Cationic star-shaped polymer as an siRNA carrier for reducing MMP-9 expression in skin fibroblast cells and promoting wound healing in diabetic rats

Abstract

Background

Excessive expression of matrix metalloproteinase-9 (MMP-9) is deleterious to the cutaneous wound-healing process in the context of diabetes. The aim of the present study was to explore whether a cationic star-shaped polymer consisting of β-cyclodextrin (β-CD) core and poly(amidoamine) dendron arms (β-CD-[D ₃] ₇) could be used as the gene carrier of small interfering RNA (siRNA) to reduce MMP-9 expression for enhanced diabetic wound healing.

Methods

The cytotoxicity of β-CD-(D ₃) ₇ was investigated by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay (MMT) method in the rat CRL1213 skin fibroblast cell line. The transfection efficiency of β-CD-(D ₃) ₇/MMP-9-small interfering RNA (siRNA) complexes was determined by confocal microscopy and flow cytometry. Quantitative real time (RT) polymerase chain reaction was performed to measure the gene expression of MMP-9 after the transfection by β-CD-(D ₃) ₇/MMP-9-siRNA complexes. The β-CD-(D ₃) ₇/MMP-9-siRNA complexes were injected on the wounds of streptozocin-induced diabetic rats. Wound closure was measured on days 4 and 7 post-wounding.

Results

β-CD-(D ₃) ₇ exhibited low cytotoxicity in fibroblast cells, and easily formed the complexes with MMP-9-siRNA. The β-CD-(D ₃) ₇/MMP-9-siRNA complexes were readily taken up by fibroblast cells, resulting in the downregulation of MMP-9 gene expression ( P<0.01). Animal experiments revealed that the treatment by β-CD-(D ₃) ₇/MMP-9-siRNA complexes enhanced wound closure in diabetic rats on day 7 post-wounding ( P<0.05).

Conclusion

β-CD-(D ₃) ₇ may be used as an efficient carrier for the delivery of MMP-9-siRNA to reduce MMP-9 expression in skin fibroblast cells and promote wound healing in diabetic rats.