剪力對內皮細胞中硫化氫生成之調控與訊息傳導之影響

Regulation of H2S Production and H2S-Dependent Signal Transduction in Endothelial Cells by Shear Stress

陳長廷(Chang-Ting Chen)

指導教授 ： 謝學真

共同指導教授 ： 王寧(Danny Ling Wang)

國立臺灣大學/工學院/化學工程學研究所/碩士(2011年)

https://doi.org/10.6342/NTU.2011.03297

摘要

硫化氫是近年來受到許多研究關注的細胞內氣體傳導分子(gasotransmitter)，在心血管系統中，硫化氫產生許多的保護作用。層流剪力(laminar shear stress)亦是提供心血管保護作用的因子之一，且剪力與硫化氫所誘發的訊息傳導多有類似之處。然而，目前關於剪力對硫化氫生成量的影響，乃至於硫化氫在剪力誘導的訊息傳導方面的知識非常有限，因此，本論文將對此課題作深入討論，主要討論方向有兩點：第一，剪力是否增加細胞內硫化氫生成量；第二，硫化氫對剪力誘導的訊息傳導之影響。

由於在心血管系統中血液流動所產生的剪力最主要作用在血管內皮細胞(vascular endothelial cells)上，因此，本論文主要以人類臍帶靜脈內皮細胞(HUVECs)以及牛大動脈內皮細胞(BAECs)為研究對象。實驗發現，層流剪力在作用30分鐘之後，會使BAECs內的硫化氫生成速率提升5倍。然而，三種可能參與硫化氫生成的酵素：cystathionine-γ-lyase (CSE)、cystathionine-β-synthase (CBS)與3-mercapto-sulfurtransferase (3-MST)的蛋白質量在剪力作用30分鐘後皆無明顯變化，顯示剪力可能透過增加上述硫化氫生成酵素之活性來提高硫化氫之生成量。至於探討硫化氫對剪力誘導的訊息傳導之影響方面，主要使用CSE、CBS與3-MST的siRNA分別抑制其表現量，再觀察剪力刺激下細胞內訊息傳導的變化。結果發現pAkt在受剪力刺激後所被誘導的程度會被上述三種siRNA所抑制，可以推論剪力作用所造成的pAkt上升，部分是由於細胞內硫化氫增高所導致。

綜合以上研究結果，可以發現剪力所產生的心血管保護作用可能部份來自於內皮細胞中硫化氫濃度升高的效應，且CSE、CBS以及3-MST可能是負責硫化氫生成主要的酵素。此外，實驗結果更進一步指出pAkt可能在硫化氫相關的心血管保護作用中扮演重要的角色。

關鍵字

剪力 ； 硫化氫 ； 內皮細胞 ； 心血管保護 ； Akt

並列摘要

Hydrogen sulfide (H2S) is a novel gasotransmitter in cardiovascular system, it renders multiple cardioprotective effects. Laminar shear stress is also a cardioprotective factor in cardiovascular system, which shares many similarities in downstream signal transduction pathways with H2S. However, knowledge about the regulation of H2S production by shear stress and the role of H2S in shear-induced signal transduction is still limited. In the present study, efforts were made to investigate the effect of shear stress on H2S production. Furthermore, the role of H2S in shear-induced cell signaling is also discussed.

In cardiovascular system, shear stress exerted by blood flow mainly affects vascular endothelial cells (ECs). Thus, human umbilical vein endothelial cells (HUVECs) and bovine aortic endothelial cells (BAECs) are used in present study. After laminar shear stress stimulation for 30 minutes, H2S production rate is enhanced by 5 fold in BAECs. Three enzymes possibly involved in H2S production—cystathionine-γ-lyase (CSE), cystathionine-β-synthase (CBS), and 3-mercapto-sulfurtransferase (3-MST)—are not significantly affected by shear stress in terms of protein and mRNA level, implying the possibility that shear stress enhaces H2S production rate by increasing CSE, CBS, and 3-MST enzyme activities. In order to study the role of H2S in shear-induced cell signaling, CSE, CBS, and 3-MST were knocked down by siRNAs before HUVECs were stimulated by shear stress. Results show that shear-induced Akt phosphorylation is inhibited by CSE, CBS, and 3-MST siRNAs.
In conclusion, shear-induced cardioprotective effect might be partly a result of increased H2S concentration in ECs. Futrther, shear stress induces Akt phosphorylation in a H2S-dependent way.

並列關鍵字

shear stress ； hydrogen sulfide ； endothelial cells ； cardioprotective ； Akt

參考文獻

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