Function of human amniotic mesenchymal stem cell exosomes in repairing submandibular gland epithelial cells after radiation injury in SD rats

Zhang Ligang; Liu Tao; Yi Jie; Zhang Nini; Yao Li; Huang Guilin; Hu Xiaohua; Dai Min

doi:10.12307/2026.088

Abstract：BACKGROUND:Hypoxia preconditioning can increase extracellular vesicles,growth factors,anti-inflammatory and immunosuppressive factors in the parasecretory secretions of mesenchymal stem cells.Exosomes derived from human amniotic mesenchymal stem cells preconditioned with hypoxia are expected to play a better therapeutic role in tissue damage repair. OBJECTIVE:To observe the repair effect of exosomes from human amniotic mesenchymal stem cells preconditioned with hypoxia on radiation-induced submandibular gland epithelial cell damage. METHODS:The passage 3 human amniotic mesenchymal stem cells were divided into two groups:hypoxia and normoxia.They were pretreated with hypoxia(1%O2 by volume)and normoxia(20%O2 by volume)for 48 hours,respectively.Then,normoxic and hypoxic human amniotic mesenchymal stem cell exosomes were extracted from the culture supernatant of human amniotic mesenchymal stem cells by modified ultrahigh-speed centrifugation.The submandibular gland epithelial cells of SD newborn rats were divided into blank control group,radiation control group,normoxic drug group,and hypoxic drug group.The submandibular gland epithelial cells were irradiated with 5 Gy to establish the radiation-damaged submandibular gland epithelial cell model.On this basis,normoxic and hypoxic human amniotic mesenchymal stem cell exosomes were added to the normoxic drug group and the hypoxic drug group and co-cultured with the submandibular gland epithelial cells for 3 days.The proliferation activity of the submandibular gland epithelial cells of the rats in each group was detected by CCK-8 assay.The alpha amylase content in the supernatant of the submandibular gland epithelial cells of the rats in each group was determined by ELISA.RT-qPCR was used to detect the expression of aquaporin 5 mRNA in the submandibular gland epithelial cells of the rats in each group. RESULTS AND CONCLUSION:(1)The cell proliferation activity of the radiation control group was significantly decreased compared to the blank control group(P<0.05).The cell proliferation activity of the normoxic and hypoxic drug groups significantly increased on days 2 and 3 compared to the radiation control group(P<0.05).The cell proliferation activity of the hypoxic drug group was higher than that of the hypoxic drug group on the first day,and showed an increasing trend on the second and third days,but there was no statistical difference compared with the normoxic drug group(P>0.05).(2)The content of alpha amylase in the radiation control group was significantly reduced compared with the blank control group(P<0.05).The levels of alpha amylase in the normoxic and hypoxic drug groups were significantly higher than those in the radiation control group(P<0.05).Compared with the normoxic drug group,the hypoxic drug group showed an increasing trend in alpha amylase content,but there was no statistical difference(P>0.05).(3)The mRNA expression of aquaporin 5 in the radiation control group was significantly reduced compared to the blank control group(P<0.05).The mRNA expression of aquaporin 5 in the normoxic and hypoxic drug groups was significantly increased compared with the radiation control group(P<0.05).The mRNA expression of aquaporin 5 in the hypoxic drug group was slightly higher than that in the normoxic drug group,but there was no statistical difference(P>0.05).These results indicate that extracellular vesicles from human amniotic mesenchymal stem cells pre-treated with normoxic and hypoxic conditions have a repairing effect on submandibular gland epithelial cells damaged by radiation,but there is no significant advantage in hypoxic human amniotic mesenchymal stem cell exosomes.