Variable expression of cystatin C in cultured trans-differentiating rat hepatic stellate cells

Figure 1 CysC mRNA expression in different rat liver cell types. A: Equal amounts of total RNA samples (10 µg) isolated from cultured SECs, MFBs, HSCs, KCs, and PCs were analyzed by Northern blot. The blot was hybridized with a ³²P-labeled probe specific for rat CysC; B: total RNA samples (5 µg/lane) of trans-differentiating HSCs/MFBs at indicated time intervals were analyzed for CysC expression by Northern blot analysis. As internal loading controls (A and B), the 18S and 28S rRNA as well as signals obtained after hybridization with a GAPDH-specific probe are shown. The blots were exposed for 4 (GAPDH) or 24 h (CysC), respectively; C: protein extracts (20 µg/lane) from primary rat HSCs cultured for 1 to 7 days or rat MFBs cultured for 3 days were analyzed for CysC expression by Western blotting under reducing conditions (50 mmol/L DTT) using antibody P-14 (sc-16989). To verify cellular trans-differentiation, the blot was stripped, and tested for α-SMA expression using a mouse monoclonal antibody (clone asm-1).