoring the ability of the damaged ovary (Malekshah et al., Reference Malekshah, Moghaddam and Daraka2006; Hosseini et al., Reference Hosseini, Shirazi, Naderi, Shams-Esfandabadi, Borjian Boroujeni, Sarvari, Sadeghnia, Behzadi and Akhondi2017; Ahmadian et al., Reference Ahmadian, Sheshpari, Pazhang, Bedate, Beheshti, Abbasi, Nouri, Rahbarghazi and Mahdipour2020). Growth differentiation factor 9 (GDF9) and bone morphogenetic protein 15 (BMP15) genes, released by the oocytes, are capable of modulating primordial follicle recruitment, granulosa cell differentiation and proliferation, and steroids synthesis, and increasing preantral follicles growth (Fenwick et al., Reference Fenwick, Mora, Mansour, Baithun, Franks and Hardy2013). Bone morphogenetic proteins (BMPs), as one of the members of superfamily of the transforming growth factor-β (TGF-β) in the ovary, regulate cell proliferation, migration, and stem cell differentiation. The GDF-9 and BMP15 are expressed in each phase of follicular growth and have a role in monitoring and steroidogenesis and proliferation of granulosa cells. The Smad family is a subgroup of proteins belonging to signal transduction molecules responsible for the transmission of TGF-β signals from the cell surface into the nucleus. Smad1, Smad2, and Smad3 are receptor-activated Smads related to both activin and TGF-β signalling (Xu et al., Reference Xu, Oakley and McGee2002). This is the first report to introduce the development of mice ovarian follicles using CM derived from endometrial cells and PRP.

Materials and methods

Animals

In total, 65 healthy BALB/c female mice (∼28–30 g and 6-8 weeks old) in five groups were studied. There were 10 mice in each group and adult female mice were used for blood sampling, preparation of PRP, and conditioned medium (CM). Animals were kept in normal conditions (temperature: 21 ± 2°C and light and dark cycle: 12 h) with free accessibility to water and food in standard cages. All procedures were done according to the instruction of the Ethics Committee of Shahid Beheshti University of Medical Sciences (Reference number: IR.SBMU.RETECH.REC.1399.247).

Experimental groups

Control: received normal saline intraperitoneally, Sham (CYC): They only received cyclophosphamide (CYC), (i.p.). CM: First received cyclophosphamide and after a week of infertility, received conditioned medium i.p. (0.5 ml) (Pouya et al., Reference Pouya, Heidari, Baghaei, Asadzadeh Aghdaei, Moradi, Namaki, Zali and Hashemi2018) on 1st, 7th, and 14th days. PRP: First received cyclophosphamide and after a week of infertility, received PRP (0.5 ml) i.p. on 1st, 7th, and 14th days.CM+PRP: First, received cyclophosphamide and after a week of infertility, received (0.5 ml) i.p. on 1st, 7th, and 14th days. The list of experimental groups is given in Table 1.

Table 1. Experimental groups

Preparation of conditioned media

The required CM was obtained from endometrial cells (in the prime phase of the menstrual cycle [menstrual phase]). The culture of newly extracted stromal cells and epithelial cells (30,000 cells) was performed in the growth medium in six-well plates for 1 day, and then they were washed with phosphate-buffered saline (PBS) and subsequently substituted with growth medium (3 ml). In the next step, collecting, filtering, and sterilizing of the CM were carried out (After 2 days of culture), and finally, it was stored at −80 °C until the experiment. For concentrating the factors secreted from the cells of endometrial niche, the stromal and epithelial cells were cultured in serum-free DMEM/F-12 medium (1 ml). After that, the secreted agents in the CM (after 48 h) were concentrated (CCM) by centrifuging (4000 g for 20 min at 4 °C) through the use of Amicon ultra-15 centrifugal filter devices (EMD Millipore) with a molecular weight cutoff of 10 kDa. The amount of the concentrated protein originated from one culture well was assumed as one unit and 1 ml of concentrated (CCM) was used (Srivastava, et al; Reference Srivastava, Sengupta, Kriplani, Roy and Ghosh2013).

Preparation of platelet-rich plasma

After collecting venous blood (10 ml) from models of induced disc-degeneration by syringes (10 ml), including 3000 U of heparin, the obtained blood sample was transferred to a tube (10 ml) and subsequently centrifuged at 2400 rpm for 10 min. Next, the whole buffy coat and supernatant in the tube were collected and transferred into another tube (10 ml). After centrifuging the tube at 3600 rpm for 15 min, upper three-quarters of the supernatant in the tube were removed, and the remaining section was considered as the PRP. The PRP solution was activated with calcium gluconate to a final concentration of 1 × 108 cells/ml. (Dehghani, et al; Reference Dehghani, Aboutalebi, Esmaeilpour, Panjehshahin and Bordbar2018, Aflatoonian, etal; Reference Aflatoonian, Lotfi, Saeed and Tabibnejad2021)

Preparation of cyclophosphamide

75 mg/kg of Cy powder (Sigma, USA) was dissolved in 0.9% normal saline. Then, the injection was performed based on the animal’s weight (Dehghani, et al; Reference Dehghani, Aboutalebi, Esmaeilpour, Panjehshahin and Bordbar2018).

Induction of infertility in mice using cyclophosphamide

Mice weight was measured and Cyclophosphamide was prescribed at 75 mg/kg by i.p. injection irrespective of the reproductive cycle (single dose). (Dehghani, et al; Reference Dehghani, Aboutalebi, Esmaeilpour, Panjehshahin and Bordbar2018.)

Tissue processing

The fixation of ovarian tissue samples was conducted (overnight) in 4% (w/v) paraformaldehyde/0.02 M PBS (pH 7.2) at 4°C. After that, their washing in running water, dehydrating by gradient alcohol (70, 80, 90, 95, and 100% alcohol I, II), and transparentizing in xylene were performed. Following embedding in paraffin, each paraffinized ovary sample was sectioned (serially) at a 4–5 μM thickness. Totally, 20 slides were evaluated, these were selected from every 30 slides to prevent the evaluation of the same follicle more than once.

Immunohistochemistry

The sections were fixed in PBS comprising 4% paraformaldehyde for 20 min. In the next step, the non-specific binding sites were blocked with PBS comprising 10% horse serum, 0.3% Triton X-100, and 1% BSA for 45 min. The primary antibodies, i.e., GDF-9 (sc-12244 Santa Cruz) and GDF-9B (sc-28911 Santa Cruz), were diluted at 1:200 and 1:100 ratios, respectively. Then, the sections were incubated (overnight) at 4° C in the suitable dilutions of the mentioned antibodies. Subsequently, incubation with secondary antibody rabbit anti-goat SABC Kit was performed. After washing with PBS comprising BSA, the cells were counter stained with 40,6-diamidino-2-phenylindole (DAPI) (Sigma) for 5 min and observed under a fluorescence microscope (Zeiss, Germany) with the appropriate excitation wavelength filters.

Quantitative real-time polymerase chain reaction

Gene expression (SMAD1, SMAD2, and BMP15) was evaluated using the real-time PCR technique. GAPDH was assumed as the housekeeping gene. RNA extraction was carried out by a RNeasy Mini kit (Qiagen, Germany) and obtained RNAs were converted to cDNA using a cDNA synthesis kit (Qiagen, Germany). For each gene, the reaction mixture possessed 10 μl Master Mix SYBR Green (Biofact, Korea), 7 μl nuclease free water, 1 μl each of forward and reverse primers and 1 μl cDNA. Table 2 listed the sequences for each primer. Over 45 cycles of PCR were conducted by the Applied Biosystems™ 7500 Real-Time PCR System (95°C for 10 min, 95°C for 25s, 5°C for 50s and 60°C for 45s). In the end, the 2−ΔΔCT method was used for data analysis.

Table 2. Primer sequences

Assessment of fertility rate

The monogamous system was used for evaluation the live-birth. In this system, a pair of male and female mice from each group were kept together in a cage. A separate cage was used for each pair of mice. Mating could be established within 24 h by the formation of a waxy vaginal plug and pregnancy lasted for 19–21 days. Then, the live-birth rate was evaluated.

Statistical analysis

Statistical analysis was accomplished using Statistical and GraphPad Prism software. The differences between the groups were measured by the Welch two-sample unpaired t-test. Moreover, the statistical significance between more than two groups was investigated using the Kruskal–Wallis test. All obtained data were reported as mean ± SEM (P-value < 0.05).

Results

Expression of GDF-9 in ovaries

The GDF9 expression significantly decreased in the CYC group compared with the control group. Also, the expression level in the PRP-treated group was very low. Albeit in the CM+PRP group, GDF9 expression was lower than in the control group, it was considerably higher than in the CYC group and the PRP group (Figure 1).

Figure 1. Immunohistochemical analysis of mouse ovary tissue labelling of GDF 9, Positive staining on follicles of mouse ovary is observed (green florescent). The nuclei were counterstained and observed using 40,6-diamidino-2-phenylindole (DAPI) staining.

Gene expression assessment

The expression levels of SMAD1, SMAD2, and BMP15 in the CYC group were remarkably lower compared with the control group. In the CM+PRP group, the expression levels of the mentioned genes were higher than in the CM group and the PRP group; however, the differences between them were not significant. The expression levels of SMAD1, SMAD2, and BMP15 in the CM group were more than in the PRP and CYC groups (Figure 2).

Figure 2. mRNA expression levels of SMAD1, SMAD2 and BMP15 in experimental groups.

Live birth rate evaluation

In CYC, PRP, and CM groups, fertility and live birth were not observed. But, in the CM+PRP and control groups, live birth was observed (Figures 3 and 4).

Figure 3. Number of live births in the experimental groups.

Figure 4. Treatment with conditioned medium and PRP in infertile mice resulted in successful fertility and live birth

Discussion

The mixture of PRP and CM was more effective than other treatment groups in mice treated with cyclophosphamide. Also, live birth in the mixed group of PRP and CM was observed. The abundance of PRP-rich therapeutic molecules is beneficial for cell proliferation and injury repair (Atashi et al., Reference Atashi, Jaconi, Pittet-Cuénod and Modarressi2015). Plus, the creation of PRP-containing scaffolds for stem cell transplantation can dramatically promote the therapeutic efficacy of mesenchymal stem cell (MSC) (Chang et al., Reference Chang, Li, Chen, Wei, Yang, Shi and Liang2015; Wu et al., Reference Wu, Zhou, Liu, Zhang, Xiong, Lv, Liu, Wang, Du, Zhang and Liu2017). Recent documents have recommended that the exploitation of PRP to improve the expression of adhesion molecules could be useful for the treatment of the endometrium (Colombo et al., Reference Colombo, Fanton, Sosa, Criado Scholz, Lotti, Aragona and Lotti2017; Liu et al., Reference Liu, Yuan, Fernandes, Dziak, Ionita, Li and Yang2017). Several published articles have shown that PRP provides an inhibitory effect on mating endometritis (Reghini et al., Reference Reghini, Ramires Neto, Segabinazzi, Castro Chaves, Dell’Aqua, Bussiere, Dell’Aqua, Papa and Alvarenga2016; Segabinazzi et al., Reference Segabinazzi, Friso, Correal, Crespilho, Dell’Aqua, Miró, Papa and Alvarenga2017).

Conversely, PRP decreased IL-6 expression in menstrual blood-derived stromal cells (MenSCs) in vitro over a short time (Zhang et al., Reference Zhang, Li, Yuan and Tan2018). WNT/ β-catenin signalling has a role in adjusting the function of endometrial stem cells during menopause. Specific cytokines during menopause can elevate endometrial MSC (eMSC) proliferation. Understanding the mechanism of eMSC regulation may help in the treatment of endometrial proliferation disorders, such as Asherman’s syndrome (Xu et al., Reference Xu, Chan, Li, Ng and Yeung2020).

The results of the current work implicated that the process of inhibiting inflammation was very weak in the PRP group. PRP alone may not be effective for treating severe intrauterine adhesions; although it can dramatically regenerate the endometrium. Kim et al. (Reference Kim, Park, Paek, Lee, Song and Lyu2020) studied the human PRP injection efficacy for endometrial regeneration in a mouse model of AS damage and showed that human PRP can ameliorate endometrial morphology and diminish fibrosis degree in an AS mouse model. In addition, human PRP treatment was associated with more implantation sites (IS) and live births (Kim et al., Reference Kim, Park, Paek, Lee, Song and Lyu2020).

The results of the study by Kim and colleagues (2020) were consistent with the present study. Human PRP-based therapy may be a useful approach for improving damaged endometrial regeneration and for fertility and pregnancy rates.

In a study, Pantos et al. (Reference Pantos, Simopoulou, Pantou, Rapani, Tsioulou, Nitsos, Syrkos, Pappas, Koutsilieris and Sfakianoudis2019) evaluated postmenopausal women and premature menopause after treatment with PRP. They showed that menstrual repair after harnessing autologous ovarian PRP remedy, as well as enhancement of hormonal characteristics, reduction in follicle-stimulating hormone (FSH) levels and there was a simultaneous elevation in anti-mullerian hormone (AMH) levels. In addition, patients were reported to have normal pregnancies within 2–6 months after PRP treatment (Pantos et al., Reference Pantos, Simopoulou, Pantou, Rapani, Tsioulou, Nitsos, Syrkos, Pappas, Koutsilieris and Sfakianoudis2019).

It has been offered that randomized controlled trials (RCTs) can also be helpful in monitoring the effectiveness of PRP. This may be a good clue to the successful therapy for a particular group of patients who have considered reproductive treatment methods after menopause.

Zhang et al. (Reference Zhang, Li, Yuan and Tan2019) studied the curative effect of transplantation of MenSCs in combination with PRP in a rat model of intrauterine adhesion (IUA) and MenSCs mechanisms in endometrial regeneration. They showed a significantly improved endometrial proliferation on days 9 and 18 after MenSC transplantation treatment. They also reported an angiogenesis and morphological improvement and reduction of inflammation and collagen fibrosis in the uterus. Also, MenSCs had lesion chemotaxis and were observed around the endometrial glands. Human secretory protein gene expression of TSP-1, SDF-1, and IGF-1 were perceived in uterus. The three treatments promoted fertility rate in rats with IUA. Furthermore, the expression of genes of cell proliferation and growth, and other biological occurrences were induced in the MenSC transplant group. The signal pathway was altered and Gdf5, Wnt5a, and CTGF were significantly modulated in the treatment groups. PRP potentiated these parameters by exerting synergistic influences. MenSCs could effectively ameliorate the uterus and significantly accelerate the healing rate of endometrial damage and enhance fertility regeneration in IUA rats (Zhang et al., Reference Zhang, Li, Yuan and Tan2019).

Genes for BMP15 and GDF9, secreted by the oocytes, can regulate granulosa cell differentiation and proliferation, primordial follicle recruitment, and steroid synthesis, and increase preantral follicle growth (Fenwick et al., Reference Fenwick, Mora, Mansour, Baithun, Franks and Hardy2013).

Previous studies have reported that BMP15 in the follicular fluid of mature follicles might be synthesized by oocytes from mature follicles and regulate the expansion of cumulus cell (Yoshino et al., Reference Yoshino, McMahon, Sharma and Shimasaki2006; Sun et al., Reference Sun, Lei, Cheng, Jin, Zu, Shan, Wang, Zhang and Liu2010). Moreover, evidence suggested that GDF9 was found in the primary follicle onwards (Aaltonen et al., Reference Aaltonen, Laitinen, Vuojolainen, Jaatinen, Horelli-Kuitunen, Seppa, Louhio, Tuuri, Sjoberg, Butzow and Hovatta1999). Our study demonstrated that the levels of BMP15 and GDF9 gene expression in the PRP+CM group was increased in comparison with the CYC, PRP, and CM groups.

In summary, the results of this study showed that fertility decreased in people undergoing chemotherapy with cyclophosphamide. Treatment with CM+PRP can stimulate the growth of ovarian follicles.

Acknowledgements

This work was supported by the Department of anatomy, Shiraz University of medical sciences (grant no. 16689). This article was extracted from PhD thesis written by Neda Taghizabet. This study was supported by the Shahid Beheshti University of Medical Sciences and Shiraz University of medical sciences.

Conflict of interest

The authors have no conflict of interest to declare.

References

Aaltonen, J., Laitinen, M. P., Vuojolainen, K., Jaatinen, R., Horelli-Kuitunen, N., Seppa, L., Louhio, H., Tuuri, T., Sjoberg, J., Butzow, R. and Hovatta, O. (1999). Human growth differentiation factor 9 (GDF-9) and its novel homolog GDF-9B are expressed in oocytes during early folliculogenesis. Journal of Clinical Endocrinology and Metabolism, 84(8), 27442750. doi: 10.1210/jc.84.8.2744 Google ScholarPubMed
Ahmadian, S., Sheshpari, S., Pazhang, M., Bedate, A. M., Beheshti, R., Abbasi, M. M., Nouri, M., Rahbarghazi, R. and Mahdipour, M. (2020). Intra-ovarian injection of platelet-rich plasma into ovarian tissue promoted rejuvenation in the rat model of premature ovarian insufficiency and restored ovulation rate via angiogenesis modulation. Reproductive Biology and Endocrinology: RB&E, 18(1), 78. doi: 10.1186/s12958-020-00638-4 CrossRefGoogle ScholarPubMed
Atashi, F., Jaconi, M. E., Pittet-Cuénod, B. and Modarressi, A. (2015) Autologous platelet-rich plasma: A biological supplement to enhance adipose-derived mesenchymal stem cell expansion. Tissue Engineering. Part C, Methods, 21(3), 253262. doi: 10.1089/ten.TEC.2014.0206 CrossRefGoogle ScholarPubMed
Aflatoonian, A., Lotfi, M., Saeed, L. and Tabibnejad, N. (2021) Effects of Intraovarian Injection of Autologous Platelet-Rich Plasma on Ovarian Rejuvenation in Poor Responders and Women with Primary Ovarian Insufficiency. Reproductive Sciences. 28:20502059. doi. 10.1007/s43032-021-00483-9 CrossRefGoogle ScholarPubMed
Chang, Y., Li, J., Chen, Y., Wei, L., Yang, X., Shi, Y. and Liang, X. (2015). Autologous platelet-rich plasma promotes endometrial growth and improves pregnancy outcome during in vitro fertilization. International Journal of Clinical and Experimental Medicine, 8(1), 12861290. doi: 10.1016/4358582 Google ScholarPubMed
Chen, Y., Zhuang, Y., Chen, X. and Huang, L. (2011). Effect of human endometrial stromal cell-derived conditioned medium on uterine natural killer (uNK) cells’ proliferation and cytotoxicity. American Journal of Reproductive Immunology, 65(6), 589596. doi: 10.1111/j.1600-0897.2010.00955.x CrossRefGoogle ScholarPubMed
Colombo, G., Fanton, V., Sosa, D., Criado Scholz, E., Lotti, J., Aragona, S. E. and Lotti, T. (2017). Use of platelet rich plasma in human infertility. Journal of Biological Regulators and Homeostatic Agents, 31(2), 179182.Google ScholarPubMed
Dehghani, F., Aboutalebi, H., Esmaeilpour, T., Panjehshahin, M.R. and Bordbar, H. (2018). Effect of platelet-rich plasma (PRP) on ovarian structures in cyclophosphamide-induced ovarian failure in female rats: a stereological study. Toxicology Mechanisms and Methods, 28(9), 653659. doi: 10.1080/15376516.2018.1491662 CrossRefGoogle ScholarPubMed
Dhillon, R. S., Schwarz, E. M. and Maloney, M. D. (2012). Platelet-rich plasma therapy – Future or trend? Arthritis Research and Therapy, 14(4), 219. doi: 10.1186/ar3914 CrossRefGoogle ScholarPubMed
Fathi, R., Valojerdi, M. R., Ebrahimi, B., Eivazkhani, F., Akbarpour, M., Tahaei, L. S. et al. (2017). Fertili preservation in cancer patients: in vivo and in vitro options. Cell Journal (Yakhteh), 19(2), 173185. doi: 10.22074/cellj.2016.4880 Google Scholar
Fenwick, M. A., Mora, J. M., Mansour, Y. T., Baithun, C., Franks, S. and Hardy, K. (2013). Investigations of TGF-β signaling in preantral follicles of female mice reveal differential roles for bone morphogenetic protein 15. Endocrinology 154(9), 34233436. doi: 10.1210/en.2012-2251 CrossRefGoogle ScholarPubMed
Hosseini, L., Shirazi, A., Naderi, M. M., Shams-Esfandabadi, N., Borjian Boroujeni, S., Sarvari, A., Sadeghnia, S., Behzadi, B. and Akhondi, M. M. (2017). Platelet-rich plasma promotes the development of isolated human primordial and primary follicles to the preantral stage. Reproductive Biomedicine Online, 35(4), 343350. doi: 10.1016/j.rbmo.2017.04.007 CrossRefGoogle Scholar
Jiang, Y., Zhao, J., Qi, H. J., Li, X. L., Zhang, S. R., Song, D. W., Yu, C. Y. and Gao, J. G. (2013). Accelerated ovarian aging in mice by treatment of busulfan and cyclophosphamide. Journal of Zhejiang University. Science. B, 14(4), 318324. doi: 10.1631/jzus.B1200181 CrossRefGoogle ScholarPubMed
Kim, J. H., Park, M., Paek, J. Y., Lee, W. S., Song, H. and Lyu, S. W. (2020). Intrauterine infusion of human platelet-rich plasma improves endometrial regeneration and pregnancy outcomes in a murine model of Asherman’s syndrome. Frontiers in Physiology, 11(3), 105. doi: 10.3389/fphys.2020.00105 CrossRefGoogle Scholar
Lin, J. X., Jia, Y. D. and Zhang, C. Q. (2011). Effect of epidermal growth factor on follicle-stimulating hormone-induced proliferation of granulosa cells from chicken prehierarchical follicles. Journal of Zhejiang University. Science. B, 12(11), 875883. doi: 10.1631/jzus.B1100023 CrossRefGoogle ScholarPubMed
Liu, Z., Yuan, X., Fernandes, G., Dziak, R., Ionita, C., Li, C. and Yang, S. (2017). The combination of nano-calcium sulfate/platelet rich plasma gel scaffold with BMP2 gene-modified mesenchymal stem cells promotes bone regeneration in rat critical-sized calvarial defects. Stem Cell Research and Therapy, 8(1), 19. doi: 10.1186/s13287-017-0574-6 CrossRefGoogle ScholarPubMed
Malekshah, A. K., Moghaddam, A. E. and Daraka, S. M. (2006). Comparison of conditioned medium and direct co-culture of human granulosa cells on mouse embryo development. Indian Journal of Experimental Biology, 44(3), 189192. doi:/handle/123456789/6373Google ScholarPubMed
Meirow, D., Assad, G., Dor, J. and Rabinovici, J. (2004). The GnRH antagonist cetrorelix reduces cyclophosphamide-induced ovarian follicular destruction in mice. Human Reproduction, 19(6), 12941299. doi: 10.1093/humrep/deh257 CrossRefGoogle ScholarPubMed
Nagashima, T., Kim, J., Li, Q., Lydon, J. P., DeMayo, F. J., Lyons, K. M. and Matzuk, M. M. (2011). Connective tissue growth factor is required for normal follicle development and ovulation. Molecular Endocrinology, 25(10), 17401759. doi: 10.1210/me.2011-1045 CrossRefGoogle ScholarPubMed
Okeke, T., Anyaehie, U. and Ezenyeaku, C. (2013). Premature menopause. Annals of Medical and Health Sciences Research, 3(1), 9095. doi: 10.4103/2141–9248.109458, doi: 10.4103/2141-9248.109458 CrossRefGoogle ScholarPubMed
Pantos, K., Simopoulou, M., Pantou, A., Rapani, A., Tsioulou, P., Nitsos, N., Syrkos, S., Pappas, A., Koutsilieris, M. and Sfakianoudis, K. (2019). A case series on natural conceptions resulting in ongoing pregnancies in menopausal and prematurely menopausal women following platelet-rich plasma treatment. Cell Transplantation, 28(9–10), 13331340. doi: 10.1177/0963689719859539 CrossRefGoogle ScholarPubMed
Pouya, S., Heidari, M., Baghaei, K., Asadzadeh Aghdaei, H. A., Moradi, A., Namaki, S., Zali, M. R. and Hashemi, S. M. (2018). Study the effects of mesenchymal stem cell conditioned medium injection in mouse model of acute colitis. International Immunopharmacology, 54, 8694. doi: 10.1016/j.intimp.2017.11.001 CrossRefGoogle Scholar
Reghini, M. F., Ramires Neto, C., Segabinazzi, L. G., Castro Chaves, M. M., Dell’Aqua, Cde P., Bussiere, M. C., Dell’Aqua, J. A., Papa, F. O. and Alvarenga, M. A. (2016). Inflammatory response in chronic degenerative endometritis mares treated with platelet-rich plasma. Theriogenology, 86(2), 516522. doi: 10.1016/j.theriogenology.2016.01.029 CrossRefGoogle ScholarPubMed
Segabinazzi, L. G., Friso, A. M., Correal, S. B., Crespilho, A. M., Dell’AquaJr, J. A., Miró, J., Papa, F. O. and Alvarenga, M. A. (2017). Uterine clinical findings, fertility rate, leucocyte migration, and COX-2 protein levels in the endometrial tissue of susceptible mares treated with platelet-rich plasma before and after AI. Theriogenology, 104, 120126.CrossRefGoogle ScholarPubMed
Srivastava, A., Sengupta, J., Kriplani, A., Roy, K. K. and Ghosh, D. (2013). Profiles of cytokines secreted by isolated human endometrial cells under the influence of chorionic gonadotropin during the window of embryo implantation. Reproductive Biology and Endocrinology, 11(2), 116. doi: 10.1186/1477–7827–11–116 CrossRefGoogle ScholarPubMed
Sugiura, K., Su, Y. Q., Li, Q., Wigglesworth, K., Matzuk, M. M. and Eppig, J. J. (2009). Fibroblast growth factors and epidermal growth factor cooperate with oocyte-derived members of the TGFbeta superfamily to regulate Spry2 mRNA levels in mouse cumulus cells. Biology of Reproduction, 81(5), 833841. doi: 10.1095/biolreprod.109.078485 CrossRefGoogle ScholarPubMed
Sun, R. Z., Lei, L., Cheng, L., Jin, Z. F., Zu, S. J., Shan, Z. Y., Wang, Z. D., Zhang, J. X. and Liu, Z. H. (2010). Expression of GDF-9, BMP-15 and their receptors in mammalian ovary follicles. Journal of Molecular Histology, 41(6), 325332. doi: 10.1007/s10735-010-9294-2 CrossRefGoogle ScholarPubMed
Takehara, Y., Yabuuchi, A., Ezoe, K., Kuroda, T., Yamadera, R., Sano, C., Murata, N., Aida, T., Nakama, K., Aono, F., Aoyama, N., Kato, K. and Kato, O. (2013). The restorative effects of adipose-derived mesenchymal stem cells on damaged ovarian function. Laboratory Investigation; a Journal of Technical Methods and Pathology, 93(2), 181193. doi: 10.1038/labinvest.2012.167 CrossRefGoogle ScholarPubMed
Turner, R. M. (2003). Tales from the tail: What do we really know about sperm motility? Journal of Andrology, 24(6), 790803. doi: 10.1002/j.1939-4640.2003.tb03123.x CrossRefGoogle ScholarPubMed
Van Der Voort, D. J., van Der Weijer, P. H. and Barentsen, R. (2003). Early menopause: Increased fracture risk at older age. Osteoporosis International, 14(6), 525530. doi: 10.1007/s00198-003-1408-1 CrossRefGoogle ScholarPubMed
Wu, J., Zhou, J., Liu, C., Zhang, J., Xiong, W., Lv, Y., Liu, R., Wang, R., Du, Z., Zhang, G. and Liu, Q. (2017). A prospective study comparing platelet-rich plasma and local anesthetic (LA)/corticosteroid in intra-articular injection for the treatment of lumbar facet joint syndrome. Pain Practice, 17(7), 914924, doi: 10.1111/papr.12544 CrossRefGoogle ScholarPubMed
Xu, J., Oakley, J. and McGee, E. A. (2002). Stage-specific expression of Smad2 and Smad3 during folliculogenesis. Biology of Reproduction, 66(6), 15711578. doi: 10.1095/biolreprod66.6.1571 CrossRefGoogle ScholarPubMed
Xu, S., Chan, R. W. S., Li, T., Ng, E. H. Y. and Yeung, W. S. B. (2020). Understanding the regulatory mechanisms of endometrial cells on activities of endometrial mesenchymal stem-like cells during menstruation. Stem Cell Research and Therapy, 11(1), 239. doi: 10.1186/s13287-020-01750-3 CrossRefGoogle ScholarPubMed
Yoshino, O., McMahon, H. E., Sharma, S. and Shimasaki, S. (2006). A unique preovulatory expression pattern plays a key role in the physiological functions of BMP-15 in the mouse. Proceedings of the National Academy of Sciences of the United States of America, 103(28), 1067810683. doi: 10.1073/pnas.0600507103 CrossRefGoogle Scholar
Zhang, S., Li, P., Yuan, Z. and Tan, J. (2018). Effects of platelet-rich plasma on the activity of human menstrual blood-derived stromal cells in vitro . Stem Cell Research and Therapy, 9(1), 48. doi: 10.1186/s13287-018-0795-3 CrossRefGoogle ScholarPubMed
Zhang, S., Li, P., Yuan, Z. and Tan, J. (2019). Platelet-rich plasma improves therapeutic effects of menstrual blood-derived stromal cells in rat model of intrauterine adhesion. Stem Cell Research and Therapy, 10(1), 61. doi: 10.1186/s13287-019-1155-7 CrossRefGoogle ScholarPubMed
Figure 0

Table 1. Experimental groups

Figure 1

Table 2. Primer sequences

Figure 2

Figure 1. Immunohistochemical analysis of mouse ovary tissue labelling of GDF 9, Positive staining on follicles of mouse ovary is observed (green florescent). The nuclei were counterstained and observed using 40,6-diamidino-2-phenylindole (DAPI) staining.

Figure 3

Figure 2. mRNA expression levels of SMAD1, SMAD2 and BMP15 in experimental groups.

Figure 4

Figure 3. Number of live births in the experimental groups.

Figure 5

Figure 4. Treatment with conditioned medium and PRP in infertile mice resulted in successful fertility and live birth