MP19-03 THE ANGIOGENIC SIGNALING MOLECULE CYR61 INDUCES INCREASED NEO-VASCULARIZATION IN REGENERATED BLADDER TISSUE

Abstract

INTRODUCTION AND OBJECTIVES: Bone marrow mesenchymal stem cells (BMMSCs) are a promising alternative cell source in bladder tissue engineering especially for improving tissue angiogenesis. Obstacles remain concerning stimulation and persistence of angiogenic vessels during bladder regeneration. The pleiotropic effects of CYR61 manifest in the regulation of inflammation, tissue repair, and angiogenesis. Here the effects of CYR61 on bladder tissue regeneration are evaluated by grafting scaffolds seeded with modified human BMMSCs that either overexpress (OX) or have limited expression of CYR61 in a nude rat bladder augmentation model. METHODS: Human BMMSCs were modified to either OX CYR61 or limit CYR61 expression by gene knockdown (KD) utilizing small interfering RNA constructs. Western blot confirmed levels of protein expression. Modified BMMSCs were seeded at 1.5 10 cells/cm onto poly (1,8-octanediol-co-citrate) (POC) scaffolds 7e8 days prior to use. Urodynamics (UDS) were obtained followed by a 50e60% partial cystectomy with bladder augmentation using the cell/scaffold composites in nude rats (n1⁄45 per group). At sacrifice (4 and 10 weeks) animals underwent repeat UDS, capillarioscopy, and harvest of augmented bladders. Vessel characteristics and muscle content were quantified with Trichrome stain. Peripheral nerve regeneration was quantified with neuron specific b III tubulin immunofluorescence staining. RESULTS: At 4 weeks, CYR61 OX, as compared to KD, resulted in significantly increased vessel number (249.9 22.3 vs. 108.8 5.5 vessels/mm, p<0.001) and muscle content (42.3 1.3% vs. 36.1 1.6%, p<0.05). Previously published data from our laboratory has shown far fewer vessels (POC 63.8 5.4, unmanipulated MSCs 83.4 15.8 vessels/mm) and decreased muscle content in 4 week controls (POC 9.3 1.9%, unmanipulated MSCs 38.4 1%). CYR61 KD demonstrated significant loss of muscle content from 36.1 1.6% at 4 weeks down to 25.0 2.7% at 10 weeks (p<0.05). At 4 and 10 weeks, capillariscopy and UDS demonstrated functional bladders and capillaries in all animals. At 4 weeks CYR61 OX resulted in primitive nerve in-growth of 664.1 87.9 mm into regenerated tissue (mean length 39.3 5.9 mm). No nerve elements were noted in CYR61 KDs. CONCLUSIONS: CYR61 is a potent extracellular signaling molecule that increases functional vasculature, preserves muscle content from 4 to 10 weeks, and induces the growth of neural elements at 4 weeks in regenerated bladder tissue.