Differentiation of Adipose-Derived Stem Cells into Smooth Muscle Cells in an Internal Anal Sphincter-Targeting Anal Incontinence Rat Model.

Abstract

Studies on development of an anal incontinence (AI) model targeting smooth muscle cells (SMCs) of the internal anal sphincter (IAS) have not been reported. The differentiation of implanted human adipose-derived stem cells (hADScs) into SMCs in an IAS-targeting AI model has also not been demonstrated. We aimed to develop an IAS-targeting AI animal model and to determine the differentiation of hADScs into SMCs in an established model. The IAS-targeting AI model was developed by inducing cryoinjury at the inner side of the muscular layer via posterior intersphincteric dissection in Sprague-Dawley rats. Dil-stained hADScs were implanted at the IAS injury site. Multiple markers for SMCs were used to confirm molecular changes before and after cell implantation. Analyses were performed using H&E, immunofluorescence, Masson's trichrome staining, and quantitative RT-PCR. Impaired smooth muscle layers accompanying other intact layers were identified in the cryoinjury group. Specific SMC markers, including SM22α, calponin, caldesmon, SMMHC, smoothelin, and SDF-1 were significantly decreased in the cryoinjured group compared with levels in the control group. However, CoL1A1 was increased significantly in the cryoinjured group. In the hADSc-treated group, higher levels of SMMHC, smoothelin, SM22α, and α-SMA were observed at two weeks after implantation than at one week after implantation. Cell tracking revealed that Dil-stained cells were located at the site of augmented SMCs. This study first demonstrated that implanted hADSc restored impaired SMCs at the injury site, showing stem cell fate corresponding to the established IAS-specific AI model.