Investigation of expression level of Klotho gene in the aging process

Investigation of Expression Level of Klotho Gene in the Aging Process Aging is the anatomical and physiological changes seen in living organisms over time. At the molecular level, genomic instability (especially DNA damage), telomere shortening and epigenetic changes are seen in aging, while stem cells depletion, increase in oxidative stress caused by deterioration of the function of mitochondria, deterioration of intercellular communication, increase in lysosomal enzyme activity in cells and cessation of cell proliferation are observed. The ovary is the reproductive endocrine organ that performs reproductive and hormonal functions in women. There is a natural decline in female fertility with aging. ROS play a role in many physiological and pathological activities in the ovary, from oocyte maturation to fertilization. Excessive increase in ROS leads to OS and causes ovarian aging. NRF2 is a transcription factor that protects cells from oxidative stress by regulating antioxidant and cytoprotective genes. It has been shown that increased NRF2 gene expression in ovarian cells triggers antioxidant genes and protects ovarian function. Klotho is an aging suppressor gene. Studies have shown that an increase in Klotho gene expression prolongs lifespan, and a decrease in Klotho gene expression accelerates the aging process. IL-1β is a potent proinflammatory cytokine and gene expression increases with aging. IL-6 is a multifunctional cytokine that regulates cell proliferation, survival and differentiation. IL-6 is involved in the formation of various diseases generally associated with inflammation, chronic immune diseases, neurological diseases and tissue aging, and gene expression increases with aging. The p53 protein is a transcription factor encoded by the TP53 gene, a tumor suppressor gene. The p53 gene, which controls the cell cycle process and programmed cell death (apoptosis), plays a very important role in determining the cell's response in various states of cellular stress, such as DNA damage, nutrient deficiency, and hypoxia, and p53 gene expression increases with aging. AKT is a serine-threonine kinase involved in cellular signaling pathways. The PI3K/AKT signaling pathway is extensively active in cells and functions in cell growth, proliferation, differentiation and apoptosis. mTOR is a serine-threonine kinase and is involved in many biological processes, including cell growth, cellular senescence, and lifespan. Epithelial ovarian stem cells are the main source of oocyte cell production in adults. OSE cells have stem cell properties. OSE cells play a role in the proliferation of epithelial cells, the production and regeneration of follicle, granulosa and theca cells. SOX2, OCT4 and NANOG are transcription factors required for self-renewal or pluripotency of undifferentiated embryonic stem cells. High levels of H2O2 cause oxidative stress in cells, leading to premature aging. In this project, the cells were aged in culture medium by applying H2O2 in the human ovarian epithelial cell line (IHOEC). The expressions of Klotho, NRF2, mTOR, p53, IL-6, IL-1β, OCT4, NANOG, SOX2 and AKT genes were examined by real-time PCR in HO treated cells. In this study, both Klotho and NRF2 gene expression were decreased during the natural aging process. In IHOEC cells, mTOR, p53, IL-6, IL-1β, OCT4, NANOG, SOX2, AKT gene expressions increased in aging induced by H2O2. The investigation of Klotho and NRF2 gene expression in the IHOEC cell line in this study is the first study for this cell line. In the context of this thesis, healthy ovarian surface epithelial cells: 1- Creation of two separate senescence cell models for two senescence types (a-senescence caused by stress, b-senescence formed by advanced passage) and verification of these two senescence models by SABG and SAHF trials, 2- It was aimed to investigate the expressions of antioxidant pathway (alpha-Klotho, NRF2), SASP markers (p53, IL6, IL1-beta), stem cell (Oct4, Nanog, Sox2) and survival (mTOR, AKT)-related genes in two different senescence models created by RT-qPCR method. Keywords: Age, Klotho gene expression, Ovum aging, NRF2, ROS