Basic Research Of The Treatment For Diabetic Cystopathy With NGF/P407 & Adipose Tissue—Derived Stem Cells

ObjectiveOur goals are to create a rat model of DCP and to investigate the function and mechanism of NGF/P407 & ADSCs in treating DCP.MethodsForty-five male Sprague—Dawley rats(12 weeks old), forty rats were randomly divided into five groups:one of the groups is control group (N group, n=8), fed with regular food. The other groups are to create a rat model of DM Ⅱ:fed with high fat diet(HFD)for 1 month, followed by two intraperitoneal injections of streptozotocin(STZ,30mg/kg). HFD was given until the end of study. Three months after STZ injection, monitored fasting blood sugar for every rats. And then adipose tissue was harvested via laparotomy to isolate and culture ADSC.ADSC were labeled with 10 uM 5-ethynyl-2-deoxyuridine(EdU).Prepared the NGF/P407. The 32 diabetic rats in Diabetic group were randomly divided into four groups: DM group(n=8):the rats received injection of phosphate buffered saline(PBS)1 ml into the bladder wall via laparotomy; A group(n=8):the rats received injection of NGF/P407 1 ml into the bladder wall via laparotomy; B group(n=8):the rats received 3x106 autologous ADSC in 1 ml PBS by injection into the bladder wall via laparotomy. C group(n=8):the rats received 3x106 autologous ADSCs and NGF/P407 in 1 ml PBS by injection into the bladder wall via laparotomy. One month after injection, all rats were performed conscious cystometry and were killed for histology examination. Including Immunofluorescence and Immunohistochemical staining for TUNEL, nerve distribution and tracking EdU,Masson, quantification of vascularity,and western blot for analysis of NGF in the bladders.ResultsRats in the diabetic group had higher mean body weight, significantly higher plasma glucose level. All rats in the N group manifested a normal voiding pattern, the numbers of normal voiding patterns in A/B groups, significantly more than DM group.and C group significantly more than A/B group. For tracking of EdU—positive cells,some positive nuclei appeared to be colocalized with SMA staining. Apoptotic cells were significantly more in DM group compared to N group. After treatments, apoptosis became significantly less, and C group significantly more than A/B group. Compared to DM group, CD31 staining was intensely positive in the submucosal layer of N/A/B/C groups. The level of NGF in A/C groups were significantly more than other groups.ConclusionBoth of NGF/P407 and ADSC could enhance the regeneration process of diabetic bladder dysfunction, ADCS could not only differentiate into smooth muscle cells, but also inhibit apoptosis and improve vascularity. While the NGF/P407 can improve the level of NGF and the receptors of Trk A and p75NTR. NGF/P407 combined with ADSC to treat diabetic cystopathy is more effective.