Figure 1. Evaluation of the fibrotic markers and expressions of the transglutaminase (TG) isozymes during liver fibrosis.

To induce liver fibrosis, the common bile duct was double-ligated and cut between the ligatures in 8 weeks-old ICR mice. The mice were then sacrificed on days 3, 7, and 14 after surgery (n = 3 mice). (A) Liver sections were fixed in 4% paraformaldehyde, and then stained using hematoxylin and eosin (HE), Sirius Red (using the Sirius Red Collagen Detection Kit), and immunohistochemistry (using anti-α smooth muscle actin (αSMA) antibody and rabbit non-immune immunoglobulin G (NI-IgG) as the negative control). The red and green colors indicate the fibrillar collagen (type I to V collagen) and non-collagenous protein, respectively. Bar = 100 μm. (B) Hydroxyproline (HDP) contents were evaluated in the liver on each indicated day after bile duct ligation (BDL). (*P < 0.05; **P < 0.01) (C and D) The mRNA expression levels of the fibrotic markers (Collagen Iα1 (ColA1), αSMA, transforming growth factor (TGF)-β1, GAPDH and the TG family (FXIIIa and TG1–7) were confirmed by RT-PCR. The successful detections for TG3–7 using each primer pair are confirmed in the other tissue extracts. (E) The protein levels in the whole lysate from the liver tissue were analyzed by immunoblotting using each antibody and glyceraldehydes 3-phosphate dehydrogenase (GAPDH) as a loading control for each sample.