TNF-α抑制TGF-β引發CTGF表現之研究

Abstract

TGF-β調控許多生理功能，例如細胞生長、細胞凋亡、細胞分化及細胞外間質 (ECM) 之合成等。CTGF為TGF-β 所調控之基因，可調節ECM相關蛋白之表現。雖然已有研究指出TNF-α可抑制TGF-β引發之CTGF表現，但詳細機轉仍不清楚。本論文發現，在WT MEF細胞，TNF-α可抑制TGF-β引發之CTGF表現，而此抑制作用在p65-/- MEF細胞消失，表示p65扮演重要角色。長時間 (8小時) 處理TNF-α可增加抑制性Smad之Smad7的表現，且阻斷Smad2之磷酸化而抑制TGF-β之signaling，在p65-/- MEF細胞，TNF-α無法引發Smad7之表現，亦不抑制Smad2之磷酸化；短時間 (1小時) 處理TNF-α可活化NF-κB signaling，使p65入核與Smad4競爭p300，造成Smad複合體與CTGF promoter結合時無法吸引p300，因而降低CTGF之表現。因此TNF-α是透過這兩種機制抑制TGF-β所引發之CTGF表現。

TGF-β regulates diverse biologic effects including cell growth, cell death or apoptosis, cell differentiation, and extracellular matrix (ECM) synthesis. Connective tissue growth factor (CTGF), which is induced by TGF-β, has been reported to mediate stimulatory action of TGF-β-induced ECM. Although TNF-α was reported to suppress the TGF-β-induced CTGF gene expression, the molecular mechanism is not well- clarified. In this study, we found the inhibitory effect of TNF-α on TGF-β-induced CTGF expression in WT MEF cells, and the attenuation was abrogated in p65-/- MEF cells, suggesting the role of p65 in TNF-α-mediated inhibition. Long-term (8 hours) treatment of TNF-α induced the expression of inhibitory Smad (Smad7) and the inhibition of TGF-β-induced Smad2 phosphorylation, thereby inhibiting TGF-β signaling which were not seen in p65-/- MEF cells. Short-term (1 hours) treatment of TNF-α activated NF-κB, and p65 preferentially interacted with p300, thereby disrupting TGF-β-induced interaction of Smad4 with p300 on the CTGF promoter. Two molecular mechanisms of TNF-α inhibition are explored. One is the induction of Smad7 expression, and the other is the switch of binding preference of p300 from Smad4 to p65, thereby leading to the inhibition of TGF-β-induced CTGF expression.