Evaluation of the antifibrotic potency by knocking down SPARC, CCR2 and SMAD3

BACKGROUND: The genes of SPARC, CCR2, and SMAD3 are implicated in orchestrating inflammatory response that leads to fibrosis in scleroderma and other fibrotic disorders. The aim of the studies is to evaluate synergistic anti-fibrotic potency of the siRNAs of these genes. METHODS: The efficacy of the siRNA-combination was evaluated in bleomycin-induced mouse fibrosis. The pathological changes of skin and lungs of the mice were assessed by hematoxylin and eosin and Masson's trichrome stains. The expression of inflammation and fibrosis associated genes and proteins in the tissues were assessed by real-time RT-PCR, RNA sequencing, Western blots and ELISA. Non-crosslinked fibrillar collagen was measured by the Sircol colorimetric assay. FINDINGS: The applications of the combined siRNAs in bleomycin-induced mice achieved favorable anti-inflammatory and anti-fibrotic effects. Activation of fibroblasts was suppressed in parallel with inhibition of inflammation evidenced by reduced inflammatory cells and proinflammatory cytokines in the BALF and/or the tissues by the treatment. Aberrant expression of the genes normally expressed in fibroblasts, monocytes/ macrophage, endothelial and epithelial cells were significantly restrained after the treatment. In addition, transcriptome profiles indicated that some bleomycin-induced alterations of multiple biological pathways were recovered to varying degrees by the treatment. INTERPRETATION: The application of the combined siRNAs of SPARC, CCR2, and SMAD3 genes ameliorated inflammation and fibrosis in bleomycin-induced mice. It systemically reinstated multiple biopathways, probably through controlling on different cell types including fibroblasts, monocytes/macrophages, endothelial cells and others. The multi-target-combined therapeutic approach examined herein may represent a novel and effective therapy for fibrosis.