Expression analysis of CCN1/CYR61 in primary liver cells and in models of experimental liver fibrogenesis

Objective: Cysteine-rich protein 61 (CCN1/CYR61) is a matricellular protein of the CCN family that comprises six secreted CCN proteins in mammals [1]. In adults, CCN1/CYR61 expression is associated with inflammation and injury repair. Recent studies showed that CCN1/CYR61 limits fibrosis in models of cutaneous wound healing by inducing cellular senescence in myofibroblasts of the granulation tissue which thereby transform into an extracellular matrix (ECM)-degrading phenotype. In fibrotic livers, hepatic stellate cells (HSC) and periportal myofibroblasts (pMF) are considered to produce ECM. Design and Results: We here measured CCN1/CYR61 expression in HSC and pMF. We found a dramatic increase during activation of HSC and a subsequent decline in fully transdifferentiated myofibroblasts (MFB). In contrast, CCN1/CYR61 levels in primary hepatocytes were only marginal compared to the levels found in HSC and pMF. Quantitative RT-PCR using RNA isolated from livers of animals that were subjected to bile duct ligation (BDL) and repeatedly applications of CCl4 showed increased CCN1/CYR61 mRNA quantities compared to livers taken from untreated controls. In these models, elevated levels of CCN1/CYR61 were particularly found during early periods of liver injury, while the expression declined during prolonged phases of fibrogenesis. In line, Western blot analysis further revealed that the protein quantities of CCN1/CYR61 declined at later stages of BDL. The initial CCN1/CYR61 upregulation correlated well with the expression of α-smooth muscle actin (α-SMA) and collagen type I (col1α) in early stages of in vivo liver fibrogenesis. Likewise, we observed a correlation to the acute phase response protein Lipocalin 2 (LCN2) that was recently shown to be linked to acute and chronic liver damage and controlled by interleukin-1beta via the nuclear factor-kappaB (NF-kappaB) pathway [2]. Immunohistochemistry confirmed the initial elevation and subsequent decline of CCN1/CYR61 expression in respective injury models. Conclusions: CCN1/CYR61 is obviously linked to the pathogenesis of hepatic fibrogenesis and most likely involved in the control of senescence in transdifferentiated MFB and pMF.