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Analysis of Apoptosis in Cultured Human Vitrified Ovarian Tissue in the Presence of Leukemia Inhibitory Factor.

ABSTRACT: Background:For improving the human ovarian tissue culture, this study was designed to assess the incidence of apoptosis in this tissue following vitrification and in vitro culture in the presence of leukemia inhibitory factor (LIF) as an anti-apoptotic factor. Methods:After collecting the ovarian tissue samples they were divided into non-vitrified and vitrified groups and cultured for 14 days in the presence and absence of LIF then morphological, ultrastructural and steroidogenesis studies, TUNEL and caspase-3/7 assays, and apoptosis analysis by real time RT-PCR were done in all groups. The data were analyzed by independent t-tests and the real time RT-PCR results were compared by one-way ANOVA (p-values of <0.05 were considered significant). Results:No significant difference was observed between non-vitrified and vitrified groups in normality rate of follicles, the levels of hormones, TUNEL positive cells and caspase-3/7 activity. But in all LIF-treated groups, the levels of 17-? estradiol and progesterone were higher and TUNEL signals and caspase-3/7 activity were lower than non-LIF treated groups. The expression of Fas and FasL genes was higher in vitrified group in comparison with non-vitrified group but the expression of other genes was not significantly different. In LIF-treated groups, the expression of pro-apoptotic genes was significantly lower and the expression of anti-apoptotic genes was higher than non-LIF treated group. Conclusion:The vitrification of human ovarian tissue did not increase the incidence of apoptosis at the morphological and molecular levels during long term culture and LIF improves the survival and development of cultured follicles.

SUBMITTER: Abdollahi M

PROVIDER: S-EPMC6328982 | biostudies-literature | 2018 Oct-Dec

REPOSITORIES: biostudies-literature

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Analysis of Apoptosis in Cultured Human Vitrified Ovarian Tissue in the Presence of Leukemia Inhibitory Factor.

Abdollahi Maasoume M Salehnia Mojdeh M Salehpour Saghar S Pour-Beiranvand Shahram S

Journal of reproduction & infertility 20181001 4

<h4>Background</h4>For improving the human ovarian tissue culture, this study was designed to assess the incidence of apoptosis in this tissue following vitrification and <i>in vitro</i> culture in the presence of leukemia inhibitory factor (LIF) as an anti-apoptotic factor.<h4>Methods</h4>After collecting the ovarian tissue samples they were divided into non-vitrified and vitrified groups and cultured for 14 days in the presence and absence of LIF then morphological, ultrastructural and steroid ...[more]

PMID: 30746334

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Project description:ObjectiveAutograft transplantation of vitrified cortical ovarian tissue is an acceptable clinical technique for fertility preservation in women. Xenograft transplantation into animal models could be useful for evaluating the safety of human vitrified ovarian tissue. This study targeted to evaluate impact of vitrification on expression of the genes associated with folliculogenesis after xenograft transplantation of human vitrified ovarian tissue to γ-irradiated mice.Materials and methodsIn this experimental study, ovarian biopsies were gathered from six transsexual persons. The cortical section of ovarian biopsies was separated and chopped into small pieces. These pieces were randomly divided into vitrified and non-vitrified groups. In each group some pieces were considered as non-transplanted tissues and the others were transplanted to γ-irradiated female National Medical Research Institute (NMRI) mice. Before and after two weeks of xenograft transplantation, histological assessment and evaluation of the expression of folliculogenesisassociated genes (FIGLA, GDF-9, KL and FSHR) were performed in both vitrified and non-vitrified groups.ResultsPercentage of the normal follicles and expression of the all examined genes from transplanted and nontransplanted tissue were similar in both vitrified and non-vitrified groups (P>0.05). After transplantation, the normal follicle rate was significantly decreased and among the folliculogenesis-associated genes, expression of GDF-9 gene was significantly increased, rather than before transplantation in vitrified and non-vitrified tissues (P<0.05).ConclusionThe vitrification method using dimethyl solphoxide and ethylene glycol (EG) had no remarkable effect on the normal follicular rate and expression of folliculogenesis-associated genes after two weeks human ovarian tissue xenografting. In addition, transplantation process can cause a significant decrease in normal follicular rate and expression of GDF-9 gene.

Project description:Ovarian carcinoma-associated mesenchymal stem cells (CA-MSC) produce not only high levels of interleukin-6 (IL6) but also the related cytokine leukemia inhibitory factor (LIF). IL6-mediated activation of STAT3 is implicated as a critical therapeutic target for cancer therapy. Less is known about the role of LIF, which can similarly activate STAT3, in ovarian cancer. We therefore sought to evaluate the tumorigenic effects of CA-MSC paracrine LIF signaling and the redundancy of IL6 and LIF in activating ovarian cancer STAT3 mediated cancer growth. As expected, we found that both IL6 and LIF induce STAT3 phosphorylation in tumor cells. In addition, both IL6 and LIF increased the percentage of ALDH+ ovarian cancer stem-like cells (CSC). Supporting redundancy of function by the two cytokines, CA-MSC induced STAT3 phosphorylation and increased cancer cell "stemness". This effect was not inhibited by LIF or IL6 blocking antibodies alone, but was prevented by dual IL6/LIF blockade or JAK2 inhibition. Similarly, small hairpin RNA (shRNA)-mediated reduction of IL6 or LIF in CA-MSC partially decreased but could not completely abrograte the ability of CA-MSC to induce STAT3 phosphorylation and stemness. Importantly, the in vivo pro-tumorigenic effect of CA-MSC is abrogated by dual blockade with the JAK2 inhibitor ruxolitinib to a much greater extent than treatment with anti-IL6 or anti-LIF antibody alone. Ruxolitinib treatment also improves survival in the immunocompetent ovarian cancer mouse model system with ID8 tumor cells plus MSC. Ruxolitinib-treated tumors in both the immunocompromised and immunocompetent animal models demonstrate decreased phospho-STAT3, indicating on-target activity. In conclusion, CA-MSC activate ovarian cancer cell STAT3 signaling via IL6 and LIF and increase tumor cell stemness. This functional redundancy suggests that therapeutic targeting of a single cytokine may be less effective than strategies such as dual inhibitor therapy or targeting shared downstream factors of the JAK/STAT pathway.

Dataset Information

Analysis of Apoptosis in Cultured Human Vitrified Ovarian Tissue in the Presence of Leukemia Inhibitory Factor.

Publications

Analysis of Apoptosis in Cultured Human Vitrified Ovarian Tissue in the Presence of Leukemia Inhibitory Factor.

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