Human MMP Antibody Array from Abcam was used to profile MMPs in macrophages. Gelatin or casein zymography was performed using 10% SDS-polyacrylamide gels (SDS-PAGE) containing gelatin (1 mg/ml) or Casein (1 mg/ml) as substrate. HSCs migration assay was performed using Boyden chamber as described previously (Guo et al., 2007, McGarrigle et al., 2006, Shan et al., 2006 and Yang and Huang, 2005). Real-time PCR with SYBR green was performed using iTaq™ universal SYBR® Green supermix (BIO-RAD) and a 7500 Real-Time PCR System (Applied Biosystems). Collagen, type I, alpha 1 (COL1A1), alpha smooth muscle actin (α-SMA) expression was determined by immunoblot analysis.
We first profiled the expression of all MMPs in primary murine bone marrow-derived macrophages (BMDMs) and differentiated THP-1 cells and found that MMP-8, -10, & -13, were significantly overexpressed after 12 h of lipopolysaccharide (LPS) treatment. Based on this pattern of expression, we speculated that macrophage MMP-8,-10, &-13 might play a non-canonical role in HSCs activation. Further, we found that exogenous active MMP-8 (Collagenase-2) treated HSC shows markedly increased migration and COL1A1 expression as compared to MMP-10 and MMP-13 treated HSCs. Thus, macrophage MMP-8 (Collagenase-2) expression in macrophages emerges as an important moderator of HSC cell migration and invasion.
These findings suggest that macrophage MMP-8 promotes HSC activation and might have a role in liver disease progression. MMP-8 targeting in the liver may have therapeutic potential in alcoholic liver disease (ALD).