| Eospalax cansus(E.cansus)belongs to the Rodentia,Spalacidae,Myospalactinae,Eospalax,It is widely distributed in the Loess Plateau of Gansu,southern Ningxia,Shanxi,eastern Qinghai and other regions,where forests,farmLands,and barren slopes habit,is a typical subterranean rodent.Because they lived in underground oxygen deficient tunnels for many years,they evolved many similar characteristics.There was a significant difference in morphology and physiology from the ground mouse,which evolved into the unique hypoxia adaption mechanism of subterranean mice.Oxygen is a necessary condition for animal survival.Hypoxia can cause damage to the animal or lead to death.The brain is an organ with high oxygen consumption in the animal body and accounts for 20%-30%of the total oxygen consumption,and is highly sensitive to hypoxia.Under hypoxia,hypoxia-inducible transcription factor(HIF),which is an oxygen-dependent transcriptional activator,is produced.The activity of HIF is mainly caused by oxygen through the reg ? Lation of HIF-1?,which is necessary for growth dependent cell growth.In the hypoxic state,inhibition of HIF-1? expression can accelerate cell death.Mammalian target of rapamycin(mTOR)is an atypical serine/threonine protein kinase that plays an important role in cell cycle reg ?Lation,proliferation,cell growth,and differentiation.It can integrate extracell?Lar signals and phosphorylate downstream target proteins,such as S6K1 and 4E-BP1 to influence protein translation and gene transcription,there by participating in the reg?gLation of cell proliferation and growth.The mTOR signaling pathways is a major reg ? Lator of physiological activities such as growth,development,metabolism,immunity,and cancer.when externally stim ? gLated,the mTOR signaling pathway is activated,and TOR phosphorylates the downstream target protein 4E-BP1(binding of eukaryotic translation inititation factor 4E protein 1).The inhibition of EIF4E(eukearyotic translation initation factor 4E)by 4E-BP1was released,and EIF4E recruited the 40s ribosomal subunit to bind to the 5 'end of the mRNA to begin translation of the protein.Related studies have shown that cell in the hypoxia are affected by protein synthesis,and even cause cell apoptosis,affecting the normal physiological metabolism of the body.EKcansus lives in a hypoxia environment and still maintain normal physiologiy.We hypothesize that the mTOR signaling pathway plays an important role in the hypoxia stress of E.cansus.To this end,this study used E.cansus as a research object,use SD rats as controls,using real-time quantitative PCR and Westen Blot technology to detect expression of mRNA and protein in the brain of E.cansus and SD rate for HIF-1? and mTOR signaling pathways downstream,such as 4E-BPI,EIF4E at 21%,10.5%and 6.5%oxygen.The brain tissues of E.cansu and SD rate under three oxygen conditions were observe by HE staining and TUNEL.Morphological changes in cella and apoptosis.The above experiment res?Lts are as follows:1.The mRNA expression of HIF-1?,mTOR and its downstream factors,4E-BP1 and EIF4E mRNA in the brain of E.cansusAt 21%and 10.5%oxygen,the mRNA expression of HIF-1? in E.cansus was higher than SD rats,and was significantly higher than SD rats in 21%oxygen concentration.The expression of HIF-1? in SD rats was significantly lower than E.cansus in 21%and 10.5%oxygen.With the decrease of oxygen concentration,the expression of HIF-1? in SD rats showed an increasing trend,but there was no significant difference in different concentrations.The mRNA expression of mTOR in 21%,10.5%and 6.5%of the oxygen,the expression of mTOR in E.cansus was significantly higher than SD rats.the expression in E.cansus under 6.5%significantly higher than under the 21%and 10.5%.SD rats were significantly increased at 6.5%oxygen concentration,and there was no significant difference between 10.5%oxygen and 21%oxygen.4E-BP1 mRNA expression in E.cansus with the reducing of oxygen is reduced,and extremely significant differences between the three groups,the same oxygen concentration,the expression of 4E-BP1 in E.cansus and SD rats were extremely significant differences,under 21%and 10.5%oxygen significantly higher than SD rats,under 6.5%oxygen,the expression of 4E-BP1 in E.cansus is significantly lower than SD rats.The mRNA expression of EIF4E in the E.cansus decreased with the decrease of oxygen concentration,and the expression under 6.5%oxygen was significantly lower than 21%oxygen,and there was no significant differences under 10.5%oxygen.SD rats under three oxygen difference was no significant,under 21%,the expression of EIF4E in E.cansus is significantly higher than SD rats,10.5%oxygen higher than SD rats,under 6.5%oxygen,E.cansus is slightly lower than SD rats,but there was no significant difference.2.The ptotein expression of HIF-1?,4E-BP1,and EIF4E in E.cansus:In E.cansus,the expression of HIF-1? protein under 10.5%oxygen was significantly higher than 21%oxygen,and the change was not obvious at 6.5%oxygen,slightly higher than 21%oxygen concentration.With the decrease of oxygen concentration,the expression of HIF-1? in SD rats increased gradually,and the expression at 6.5%oxygen was significantly higher at 21%and 10.5%oxygen.At different oxygen concentrations,the expression of HIF-1? protein in SD rats was consistently lower than E.cansus,at 21%and 10.5%oxygen the expression of HIF-1?significantly higher than that SD rats.With the decrease of oxygen concentration,the expression of P-4EBP1 protein in E.cansus showed a decreasing trend,and the difference was significant.The expression of P-4EBP1 protein in SD rats showed an increasing trend,with a significant difference between the expression quantity of 6.5%oxygen and 21%and 10.5%.In the 21%oxygen,the expression in E.cansus was significantly higher than SD rats,and under the 10.5%oxygen was lower than SD rats,but the difference was no significant,and at 6.5%oxygen concentration was significantly lower than SD rats.As the oxygen concentration decreased,EIF4E protein expression in E.cansus has no obvious change.In SD rats,the expression under 6.5%oxygen compared with 21%and 10.5%oxygen is increases,at 10.5%is slightly less than 21%oxygen,but there was no significant difference.3.Under hypoxia stress,HE staining and apoptosis in E.cansus and SD rats.According to the res?Lts of HE staining,under oxygen concentration 6.5%,E.cansus and SD rats compared with 6.5%cells is small,just the cytoplasm shrinkage change cell volume,not inflammation or autophagy,remain cells of normal physiological function.SD rats had inflammatory infiltration at 6.5%oxygen,and the normal physiological function of the cells was obstructed.According to the res?Lts of TUNEL,it was found that the apoptosis of brain cells of rats and SD rats was not obvious under hypoxia stim?Lation.The research res?Lts indicate that hypoxia stim?Lus E.cansus HIF-1? expression,actived the mTOR signaling pathways,the mRNA expression of mTOR increase,reduce the E.cansus 4E-BP1 and EIF4E mRNA express,specification under 6.5%,mTOR in high expression of transcription,reduce 4E-BP1 protein phosphorylation,reduce the expression of transcription EIF4E,but EIF4E changes in protein expression level was not significant.HE staining showed that the brain cells of SD rats had inflammatory infiltration at the 6.5%oxygen,and such phenomenon no found in E.cansus.It was confirmed that the E.cansus had better hypoxic tolerance than SD rats.E.cansus have different reg?Latory mechanisms compared with SD rats.This is caused by long-term evolution.Protein translation is one of the most energy consumption process of cells,in order to adapt to low oxygen,E.cansus reduce the rate of protein synthesis in the body to reduce energy consumption,the available energy is used to maintain the balance of ion balance and osmotic pressure in the body,prevent cell swelling and necrosis associated with cell membrane depolarization,better protect E.cansus brain cells from hypoxia damage. |
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