FIGURE 4.

CCN1 stimulates liver macrophage efferocytosis and enhances Tgfb1 expression. Liver macrophages and neutrophils (PMN) were isolated from Ccn1 ^WT/WT and Ccn1 ^D125A/D125A mice 48 hours after CCl₄ injury. (A) Apoptotic neutrophils (PMN; stained blue with DAPI) were pretreated with BSA or CCN1 protein (4 μg/ml) for 60 min and incubated with macrophages (stained with CellTracker Green) for 90 min. The field of view in each panel of Figure 4A is 35 × 35 μm. The width of each cell is approximately 20 μm. (B) The number of DAPI‐positive macrophages was counted and expressed as a percentage of total macrophages. (C) The expression of Tgfb1 was assessed by qRT‐PCR. The vehicle samples contained no PMN. (D) Apoptotic neutrophils from Ccn1 ^WT/WT and Ccn1 ^D125A/D125A were pretreated with the indicated proteins for 60 min before incubation with macrophages from Ccn1 ^WT/WT (left panel) and Ccn1 ^D125A/D125A (right panel) mice, and the expression of Tgfb1 was assessed by qRT‐PCR. Data represent means ± SD. *p < 0.033, **p < 0.002; Student t test. BSA, bovine serum albumin; CCl₄, carbon tetrachloride; CCN1, cellular communication network factor 1; DAPI, 4′,6‐diamidino‐2‐phenylindole; MФ, macrophage; mRNA, messenger RNA; PMN, polymorphonuclear; qRT‐PCR, quantitative reverse‐transcription polymerase chain reaction; Rel., relative; Tgfb1, transforming growth factor‐β1; WT, wild type; αSMA, alpha smooth muscle actin.