基于HIF-1α信号途径研究硫化氢对缺氧诱导Aβ生成和聚积的抑制作用及机制

Overproduction of amyliod-β peptide(Aβ) is core pathogenic mechanism of Alzheimer's disease (AD), but there is not effective intervention to prevent Aβ overproduction. It is known that hypoxia increase the expression of β-site amyloid precursor protein cleaving enzyme 1(BACE1) and APH-1A, a component of γ-secretase, to activate β- and γ-secretase by elevating HIF-1α expression, and then promote Aβ production. It was reported that the levels of hydrogen sulfide were severely decreased in the brains of Alzheimer's disease patients compared with the brains of the age matched normal individuals and the treatment of hydrogen sulfide attenuated Alzheimer-like pathological changes. Further, research confirmed that hydrogen sulfide inhibited HIF-1α overexpression during hypoxia. However, whether hydrogen sulfide inhibited hypoxia-induced overexpression of BACE1 and APH-1A through regulating HIF-1α signaling pathway or inhibited the activation of β and γ secretase to prevent Aβ production is not yet clear. In preliminary experiments, we found that hypoxia decreased the levels of hydrogen sulfide in neurons and hydrogen sulfide prevented Aβ overproduction induced by hypoxia. Based on the above information, we speculate that hydrogen sulfide decrease the overexpression of BACE1 and APH-1A by reducing HIF-1α overexpression during hypoxia to inhibit β and γ secretase, so hydrogen sulfide reduces Aβ production. To confirm above-mentioned hypothesis, we'll employ LSCM, Immunohistochemistry, Western blot, qPCR, ELISA and RNAi methods to illuminate mechanisms and inhibitory action of hydrogen sulfide on Aβ overproduction during hypoxia. The project shall provide new ideas for prevention and treatment of Alzheimer's disease.

Aβ过量生成是阿尔茨海默病(AD)核心致病机制，但缺乏有效干预措施。研究表明缺氧通过HIF-1α，上调BACE1和APH-1A表达，激活β、γ分泌酶，使Aβ生成增多。调查发现AD患者H2S浓度低于对照组，且H2S对AD样病变具有抑制作用。文献证实H2S抑制HIF-1α过表达，但能否通过调控HIF-1α信号途径抑制缺氧诱导BACE1、APH-1A高表达和β、γ分泌酶的激活，阻止Aβ过量生成，目前尚不清楚。课题组预实验发现缺氧降低H2S浓度，NaHS预处理抑制缺氧时Aβ的过量生成。基于此，申请者推测硫化氢通过抑制缺氧时HIF-1α的过表达，减少BACE1和APH-1A表达，抑制β和γ分泌酶的激活，减少Aβ的生成和聚积。为验证假设，拟以海马神经元和转基因鼠为研究对象，采用激光共聚焦、免疫组化、Wb、qPCR、 ELISA和RNAi等技术，阐明上述问题。研究成果将为AD临床防治提供新思路。

Aβ过量生成是阿尔茨海默病(AD)核心致病机制，但缺乏有效干预措施。研究表明缺氧通过HIF-1α，上调BACE1和APH-1A表达，激活β、γ分泌酶，使Aβ生成增多。调查发现AD患者H2S浓度低于对照组，且H2S对AD样病变具有抑制作用。文献证实H2S抑制HIF-1α过表达，但能否通过调控HIF-1α信号途径抑制缺氧诱导BACE1、APH-1A高表达和β、γ分泌酶的激活，阻止Aβ过量生成，目前尚不清楚。本项目采用激光共聚焦、免疫组化、免疫印迹、ELISA和RNAi等技术，研究发现：（1）缺氧使脑组织硫化氢浓度降低；（2）硫化氢阻止缺氧时HIF-1a的高表达；（3）硫化氢抑制缺氧诱导BACE1和APH-1A的高表达；（4）硫化氢减轻缺氧诱导β和γ分泌酶活性的升高；（5）硫化氢减少缺氧诱导Aβ含量的增加；（6）过表达HIF-1a增高β和γ分泌酶的活性；（7）下调HIF-1a的表达抑制缺氧诱导β和γ分泌酶活性的增高；（8）硫化氢减少缺氧所致转基因鼠老年斑数量的增加。上述结果表明硫化氢通过抑制HIF-1α高表达，减少缺氧诱导BACE1和APH-1A的高表达，抑制β和γ分泌酶的激活，从而减少缺氧时Aβ的过量生成和聚积。本项目采用多种技术，从多个角度阐明了硫化氢在缺氧诱导Aβ过量生成和聚积的作用及其机制，研究成果为临床AD防治提供新思路和新靶点。