阿拉善双峰驼独特生理特征的调控机制研究

The camel is an new important model animal to study the high-salt metabolish and hyperglycemia. After a long drought desert environment of adaptive evolution, the camel formed the unique physiological and biochemical characteristics such as halophilic, high concentration of urine and hyperglycemia. Existing genome and transcriptome data section explains the salt metabolism, signification of hyperglycemia, but its specific regulatory mechanism is not clear. This study intends to treat the camels with different two methods. One group banned to drink water and the another group treated with salt stress. Then, we collect the kidneys, livers and small intestinal, which were the main tissues to take place the metabolish and transportation of salt and sugar, from the experiments and control camels. Through the lncRNA sequencing, miRNA sequencing，bioimformatics analysis and functional verification experiments, we may find the ncRNAs which participate in regulating the the metabolish and transportation of salt and sugar. The results maybe explaine the formation and regulatory mechanism of unique physical characteristics in Camel, which will supply the reference to disease of brain and cardiovascular system derived from hyperglycemia and high salt intake.

骆驼是研究高盐代谢和高血糖转运和代谢的新型重要模式动物。经过长期沙漠干旱环境下的适应性进化，骆驼形成了嗜盐、高浓缩尿和高血糖等独特生理生化特性。已有的基因组、转录组数据部分的解释了盐代谢、高糖存在的意义，但其具体调控机制不清楚，本研究拟通过禁水和盐胁迫实验处理，采集处理组和对照组盐、糖代谢和转运比较集中的组织肾脏、肝脏和小肠，经lncRNA测序、miRNA测序、生物信息学分析和功能验证试验，揭示骆驼中参与盐、糖转运和代谢的ncRNAs种类、功能和调节方式，解释骆驼嗜盐和高血糖等独特生理特征形成的原因和调控机制，为人类的高血糖以及由摄入高盐引起的心脑血管疾病的治疗提供参考。

骆驼是久居沙漠的大型哺乳动物，其作为动物抗逆性研究的良好生物资源拥有着极其重要的科研和应用价值。本试验对9峰雌性双峰驼随机分为盐胁迫组、禁水胁迫组和自由饮食组。经24天处理后分别采集各骆驼的肾皮质、肾髓质、肝脏和回肠进行了转录组测序，从中筛选出各组织差异表达的mRNA、miRNA和lncRNA。结合GO，KEGG通路富集和文献分析，进一步选出与盐、糖代谢和水代谢相关的基因作为候选基因，进行qRT-PCR验证，结果与测序结果的差异表达趋势一致。之后对候选ceRNA基因SLC14A1、LNC003834、miRNA-34a进行FISH和Luciferase检测，对候选抗氧化基因SLC6A1、PCBP2、PEX5L进行RNAi和ROS检测。综合结果，建立了骆驼抗旱耐盐调控模型。具体为：肾髓质通过ceRNA机制上调UTB表达来降低Na+重吸收量，同时上调GAT1、PEX5L和PCBP2表达来增强细胞的抗氧化能力，以达到耐高盐的目的；通过下调ATP柠檬酸合酶和HBB表达来降低细胞呼吸和新陈代谢，同时上调PCBP2表达来增强细胞的抗氧化能力，从而起到抗干旱的作用。肝脏实现耐高盐是通过上调TEN-1表达来提高细胞基底膜的渗透屏障作用，上调CDH11和PKP4的表达来增加细胞黏附并降低Na+进细胞所需的表面积，下调HBB表达来降低细胞呼吸和新陈代谢，下调SDH表达来降低糖异生并防止细胞内糖沉积；实现耐高血糖和抗干旱是通过下调UPP2和SDH表达来降低糖异生和糖原合成，上调PLIN2表达来促进脂滴积累和类乳脂球分泌出细胞，下调HBB表达来降低细胞呼吸和新陈代谢。回肠耐高盐是通过下调AQP5表达来防止因细胞外渗透压过高引起的失水，下调MUC6表达来降低经Na+/Ca2+交换体的粘蛋白分泌并防止大量Na+进细胞来完成的；耐高血糖和抗干旱的实现是通过下调AQP5的基因表达来防止细胞内水流失，下调MUC6的基因表达来降低经Na+/Ca2+交换体的粘蛋白分泌并防止Ca2+流失，其中Ca2+能促进脂肪酸合酶FAS表达和活性，上调线粒体肌酸激酶U型的基因表达来形成能量缓冲作用，从而减缓大量葡萄糖经钠依赖性葡萄糖协同转运蛋白进入细胞带来的糖负荷。该调控模型为揭示骆驼适应沙漠环境的独特机制和解决人类高血压、高血糖等疾病提供了分子理论基础参考。