内源性硫化氢与重症急性胰腺炎结肠动力的相关研究

Inhibition of intestinal motility is one of complications in the early phase of severe acute pancreatitis(SAP) and plays a critical role initiating the development of SAP. However, the mechanisms of SAP-induced dysmotility are largely unknown. Hydrogen sulfide (H2S) has recently been identified as a new "gasotransmitter''. Cystathionine β-synthase(CBS) and systathionine -γ-lyase (CSE) are two important enzymes for generation of endogenous H2S. They have been shown to be expressed in the gastrointestinal tract. H2S can inhibit intestinal motility and the mechanism through which H2S exerts its relaxant properties is related to the direct opening of ATP-sensitive potassium (KATP) channels located in the smooth muscle cells. By now, little is known about the link between endogenous synthesis of H2S and the colonic motility in SAP. The project intends to explore the potential role of colonic endogenous H2S and the two enzymes(CSE and CBS) for H2S synthesis on colonic dysmotility in a rat model of SAP, using a series of methods such as electrophysiology, cell and molecular biology technologies, from multiple levels such as entirety, tissues, cells, ion channels. We also examine the potential mechanism of inflammatory mediators-PI3K/Akt signaling pathway-transcription factor Spl on the enzymes for H2S synthesis (mainly CSE), using a lot of technologies such as RNA interference. This will provide a theoretical basis for the prevention and treatment of SAP-induced colonic dysmotility .

肠道动力减退是重症急性胰腺炎(SAP)早期病程中常见并发症，对SAP发展与转归至关重要，至今其发生机制尚未完全阐明，临床缺乏有效防治手段。硫化氢是近年来发现的一种新型气体信号分子。消化道组织分布硫化氢合成酶(CSE和CBS)，能合成内源性硫化氢。硫化氢通过激活平滑肌细胞膜ATP敏感性钾通道而抑制肠道动力。目前未见内源性硫化氢与SAP肠道动力相关报道。本项目拟构建SAP结肠动力减退大鼠模型，采用一系列电生理学、细胞、分子生物学方法，从整体、组织、细胞、分子、离子通道等多个层面上探讨内源性硫化氢/ 硫化氢合成酶(CSE和CBS)对SAP结肠动力的影响；同时运用RNA干扰等技术，从正反两方面揭示炎症介质-PI3K/Akt信号通路-转录因子Spl对硫化氢合成酶(主要为CSE)表达的调控机制，为SAP结肠动力紊乱防治的新策略提供理论基础。

肠道动力减退是重症急性胰腺炎(SAP)早期病程中常见并发症，对SAP发展与转归至关重要，至今其发生机制尚未完全阐明。消化道组织分布硫化氢合成酶(CSE和CBS)，能合成内源性硫化氢(H2S)；H2S可抑制结肠动力。本项目拟构建SAP结肠动力减退大鼠模型，探讨内源性硫化氢与SAP肠道动力关系以及炎症介质-PI3K/Akt信号通路-Spl对CSE表达的调控机制。发现：①SAP大鼠结肠动力减退，与CBS和CSE表达升高内源性H2S合成增加相关；使用PAG后CSE表达减少，可部分逆转SAP结肠动力紊乱。②SAP血浆TNF-α、IL-6和IL-1β的浓度增加；使用CSE抑制剂PAG后H2S合成减少，其浓度显著降低，提示H2S和炎症因子之间可能存在相关性。③全细胞膜片钳发现IL-6可减少结肠平滑肌细胞L-Ca通道电流密度，从而导致平滑肌的收缩减弱。提示SAP动力减退亦与IL-6合成增加，Ca2+内流减少相关。④SAP大鼠血浆、TNF-α和IL-6分别培养肠道平滑肌细胞及肠神经细胞，均发现CSE和CBS表达上调，提示在SAP时炎症因子可增加内源性H2S的合成。⑤SAP大鼠血浆、TNF-α和IL-6，促进肠道平滑肌细胞及肠神经细胞PI3K/Akt/ Sp1/CSE的表达升高；采用PI3K的抑制剂LY294002及使用转染Sp1的siRNA抑制了Sp1活性后，CSE合成减少；提示SAP时炎症介质的产生增加激活了PI3K/Akt通路，通过上调Sp1的表达从而使内源性H2S产生增加。以上结果表明：SAP结肠动力减退，与局部内源性H2S合成增加有关，亦可能与IL-6合成增加导致结肠平滑肌Ca2+内流减少相关；SAP时炎症因子激活PI3K/Akt/Sp1的信号通路可使结肠内源性H2S的生成增加。