硫化氢前体L-半胱氨酸对结肠动力的影响及机制
全晓静¹, 邹百仓¹, 秦斌¹, 戴菲¹, 罗和生², 王进海^1,*
1西安交通大学医学院第二附属医院消化科，西安 710004；²武汉大学人民医院消化科，武汉 430060
摘要
本文旨在研究硫化氢(hydrogen sulfide, H2S)前体L-半胱氨酸对大鼠结肠动力的影响，以阐明其对结肠收缩的调节作用及机制。采用免疫组织化学染色和免疫印迹实验检测内源性H2S合成酶胱硫醚-γ-裂解酶(cystathionine-γ-lyase, CSE)和胱硫醚-β-合成酶(cystathionine-β-synthase, CBS)在大鼠近端结肠的表达情况；采用生理记录仪检测结肠平滑肌收缩活动的变化；利用膜片钳实验检测结肠平滑肌细胞离子通道电流。结果显示，CBS和CSE在大鼠近端结肠黏膜层、平滑肌层及肌间神经丛均有表达；L-半胱氨酸以浓度依赖的方式抑制近端结肠纵行平滑肌收缩，H₂S合成酶抑制剂氨基氧乙酸(aminooxyacetate acid, AOAA)和炔丙基甘氨酸(propargylglycine, PAG)孵育纵行平滑肌后，L-半胱氨酸半最大效应浓度(concentration for 50% of maximal effect, EC50)相比对照组显著下降(P < 0.05)；而L-半胱氨酸对结肠环形肌收缩具有抑制和促进双重调节作用，AOAA和PAG预处理可阻断其对环形肌的兴奋作用；H₂S外源性供体NaHS在低浓度时促进平滑肌细胞L型钙通道开放(P < 0.01)，而在高浓度时抑制L型钙通道电流(I_Ca,L) (P < 0.05)；与NaHS不同，L-半胱氨酸浓度依赖性抑制I_Ca,L (P < 0.01)；NaHS抑制大电导钙激活钾电流(I_BKCa)，而L-半胱氨酸对IBKCa无明显作用。以上结果提示，L-半胱氨酸对大鼠结肠平滑肌收缩具有潜在双重调节作用，其中抑制作用由L型钙通道介导，而促进作用可能是由内源性生成的H₂S介导。


关键词： L-半胱氨酸; 硫化氢; 离子通道; 膜片钳; 大鼠
分类号：R333;Q48

Effect of L-cysteine on colonic motility and the underlying mechanism
QUAN Xiao-Jing¹, ZOU Bai-Cang¹, QIN Bin¹, DAI Fei¹, LUO He-Sheng², WANG Jin-Hai^1,*
1Department of Gastroenterology, Second Affiliated Hospital of Xi’an Jiaotong University, Xi’an 710004, China；²Department of Gastroenterology, Renmin Hospital of Wuhan University, Wuhan 430060, China
Abstract
The purpose of the present study is to investigate the effect of L-cysteine on colonic motility and the underlying mechanism. Immunohistochemical staining and Western blot were used to detect the localization of the H2S-generating enzymes cystathionine-β-synthase (CBS) and cystathionine-γ-lyase (CSE). Organ bath system was used to observe the muscle contractile activities. Whole-cell patch-clamp technique was applied to record ionic channels currents in colonic smooth muscle cells. The results showed that both CBS and CSE were localized in mucosa, longitudinal and circular muscle and enteric neurons. L-cysteine had a dual effect on colonic contraction, and the excitatory effect was blocked by pretreatment with CBS inhibitor aminooxyacetate acid (AOAA) and CSE inhibitor propargylglycine (PAG); L-cysteine concentration-dependently inhibited L-type calcium channel current (I_Ca,L) without changing the characteristic of L-type calcium channel (P < 0.01); In contrast, the exogenous H₂S donor NaHS increased I_Ca,L at concentration of 100 μmol/L, but inhibited I_Ca,L and modified the channel characteristics at concentration of 300 μmol/L (P < 0.05); Furthermore, L-cysteine had no effect on large conductance calcium channel current (I_BKCa), but NaHS significantly inhibited I_BKCa (P < 0.05). These results suggest that L-cysteine has a potential dual effect on colonic smooth muscle and the inhibitory effect might be directly mediated by L-type calcium channel while the excitatory effect might be mediated by endogenous H₂S.


Key words: L-cysteine; hydrogen sulfide; ion channel; patch-clamp technique; rat

收稿日期：2019-10-20　　录用日期：2020-03-19
通讯作者：王进海　　E-mail: xhnkwjh@126.com
DOI: 10.13294/j.aps.2020.0037
引用本文：
全晓静, 邹百仓, 秦斌, 戴菲, 罗和生, 王进海. 硫化氢前体L-半胱氨酸对结肠动力的影响及机制[J]. 生理学报 2020; 72 (3): 299-307.
QUAN Xiao-Jing, ZOU Bai-Cang, QIN Bin, DAI Fei, LUO He-Sheng, WANG Jin-Hai. Effect of L-cysteine on colonic motility and the underlying mechanism. Acta Physiol Sin 2020; 72 (3): 299-307 (in Chinese with English abstract).



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