Fascia tissue engineering with human adipose-derived stem cells in a murine model: Implications for pelvic floor reconstruction.

BACKGROUND/PURPOSE: Mesh-augmented vaginal surgery for treatment of pelvic organ prolapse (POP) does not meet patients' needs. This study aims to test the hypothesis that fascia tissue engineering using adipose-derived stem cells (ADSCs) might be a potential therapeutic strategy for reconstructing the pelvic floor. METHODS: Human ADSCs were isolated, differentiated, and characterized in vitro. Both ADSCs and fibroblastic-differentiated ADSCs were used to fabricate tissue-engineered fascia equivalents, which were then transplanted under the back skin of experimental nude mice. RESULTS: ADSCs prepared in our laboratory were characterized as a group of mesenchymal stem cells. In vitro fibroblastic differentiation of ADSCs showed significantly increased gene expression of cellular collagen type I and elastin (p < 0.05) concomitantly with morphological changes. By contrast, ADSCs cultured in control medium did not demonstrate these changes. Both of the engrafted fascia equivalents could be traced up to 12 weeks after transplantation in the subsequent animal study. Furthermore, the histological outcomes differed with a thin (111.0 ± 19.8 µm) lamellar connective tissue or a thick (414.3 ± 114.9 µm) adhesive fibrous tissue formation between the transplantation of ADSCs and fibroblastic-differentiated ADSCs, respectively. Nonetheless, the implantation of a scaffold without cell seeding (the control group) resulted in a thin (102.0 ± 17.1 µm) fibrotic band and tissue contracture. CONCLUSION: Our results suggest the ADSC-seeded implant is better than the implant alone in enhancing tissue regeneration after transplantation. ADSCs with or without fibroblastic differentiation might have a potential but different role in fascia tissue engineering to repair POP in the future.

Tissue-engineered fascia from vaginal fibroblasts for patients needing reconstructive pelvic surgery.

INTRODUCTION AND HYPOTHESIS: Mesh-augmented reconstructive surgery for pelvic organ prolapse (POP) does not meet clinical expectations. A tissue-engineered fascia equivalent needs to be developed. METHODS: Human vaginal fibroblasts (HVFs) from 10 patients were characterized in vitro. Eligible HVFs and a biodegradable scaffold were used to fabricate a fascia equivalent, which was then transplanted in vivo. RESULTS: The cultured HVFs were divided into high (n = 6) or low (n = 4) collagen I/III ratio groups. Cells of the high-ratio group exhibited significantly higher proliferation potential than those of the low-ratio group (P < 0.05). A fascia equivalent was made with HVFs of the high-ratio group. In the subsequent animal study, a well-organized neo-fascia formation containing HVFs could be traced up to 12 weeks after transplantation. CONCLUSIONS: Our results suggest that a tissue-engineered fascia could be developed from HVFs in vitro and in vivo, which might be an effective treatment for POP in the future.

Effect of EGCG, a major component of green tea, on the expression of Ets-1, c-Fos, and c-Jun during angiogenesis in vitro.

In this study, we used rat aortic endothelial cells and human umbilical vein endothelial cells growing in collagen gel as a model system to study the tea catechin, (-)-epigallocatechin (EGCG), on the differential expression of transcription factors, Ets-1, c-Fos, and c-Jun during endothelial morphogenesis in vitro. Cells growing in collagen gel from 0 to 2 h remained spherical. After 6 h, the cells became elongated and underwent morphogenesis. At 24 h, cells started to organize to form capillary-like tubular structures. At 48 h, most cells in the gel formed a network of branching and tubular structures. Immunohistochemistry and immunofluorescence microscopy showed that the reaction products of Ets-1, c-Fos, and c-Jun presented predominantly in the nucleus. No reaction products appeared in the cells that were organized to form capillary-like tubular structures. After adding EGCG to the collagen gel, cells became elongated in the first 6 h and then remained quiescent. No tubular structure was formed. Western blotting showed that the levels of Ets-1, c-Fos, and c-Jun reached the highest levels at 12-24 h, decreasing to the basal level at 48 h. After adding EGCG to the collagen gel, levels were lower than for the non-EGCG-treated groups. These results indicated that the morphogenesis of endothelial cells in collagen gel was inhibited by EGCG through the down-regulation of Ets-1, c-Fos, and c-Jun.

Use of gain-of-function study to delineate the roles of crumbs in Drosophila eye development.

The cell polarity gene, crumbs (crb)), has been shown to participate in the development and degeneration of the Drosophila retina. Mutations in CRB1, the human homologue of Drosophila crb, also result in retinitis pigmentosa and Leber congenital amaurosis. In this study, we used the gain-of-function approach to delineate the roles of CRB in developing Drosophila eye. In the third-instar larval stage, eye development is initiated with photoreceptor differentiation and positioning of photoreceptor nuclei in the apical cellular compartment of retinal epithelium. In the pupal stage, differentiated photoreceptors begin to form the photosensitive structures, the rhabdomeres, at their apical surface. Using GMR-Gal4 to drive overexpression of the Crb protein at the third-instar eye disc, we found that differentiation of photoreceptors was disrupted and the nuclei of differentiated photoreceptors failed to occupy the apical compartment. Using HS-Gal4 to drive Crb overexpression in pupal eyes resulted in interference with extension of the adherens junctions and construction of the rhabdomeres, and these defects were stage-dependent. This gain-of-function study has enabled us to delineate the roles of Crb at selective stages of eye development in Drosophila.