Jessica Hannick

The Role of Genetically Modified Mesenchymal Stem Cells in Urinary Bladder Regeneration.

Recent studies have demonstrated that mesenchymal stem cells (MSCs) combined with CD34+ hematopoietic/stem progenitor cells (HSPCs) can function as surrogate urinary bladder cells to synergistically promote multi-faceted bladder tissue regeneration. However, the molecular pathways governing these events are unknown. The pleiotropic effects of Wnt5a and Cyr61 are known to affect aspects of hematopoiesis, angiogenesis, and muscle and nerve regeneration. Within this study, the effects of Cyr61 and Wnt5a on bladder tissue regeneration were evaluated by grafting scaffolds containing modified human bone marrow derived MSCs. These cell lines were engineered to independently over-express Wnt5a or Cyr61, or to exhibit reduced expression of Cyr61 within the context of a nude rat bladder augmentation model. At 4 weeks post-surgery, data demonstrated increased vessel number (~250 vs ~109 vessels/mm2) and bladder smooth muscle content (~42% vs ~36%) in Cyr61OX (over-expressing) vs Cyr61KD (knock-down) groups. Muscle content decreased to ~25% at 10 weeks in Cyr61KD groups. Wnt5aOX resulted in high numbers of vessels and muscle content (~206 vessels/mm2 and ~51%, respectively) at 4 weeks. Over-expressing cell constructs resulted in peripheral nerve regeneration while Cyr61KD animals were devoid of peripheral nerve regeneration at 4 weeks. At 10 weeks post-grafting, peripheral nerve regeneration was at a minimal level for both Cyr61OX and Wnt5aOX cell lines. Blood vessel and bladder functionality were evident at both time-points in all animals. Results from this study indicate that MSC-based Cyr61OX and Wnt5aOX cell lines play pivotal roles with regards to increasing the levels of functional vasculature, influencing muscle regeneration, and the regeneration of peripheral nerves in a model of bladder augmentation. Wnt5aOX constructs closely approximated the outcomes previously observed with the co-transplantation of MSCs with CD34+ HSPCs and may be specifically targeted as an alternate means to achieve functional bladder regeneration.

Trauma to the Pelvis: Injuries to the Rectum and Genitourinary Organs

Pelvic trauma is associated with high mortality rates. Blunt pelvic injuries from high-energy mechanisms are often associated with pelvic fractures and injuries to the rectum and genitourinary (GU) tract. In addition, due to close anatomic proximity, penetrating pelvic trauma can injure the bony pelvis, rectum, and GU tract concomitantly. As a result, the assessment and management of pelvic trauma requires a multifaceted approach involving orthopedics, trauma surgery and urology.

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

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.