Fig. 7

Recombinant CCL5 rescued the impaired muscle regeneration of C3aR−/− mouse. a At day 0, 1, 3 after injury, mRNA levels of cytokines CCL5 in WT and C3aR−/− muscles were accessed by real-time PCR. (N = 4 in each group). b At day 1 after CTX injury, WT (N = 8) and C3aR−/− (N = 4) muscle extract were seeded in the lower chamber and WT monocytes (5 × 10⁴/well) were seeded in the upper chamber. Twelve hours after co-culture, migrated cells from upper chamber were counted by DAPI staining (blue). (Bars, 50 μm, ten filed was collected per sample). c Schematic diagram of the protocol. Recombinant mouse CCL5 (0.1 mg kg⁻¹) was intramuscular injected to C3aR−/− muscle at day 0 and day 2 after CTX injury. At day 3 after injury, TA muscle was harvested to examine the inflammation response, and at day 30 after injury, TA muscle was harvested to examine the regeneration. d mRNA levels of CD45 and F4/80 in WT, C3aR−/− and C3aR−/− with rCCL5 muscles were accessed by real-time PCR. (N = 4 in each group). e CD45⁺ leukocytes and F4/80⁺ macrophages in muscles were detected by FACS. The right graph indicated the percentages of CD45⁺ leukocytes and F4/80⁺ macrophages (N = 4 in each group, graphs show representatives of two independent experiments). f mRNA levels of MyoD in WT, C3aR−/− and C3aR−/−with rCCL5 muscles were accessed by real-time PCR. (N = 4 in each group). g Masson staining was used to detect the collagen deposition (green color) in muscle (at day 15 after injury) of WT and C3aR−/− mice, the percentages of fibrosis area per field were analyzed. (Bars, 50 μm; N = 4 in each group). h The mean myofiber CSA in injured muscles from WT, C3aR−/− and C3aR−/− with recombinant CCL5 mice at day 30 after injury were measured. (N = 4 in each group, bars, 50 μm). Data are expressed as the mean ± s.e.m. *P < 0.05, **P < 0.01, unpaired t-test, two-tailed