Isolation and Enrichment of Mouse Female Germ Line Stem Cells

Objective: The existence of female germ-line stem cells (FGSCs) has been the subject of a wide range of recent studies. Successful isolation and culture of FGSCs could facilitate studies on regenerative medicine and infertility treatments in the near future. Our aim in the present study was evaluation of the most commonly used techniques in enrichment of FGSCs and in establishment of the best procedure. Materials and Methods: In this experimental study, after digesting neonate ovary from C57BI/6 mice, we performed 2 different isolation experiments: magnetic activated cell sorting (MACS) and pre-plating. MACS was applied using two different antibodies against mouse vasa homolog (MVH) and stage-specific embryonic antigen-1 (SSEA1) markers. After the cells were passaged and proliferated in vitro, colony-forming cells were characterized using reverse transcription-polymerase chain reaction (RT-PCR) (for analysis of expression of Oct4, Nanog, C-kit, Fragilis, Mvh, Dazl, Scp3 and Zp3), alkaline phosphatase (AP) activity test and imnnunocytochemistry. Results: Data showed that colonies can be seen more frequently in pre-plating technique than that in MACS. Using the SSEA1 antibody with MACS, 1.98 +/- 0.49% (Mean +/- SDV) positive cells were yield as compared to the total cells sorted. The colonies formed after pre-plating expressed pluripotency and germ stem cell markers (Oct4, Nanog, C-kit, Fragilis, Mvh and Dazl) whereas did not express Zp3 and Scp3 at the mRNA level. Immunocytochemistry in these colonies further confirmed the presence of OCT4 and MVH proteins, and AP activity measured by AP-kit showed positive reaction. Conclusion: We established a simple and an efficient pre-plating technique to culture and to enrich FGSCs from neonatal mouse ovaries

Effect of garlic (Allium sativum) on male fertility: a systematic review

Introduction: Fertility in men mainly depends on the number, quality, motility, and morphology of the sperms, and disruption of each of these factors leads to infertility. A large number of couples suffer from infertility problems. Among the various therapies, medicinal herbs are used in many countries to treat male infertility. Current systematic review was conducted to study the effects of garlic on male fertility. Methods: The information of this systematic review was collected by searching the key words: treatment, fertility, infertility, male, herbal medicine, garlic, Allium sativum, medicinal plant, sperm, sex hormones, testis and spermatogenesis in international databases such as: Web of Science (ISI), PubMed, Scopus and Embase until March 2018. This study was conducted in accordance with the PRISMA statement for systematic reviews and meta- analysis. and the SYRCLE risk of bias tool was used for qualitative assessment. Results: A total of 18 experimental studies were included in the study. Thirteen studies evaluated garlic and 5 studies compared garlic effect with adriamycin, titanium dioxide, furan, vitamin E, N-acetylcysteine and cadmium. All studies were conducted in in vivo condition. The results of the studies indicated the potential effect of garlic on enhancing fertility and spermatogenesis, increasing the level of testosterone and improving the testicular structure. Conclusion: Garlic can increase fertility probably due to its antioxidant properties. However, more clinical trials are recommended

Isolation and localization of cells expressing Sca-1 in the Adult Mouse Ovary: An evidence for presence of Mesenchymal Stem cells

Objective: Recently growing evident declared that ‘neo-oogenesis’ continues in mature female life span and simultaneously another studies confirmed the presence of spermatogonial stem cells (SSCs). Even though there is agreement between scientist about SSCs population in male gender but on the other side ovarian stem cells have received raising challenges regarding the existence in the surface epithelium of ovary. Mesenchymal stem cells (MSCs) are the most applicable source of stem cells and the common marker of MSCs is Sca-1 so the purpose of this study was to clarify the incidence of stem cells in the surface epithelium of ovary Methods: forty C57BL6 mice were sacrificed and the ovary carefully excised from its surrounding fat tissue, after mechanical and enzymatic digestion cells were stained with Sca1 to estimate the incidence of positive stem cells (SCs) population fluorescence activated cell sorting (FACS). Part of digested cells used for RT-PCR, also histological section prepared for immunohistochemical (IHC) staining of Sca-1 in ovarian surface epithelium (OSE) FACS. Results: The gene expression of Sca-1 was confirmed in the ovarian tissue. As well, localization of Sca-1 positive cells was detected in the germinal layer of ovary and epithelial granular layer of primordial follicles. Moreover, we successfully could isolated the Sca-1positive cells through Conclusion: The present work findings confirmed an inclusive stem cell population in the ovary which can be a strong evident for regeneration of ovarian tissue in either purpose of ovulation scar and neo-oogenesis

Attitude of Medical Students toward Third-party Reproductive Techniques

Background: Pregnancy with the help of a third party, including the use of sperm, oocyte, embryo, and uterus, can be considered as an option for some infertile couples. Due to the important role of health professionals in infertility treatments, their attitudes are of particular importance in the acceptance or rejection of fertility suggestions involving the help of a third party. This study aimed to determine the attitudes of medical students at medical universities in Tehran toward third-party reproduction. Methods: This descriptive cross-sectional study was carried out at the Medical University of Tehran in 2018. Medical students (n=187) filled out the questionnaire, which consisted of two parts: the demographic characteristics of the research subjects and the questionnaire consisted of 76 questions about attitudes toward third-party reproduction. The content and face validity of the questionnaire were determined, and test-retest reliability of the questionnaire was established (0.89). Results: According to gender, participants’ attitudes toward childbearing, the importance of genetic dependency between parents and children, law issues, anonymity in donation programs, parental affection, the importance of the recipient's and donor's characteristics, surrogacy, gamete, and embryo donation were all statistically non-significant (Pvalue>0.05). According to age and also to year of entering the university, participants’ attitude only toward childbearing was statistically significant (Pvalue= 0.018 and 0.01, respectively). Conclusion: Since medical school students may set on the road to a specialty associated with infertility and its ramifications, it’s better to educate our soon-to-be health system professionals on all necessary aspects of infertility and third-party reproduction. Key Words: Attitude; Donor Conception; Infertility; Medical Students; Assisted Reproductive Technique

Attitude of Medical Students toward Third-party Reproductive Techniques

Background: Pregnancy with the help of a third party, including the use of sperm, oocyte, embryo, and uterus, can be considered as an option for some infertile couples. Due to the important role of health professionals in infertility treatments, their attitudes are of particular importance in the acceptance or rejection of fertility suggestions involving the help of a third party. This study aimed to determine the attitudes of medical students at medical universities in Tehran toward third-party reproduction. Methods: This descriptive cross-sectional study was carried out at the Medical University of Tehran in 2018. Medical students (n=187) filled out the questionnaire, which consisted of two parts: the demographic characteristics of the research subjects and the questionnaire consisted of 76 questions about attitudes toward third-party reproduction. The content and face validity of the questionnaire were determined, and test-retest reliability of the questionnaire was established (0.89). Results: According to gender, participants’ attitudes toward childbearing, the importance of genetic dependency between parents and children, law issues, anonymity in donation programs, parental affection, the importance of the recipient's and donor's characteristics, surrogacy, gamete, and embryo donation were all statistically non-significant (Pvalue>0.05). According to age and also to year of entering the university, participants’ attitude only toward childbearing was statistically significant (Pvalue= 0.018 and 0.01, respectively). Conclusion: Since medical school students may set on the road to a specialty associated with infertility and its ramifications, it’s better to educate our soon-to-be health system professionals on all necessary aspects of infertility and third-party reproduction. Key Words: Attitude; Donor Conception; Infertility; Medical Students; Assisted Reproductive Technique

Astaxanthin ameliorates inflammation, oxidative stress, and reproductive outcomes in endometriosis patients undergoing assisted reproduction: A randomized, triple-blind placebo-controlled clinical trial

PurposeIn a randomized, triple-blind, placebo-controlled clinical trial (RCT) including 50 infertile women with endometriosis candidate for assisted reproductive techniques (ART), we studied the effect of Astaxanthin (AST) on pro-inflammatory cytokines, oxidative stress (OS) markers, and early pregnancy outcomes.MethodsBefore and after 12 weeks of AST treatment (6 mg per day), blood serum and follicular fluid (FF) samples were collected from 50 infertile women with endometriosis stage III/IV undergoing ART. Pro-inflammatory cytokines (IL-1β, IL-6, and TNF-α) and OS markers (malondialdehyde [MDA], superoxide dismutase [SOD], catalase [CAT], and total antioxidant capacity [TAC]) were measured in the serum and FF. ART outcomes were also compared between the groups.ResultsIncreased serum levels of TAC (398.661 ± 57.686 vs. 364.746 ± 51.569; P = 0.004) and SOD (13.458 ± 7.276 vs. 9.040 ± 5.155; P = 0.010) were observed after AST therapy in the treatment group. Furthermore, serum MDA (14.619 ± 2.505 vs. 15.939 ± 1.512; P = 0.031) decreased significantly following antioxidant treatment. In addition, significantly lower serum levels of IL-1β (4.515 ± 0.907 vs. 6.8760 ± 0.8478; P = 0.000), IL-6 (5.516 ± 0.646 vs. 5.0543 ± 0.709; P = 0.024) and TNF-α (2.520 ± 0.525 vs. 2.968 ± 0.548; P = 0.038) were observed after AST treatment. In addition, AST supplementation led to an improved number of oocytes retrieved (14.60 ± 7.79 vs. 9.84 ± 6.44; P = 0.043), number of mature (MII) oocytes (10.48 ± 6.665 vs. 6.72 ± 4.3; P = 0.041), and high-quality embryos (4.52 ± 2.41 vs. 2.72 ± 2.40; P = 0.024).ConclusionAST pretreatment can modulate inflammation and OS in endometriosis-induced infertile patients. ART outcomes also improved after 12 weeks of AST therapy. Our results suggest that AST can be a potential therapeutic target for infertile patients with endometriosis undergoing ART

Association of dietary total antioxidant capacity, alternative healthy eating index, and dietary inflammatory index with semen quality in men seeking infertility treatment

BackgroundSince the association between dietary quality scores and semen quality remains unclear, we carried out a hospital-based cross-sectional study to investigate the association of Dietary Total Antioxidant Capacity (dTAC), Dietary Inflammatory Index (DII), and Alternative Healthy Eating Index (AHEI) scores with semen quality in men seeking infertility treatment.MethodsThis study enrolled 210 men with unexplained or idiopathic infertility. Semen samples were collected and analyzed according to the WHO 2010 criteria. Dietary data was collected using a 168-item semi-quantitative food frequency questionnaire (FFQ) developed for Tehran Lipid and Glucose Study. Multivariable logistic regression models were used to estimate the relationship between dTAC, AHEI, and DII scores with abnormal semen in crude and adjusted models.ResultsThere were no significant differences across quartile categories of the dTAC, AHEI, and DII scores regarding semen parameters. There was a trend toward a significant direct association between DII and abnormal semen risk (p = 0.01). Infertile men in the highest quartile of DII had a 2.84 times higher risk of abnormal semen in the crude model (OR: 3.84; 95% CI: 1.64–8.95); such that remained after adjusting for several potential confounders. There was no significant association between dTAC or AHEI and the risk of abnormal semen in infertile men, either before or after adjusting for potential confounders. Total energy (p = 0.05), fat (p = 0.02), saturated fat (p = 0.02), mono-saturated fat (p = 0.009), Thiamine (Vitamin B1) (p = 0.02), Niacin (Vitamin B3) (p = 0.03), Calcium (p = 0.01), and Selenium (p = 0.01) were inversely associated with semen normality.DiscussionThe study suggests that certain dietary factors may affect semen quality, and the mechanisms underlying the observed associations are likely multifactorial, involving complex interactions between diet, oxidative stress, inflammation, and hormone levels. Further research is required to confirm the results, fully elucidate the mechanisms underlying the associations, and identify specific dietary interventions that may improve male fertility outcomes

Comparison of differentiation potential of male mouse adipose tissue and bone marrow derived-mesenchymal stem cells into germ cells

Background: Recent publications about differentiation of stem cells to germ cells have motivated researchers to make new approaches to infertility. In vitro production of germ cells improves understanding differentiation process of male and female germ cells. Due to the problem of using embryonic stem cells (ESC), it’s necessary the mentioned cells be replaced with some adult multi-potent stem cells in laboratories. Objective: The aim of this study was to obtain germ cells from appropriate source beyond ESC and compare differential potentials of adipocytes derived stem cells (ADMSCs) with bone marrow derived stem cells (BMMSCs). Materials and Methods: To find multi-potential entity, after providing purified ADMSCs and BMMSCs, differentiation to osteoblast and adipocyte was confirmed by using appropriate culture medium. To confirm mesenchymal lineage production superficial markers (expression of CD90 and CD44 and non-expression of CD45 and CD31) were investigated by flowcytometry. Then the cells were differentiated to germ cells in inductive medium containing retinoic acid for 7days. To evaluate germ cells characteristic markers [Dazl (Deleted in azoospermia-like), Mvh (Mouse vasa homolog gene), Stra8 (Stimulated by retinoic acid) and Scp3 (Synaptonemal complex protein 3)] flowcytometry, imunoflorescence and real time PCR were used. Results: Both types of cells were able to differentiate into osteoblast and adipocyte cells and presentation of stem cell superficial markers (CD90, CD44) and absence of endothelial and blood cell markers (CD31, CD45) were confirmative The flowcytometry, imunoflorescence and real time PCR results showed remarkable expression of germ cells characteristic markers (Mvh, Dazl, Stra8, and Scp3). Conclusion: It was found that although ADMSCs were attained easier and also cultured and differentiated rapidly, germ cell markers were expressed in BMMSCs significantly more than ADMSCs