Department of Physiology, College of Medicine, University of Kentucky, Lexington, KY 40506-0298, USA.
Am J Physiol Regul Integr Comp Physiol. 2013 May 1;304(9):R702-11. doi: 10.1152/ajpregu.00296.2012. Epub 2013 Feb 27.
The present study investigated transient receptor potential vanilloid type 4 (TRPV4) ion channels in pancreatic stellate cells (PSCs) isolated from rats with high-fat and alcohol diet (HFA)-induced chronic pancreatitis. TRPV4 is a calcium-permeable nonselective ion channel responsive to osmotic changes, alcohol metabolites arachidonic acid, anandamide, their derivatives, and injury-related lipid mediators. Male Lewis rats were fed HFA for 6-8 wk before isolation and primary culture of PSCs. Control PSCs were harvested from rats fed standard chow. Immunoreactivity for cytoskeletal protein activation product α-smooth muscle actin (α-SMA) and platelet-derived growth factor receptor-β subunit (PDGFR-β) characterized the cells as PSCs. TRPV4 expression increased in PSCs of HFA-fed rats and control cultures after alcohol treatment (50 mM). Cell responses to activation of inducible TRPV4 were assessed with live cell calcium imaging. Threefold increased and sustained intracellular calcium mobilization responses occurred in 70% of pancreatic stellate cells from HFA-fed rats in response to TRPV4 activators arachidonic acid, lipid second messenger, phorbol ester 4 α-phorbol 12,13-didecanoate (4αPDD), and 50% hypoosmotic media compared with relatively unresponsive PSCs from control rats. Activation responses were attenuated by nonselective TRPV channel blocker ruthenium red. Tumor necrosis factor-α (TNF-α, 1 ng/ml, 16 h) increased responses to 4αPDD in control PSCs. These findings implicate TRPV4-mediated calcium responses inducible after HFA exposure and inflammation in reactive responses of activated PSCs that impair pancreatic function, such as responsiveness to cytokines and the deposition of collagen fibrosis that precipitates ductal blockage and pain.
本研究探讨了高脂和酒精饮食（HFA）诱导的慢性胰腺炎大鼠胰腺星状细胞（PSC）中瞬时受体电位香草酸 4 型（TRPV4）离子通道。TRPV4 是一种对渗透压变化、酒精代谢产物花生四烯酸、大麻素、其衍生物以及与损伤相关的脂质介质敏感的钙通透性非选择性离子通道。雄性 Lewis 大鼠在分离和原代培养 PSC 之前，用 HFA 喂养 6-8 周。对照组 PSC 从标准饲料喂养的大鼠中获得。细胞骨架蛋白激活产物α-平滑肌肌动蛋白（α-SMA）和血小板衍生生长因子受体-β亚基（PDGFR-β）的免疫反应性将细胞鉴定为 PSC。在酒精处理（50 mM）后，HFA 喂养大鼠和对照培养物中的 PSC 中 TRPV4 表达增加。使用活细胞钙成像评估诱导型 TRPV4 的细胞反应。与对照组大鼠的 PSC 相比，HFA 喂养大鼠的 PSC 对 TRPV4 激活剂花生四烯酸、脂质第二信使佛波醇 12,13-二癸酸酯（4αPDD）和 50%低渗介质的反应分别增加了三倍，并持续增加细胞内钙动员反应，而相对无反应性的 PSC 则增加了三倍。非选择性 TRPV 通道阻滞剂钌红可减弱激活反应。肿瘤坏死因子-α（TNF-α，1ng/ml，16h）增加了对照组 PSC 对 4αPDD 的反应。这些发现表明，HFA 暴露和炎症后诱导的 TRPV4 介导的钙反应以及激活的 PSC 的反应性增强参与了胰腺功能的损害，例如对细胞因子的反应性和胶原纤维的沉积，这会导致导管阻塞和疼痛。
