The Study Of Bladder Structural Remodeling And Dysfunction Caused By Lower Urinary Tract Obstruction

Objective: The bladder dysfunction caused by lower urinary tract obstruction is a difficult of treatment and research focus. The current study on the role of nitric oxide synthase (NOS) and nitric oxide (NO) in normal and obstructive bladder is controversial.This study was designed to investigate the relation of NOS subtypes in rat partial bladder outlet obstruction (PBOO) model resulted in structural remodeling and dysfunction,and oxidative stress damage,the feasibility of NOS as bladder dysfunction therapeutic targets.Methods:1. 24 SD male rats were randomly divided into four groups: normal control group, sham operation control group, partial bladder outlet obstruction persisted for 3 weeks (PBOO3W) and 6 weeks (PBOO6W). PBOO model were established by the urethral outside-stent.The rats underwent cystometry, bladder weighting, muscle collagen staining. Bladder structural remodeling and dysfunction caused by obstruction was objective reflected by above morphological and functional index.2. Real-time quantitative PCR (Q-PCR) detected NOS mRNA in the obstructive bladder tissue, immunohistochemical stain located NOS express,western blotting detected NOS monomer and dimer, spectrophotometer detected NOS subtype activity and oxidative stress levels.To explain the relationship of NOS,dysfunction and structural remodeling,oxidative stress damage in the obstructive bladder.3. NOS cofactor tetrahydrobiopterin (BH4), NO donor L-arginine (L-arg), anti-oxidants Vit-C was given to PBOO3W rats and persisted 3 weeks.Then to evaluate these groups bladder function, morphology, molecular markers,and to objective prove NOS as a protective therapeutic target of obstructive bladder is feasible.Results:1. Compared with control groups, obstructed groups bladder weight and bladder weight/body weight significantly increased (P <0.05);Masson staining demonstrated obstructed bladder muscle hypertrophy and collagen deposition increasing;obstruction caused significant maximal voiding pressure decreasing and residual volume increasing (P <0.05),voiding volume and voiding interval increasing, and even caused filling voiding in PBOO6W group.2. Neuronal NOS (nNOS) was not detected in bladder tissue by Q-PCR, immunohistochemistry and western blot.Inducible NOS (iNOS), endothelial NOS (eNOS) mRNA expression decreased in obstructed bladder;denatured iNOS and eNOS protein monomer decreased in obstructed groups but not significantly different compared with control groups(P> 0.05),non-denatured eNOS monomer/(monomer + dimer) significantly increased in obstructed groups compared with control groups (P <0.05),that eNOS protein uncoupling and inactivation was increasing. Malondialdehyde (MDA) levels significantly increased,superoxide dismutase (SOD) activity decreased significantly(P<0.05) in obstructed groups, that mean obstruction caused increasing oxidative stress damage in bladder tissues.3. Compared with PBOO6W, L-arg, Vit-C groups, BH4 intervention led to the bladder weight, bladder weight/body weight significantly decrease(P<0.05),less muscle hypertrophy collagen deposition,residual urine volume decrease,maximal voiding pressure and voiding volume significantly increase(P<0.05),eNOS monomer/(monomer + dimer) significantly lower (P<0.05), that eNOS recoupling was increasing, less oxidative stress damage(P<0.05);Vit-C group only led to oxidative stress damage mitigation; L-arg and PBOO6W groups were similar.Conclusion: The method of urethral outside-stent could successfully cause rat bladder dysfunction, structural remodeling and had better repeatability and stability. PBOO led to bladder iNOS and eNOS expression tended to decrease, but eNOS uncoupling and inactivating played a major role in structural remodeling,dysfunction and oxidative stress injury in obstructed bladder. eNOS cofactor BH4 can recouple inactivated uncoupling eNOS and protect the obstructive dysfunctional bladder.