Sex differences in CXCL4‐mediated lung damage in the bleomycin‐induced systemic sclerosis mice model

Abstract

Abstract Background Sex is an independent risk factor for systemic sclerosis (SSc). Chemokine ligand 4 (CXCL4) participates in SSc via multiple mechanisms. In the present study, we investigated the role of CXCL4 in sex‐specific lung injury in the bleomycin‐induced systemic sclerosis (BLM‐SSc) mouse model. Methods Thirty‐two 6‐week‐old CXCL4–/– C57BL/6J and wild‐type (WT) mice were divided into CXCL4–/– male, CXCL4–/– female, wild‐type male (WT male), and wild‐type female (WT female) groups (n = 8 each). BLM‐SSc mice were established by subcutaneous injections of 100 µL (2 mg/mL of bleomycin, once every other day) for 4 weeks. Thereafter, all mice were sacrificed and the extracted lung tissues were stained with hematoxylin and eosin and Masson's trichrome to evaluate the degree of inflammation and fibrosis. Enzyme‐linked immunosorbent assay was used to measure the levels of tumor necrosis factor‐α and interleukin‐6 in the lung tissue homogenate. In addition, immunofluorescence double staining detected the coexpression of S100 calcium‐binding protein A8 and matrix metalloproteinase 7. Two‐way analysis of variance was used to examine the independent effects of sex, CXCL4, and their interactions. Results Compared to CXCL4–/– mice, WT mice had significantly higher infiltration of macrophages and neutrophils, inflammation scores, and fibrosis scores (all p 0.05). Compared with that of the CXCL4–/– mice, the levels of tumor necrosis factor‐α and interleukin‐6 in the lung tissue homogenate were increased in the WT mice with higher levels in the male group. No significant sex‐based differences were observed in CXCL4–/– mice (p > 0.05). Immunofluorescence double staining revealed a significantly higher number of positive double‐stained cells in the WT male than the WT female group; however, no such cells were observed in CXCL4–/– groups. Conclusion CXCL4 may participate in the pathogenesis of sex‐based differential lung injury in the BLM‐SSc model.

Keywords

• CXCL4
• lung injury
• sex
• systemic sclerosis