The Role Of Regulating Mouse Erythropoiesis By Plateau Adaptation Related Mutant Gene Retsat^Q247R

Purposes:When mammals living on the plain are exposed to high altitude hypoxia environment,a series of altitude sickness will occur,such as dyspnea,hyper erythrocytosis,high altitude heart disease and so on.However,there are many mammals living on the plateau for generations,have been able to adapt to the plateau hypoxia environment after long-term evolution.Recently,according to the convergent evolution analysis of four plateau animals(yak,zokor,pika and Tibetan antelope),the mutation of Retsat gene encoding retinol saturase at the same amino acid site:Q247R was identified,which is speculated to be closely related to mammalian plateau adaptation.Pleanty of studies have reported that the number and morphology of the erythroblast of the four plateau animals,yak,zokor,pika,and Tibetan antelope,are significantly different in comparing with that of non-plateau animals.As a common muted gene that was identified in the plateau animals,we speculated that RetsatQ247R gene might possess its roles in the plateau hypoxia by regulating the production and function of erythroid.In addition,to further illustrate the roles of RetsatQ247R in the regulation of erythropoiesis,we also should explore its roles in the plain environment.Therefore,by combing use various environmental system,the present thesis was conducted to explore the roles of RetsatQ247R in mouse erythropoiesis,and to further illustrate its' function involved in the plateau hypoxia adaption.Methods:Firstly,by combining use the RNA-seq data and proteomics data that were published previously,we explore the expression of Retsat gene at mRNA and protein level in mouse erythroid at various developmental stages.Then,by testing blood routine,observing the morphology of the femur and spleen,and detecting the erythroid progenitor proliferation and terminal erythroid differentiation by flow cytometry,we compared the process of the specific biological events during the erythropoiesis in RetsatQ247R mice with that of the WT mice and further analyze the influence of RetsatQ247R on erythropoiesis.Results:1.Retsat was expressed in a stage specific manner during the process of mouse erythropoiesis,and the expression was dramatically upregulated in CFU-E progenitors.2.Only under hypoxic pressure(normal oxygen conditions in plain,Under normal oxygen conditions in plateau,Under the condition of 16%oxygen content in Kunming)there was no statistical difference between RetsatQ247R and WT mice in blood routine indexes,femur and spleen morphology,erythroid progenitor proliferation and terminal erythroid differentiation.3.Only after EPO stress(After EPO injection under normal oxygen conditions in plain),we did not observe statistical difference between RetsatQ247R and WT mice in blood routine indexes,femur and spleen morphology,terminal erythroid differentiation.4.Under the dual stimulation of hypoxia and injection of EPO,the spleen of RetsatQ247R mice was smaller than that of WT mice.Conclusion:Bioinformatics analysis revealed the stage specific expression of Retsat during mouse erythropoiesis that Retsat might play important roles in the regulation of mouse erythropoiesis,and RetsatQ247R might be involved in the high-altitude adaption by affecting erythropoiesis.There was no significant change of the erythropoiesis process in RetsatQ247R mice only under the stimulation of hypoxia or injection of EPO.In contrast,when the mice were treated with double stimulation by exposing in hypoxia and EPO stress,the compensatory hyperplasia of spleen in RetsatQ247R mice was significantly reduced.All these findings indicated that RetsatQ247R was involved in the regulation of hypoxia adaption.It can alleviate the pathological increase of spleen under the dual stimulation of hypoxia and injection of EPO to a certain extent.RetsatQ247R plays a regulatory role in stress hematopoiesis under hypoxia,but under what specific conditions it can effectively regulate stress hematopoiesis needs further research.In addition,based on the findings,we speculate that there may be insufficient regulation intensity under the single action of RetsatQ247R.Multiple mutations may be required for coordinated regulation in order to play an important role in altitude hypoxia adaptation by affecting erythroid development and function.In the follow-up study,themode and mechanism of synergy,need to be further explored.