Adenoviral expression of the Nephroblastoma overexpressed gene product (CCN3/NOV) induces apoptosis and inhibits epithelial-to-mesenchymal transition (EMT) in hepatocytes in vitro but fails to attenuate ongoing liver fibrogenesis in vivo

CCN3/NOV encoded by the Nephroblastoma overexpressed gene represents a matricellular protein of the CCN family. These proteins comprise six different secreted proteins associating specifically with the extracellular matrix [1]. CCN proteins lack specific high-affinity receptors but signal through integrins and proteoglycans and regulate crucial biological processes in liver including fibrosis [2]. Previously we have demonstrated that CCN3 expression increases dramatically during activation of hepatic stellate cells (HSC) and in models of experimental liver fibrosis [3]. Notably, suppression of CCN3 enhanced expression of profibrotic marker proteins including Connective tissue growth factor (CCN2/CTGF) in primary HSC and in the immortalized stellate cell line CFSC. Vice versa, the overexpression of CCN2 reduced CCN3 expression. Surprisingly, the adenoviral transfer of CCN3 enhanced CCN2 expression suggesting that the CCN protein network is highly complex and extends well beyond the classic Yin and Yang interaction. In the present study, we analysed the effects of CCN3 overexpression in a model of ongoing fibrogenesis induced by ligature of the common bile duct and in primary cultured hepatocytes. Contrasting to the findings in HSC, transfer of CCN3 resulted in marked reduced levels of CCN2 in primary hepatocytes. In addition, CCN3 prevented TGF-β-induced expression of Snail1, vimentin. fibronectin and matrix metalloproteinase 9 that are associated in vitro with EMT. We further found that Ad5-CMV-NOV significantly induce hepatocyte apoptosis as demonstrated in positive TUNEL staining and Western blot analysis of cleaved caspase–3. The infection of hepatocytes with Ad5-CMV-NOV further resulted in strong activation of the JNK and p38 MAPKs. However, the adenoviral gene transfer of CCN3 was not sufficient to attenuate ongoing fibrogenesis indicating that CCN3 treatment alone is not sufficient to inhibit the fibrotic pathway in the bile duct ligation model.

Literatur: [1] Bork P: The modular architecture of a new family of growth regulators related to connective tissue growth factor. FEBS Lett 1993, 327:125-130. [2] Weiskirchen R. CCN proteins in normal and injured liver. Front Biosci 2011;16:1939-1961. [3] Borkham-Kamphorst E, van Roeyen CR, Van de Leur E, Floege J, Weiskirchen R. CCN3/NOV small interfering RNA enhances fibrogenic gene expression in primary hepatic stellate cells and cirrhotic fat storing cell line CFSC. J Cell Commun Signal 2011, in press.

BDL - CCN2 - CCN3 - EMT - adenovirus gene transfer - animal models. - apoptosis - hepatic stellate cells - hepatocytes - inflammation - liver fibrosis