In brief: During pregnancy and delivery, the myometrium was affected by hypoxia stress, which acts as a regulator of cell proliferation. The proliferation of uterine smooth muscle cells in pregnant mice was inhibited under hypoxia, which was related to the up-regulated autophagy through the mTOR pathway.
Abstract: Hypoxia is closely associated with physiological and pathological conditions in the human body, and the myometrium is affected by hypoxic stress during pregnancy and delivery. Autophagy is a catabolic pathway involved in the regulation of apoptosis, proliferation, and migration of a variety of cells, which can be activated under hypoxia. However, the mechanism and function of autophagy in uterine smooth muscle cells remained unclear. The aim of this study was to investigate the changes in autophagy in pregnant uterine smooth muscle cells (pUSMCs) under hypoxia and the effect of autophagy on myometrial cellscell proliferation during pregnancy. In this study, primary uterine smooth muscle cells were isolated from mice in late pregnancy and cultured under normoxic and hypoxic conditions, respectively. Western blotting and immunofluorescence were used to detect the expression levels of autophagy-related proteins LC3B, P62, mTOR, and p-mTOR under different culture conditions. Cell proliferation was assessed by CCK-8 assay. In addition, 3-methyladenine (3-MA) was used to inhibit autophagy in hypoxia-treated pUSMCs, and MHY1485 was used to activate mTOR. Studies have confirmed that under hypoxic conditions, autophagy is enhanced and cell proliferative viability is reduced in pUSMCs. The autophagy inhibitor 3-MA restored cell proliferation inhibited by hypoxia. Furthermore, hypoxia in pUSMCs led to a downregulation of p-mTOR/mTOR levels. The mTOR activator MHY1485 inhibited autophagy by preventing the binding of autophagosomes to lysosomes and reversed the hypoxia-induced inhibition of cell proliferation. Collectively, our results indicate that hypoxia upregulates autophagy through the mTOR pathway in pUSMCs, thereby inhibiting cell proliferation during pregnancy.