Differences in regulation of type I collagen synthesis in primary and passaged hepatic stellate cell cultures: the role of α₅β₁-integrin

Abstract

Hepatic stellate cells (HSC) differ in their phenotype depending on the initiation and progression of their activation. Our hypothesis was that different mechanisms govern type I collagen synthesis depending on stage of HSC activation. We investigated the role of ␣5␤1-integrin as a regulator of type I collagen gene COL1A1 expression in primary and passaged HSC cultures using transgenic mouse containing type I collagen gene COL1A1 promoter linked to the chloramphenicol acetyltransferase (CAT) reporter gene. The ␣5␤1 protein levels increased during the activation and were highest in day 6 primary cultures but decreased in passaged HSC. CAT activity, reflecting COL1A1 expression, was upregulated by ␣5␤1-integrin. Inhibition of ␣5␤1-integrin by echistatin and blocking antibody resulted in reduced transgene activity only in early primary cultures (compared with the control, 53.3 Ϯ 12% echistatin and 58.8 Ϯ 7% blocking antibody, respectively, P Ͻ 0.05). Treatment of passaged HSC with either echistatin or blocking antibody had no effect. Fibronectin, an ␣5␤1-integrin ligand, increased transgene activity in primary (210 Ϯ 33%, P Ͻ 0.05) but not in passaged HSC cultures (119 Ϯ 8%). This ␣5␤1-integrin effect appears to be at least in part mediated by CCAAT enhancer binding protein-␤ (C/EBP␤), because fibronectin increased and ␣5-gene silencing by small interfering RNA decreased C/EBP␤ levels. In addition, C/EBP␤ knockout mice showed reduced type I collagen synthesis compared with wild-type littermates. Therefore ␣5␤1-integrin is an important regulator of type I collagen production in early primary HSC cultures but appears to have no direct role once the HSC are fully activated.

Fibronectin Increases Survival of Rat Hepatic Stellate Cells - A Novel Profibrogenic Mechanism of Fibronectin

“…Moreover, a number of studies have documented that FN influence HSC adhesion, proliferation, migration, and differentiation [8], induces collagen gene expression [9], wound repair, and tissue regeneration [25]. In addition of all these profibrogenic effects, in this study, we show that FN also contributes to the fibrogenic process by providing survival signals for serum depleted HSCs.…”

“…Moreover, FN interacts with cell surface receptors, including integrins [5] and non-integrin receptors [6] These integrin-mediated interactions with ECM components play crucial roles in many fundamental aspects of cell migration, growth, and differentiation [7]. FN is a profibrogenic protein, since it contributes to the organization of the ECM, helps cells attach to it, promotes activation [8] and proliferation of HSCs, induces collagen gene expression [9] and precedes the deposition of other components of the ECM in the process of liver fibrogenesis. Moreover, FN might also contribute to the liver fibrosis by increasing HSCs survival.…”

Integrins and heparan sulfate proteoglycans on hepatic stellate cells (HSC) are novel receptors for HSC‐derived exosomes

“…This represents a novel function for integrin avb3 in HSC which has previously been shown to support cell survival, adhesion to CCN2, periostin-dependent activation, and osteopontin-dependent collagen up-regulation [23,26,[30][31][32], all of which reflect the enhanced expression and a central role for integrin av or avb3 in HSC function and hepatic fibrosis [33][34][35]. Similarly, integrin a5b1 in activated HSC is associated with cell adhesion to fibronectin or CCN2, or fibronectin-dependent survival, cytoskeletal rearrangements or expression of matrix metalloproteases or collagen I [26,[36][37][38][39], but its role in binding and mediating functional effects of HSC-derived exosomes is novel. Integrin b1 on HSC also engages exosomes from liver sinusoidal endothelial cells [40] suggesting that exosomes from different cell types may compete for common integrin receptors on the same target HSC.…”

Role of c‐Jun N‐terminal kinase and p38/activation protein‐1 in interleukin‐1β‐mediated type I collagen synthesis in rat hepatic stellate cells

“…Hepatic stellate cells (HSCs) are the principal cells that generate the liver extracellular matrix (ECM), and their activation and proliferation are important factors in the development of liver fibrosis. Activated HSCs can secrete large amounts of ECM proteins (especially type I and III collagen) and tissue inhibitors of metalloproteinases, thereby leading to the accumulation of ECM in the liver (1–4).…”