Expression Of P2X Receptor Subtypes In Bladders Of Bladder Outlet Obstruction Rats

PrefaceBladder outlet obstruction is one of the common diseases in clinic, and its main harm is that it can cause bladder dysfunction, arise overactive bladder, and lead to voiding frequency, urgency, urinary retentionalso, incontinence and even upper urinary tract damage in serious cases. P2X receptor which was discovered recently is a purinergic receptor gated by ATP ion channel. At present, seven P2X receptor subtypes are discovered. Members of the P2X receptor family are widely distributed in the central and peripheral nervous system, and they play important roles in both sensory and motor functions of the bladder. In this study, rats of partial bladder outlet obstruction are created by surgical proximal urethral ligation, and are grouped according to the duration of obstruction. The mRNA expression of each P2X receptor subtype is detected by reverse transcriptive polymerase chain reaction(RT-PCR) in the bladders of both normal and BOO rats. The role of P2X receptors on vesical contraction and relaxation and the relation with bladder dysfunction is explored, which might provide new ideas for the clinical therapy of bladder dysfunction after bladder outlet obstruction.Material and Methods1. Experimental material1.1 Experimental animals: female Wistar rats of 3-4 months old and each weight is about 250g.1.2 Main experimental reagents and primers: total RNA separating reagents, RT-PCR kit, DNA polymerase and gene mRNA primers of P2X₁～P2X₇ receptor subtypes.1.3 Main experimental instruments: ophthalmic operating sets, ultraviolet spectrophotometer, PCR amplification apparalus, desk-top low temperature high speed centrifugal machine, level electrophoresis bath, electrophoresis apparalus and automatic electrophoresis gelamm image analysis system.2. Experimental methods2.1 Establish the rats model of bladder outlet obstruction.2.2 Detect the mRNA expression of P2X receptor subtypes in the bladders by reverse transcriptive polymerase chain reaction(RT-PCR).2.3 statistical analysisAll datum were expressed as the means±SD, the t test was carried out using SPSS statistical software.Results1. Results of the animal modelsTen rats from control group were all alive, while seven were alive from two-week BOO group and seven were alive from four-week BOO group. All the dead rats had hydronephrosis. Among the alive rats that were from the BOO two-week and the BOO four-week group, the number of which had hydronephrosis was two and five respectively.2. Results of RT-PCRThe mRNA expression of each P2X receptor subtype(P2X_1-7) was detected in the bladders of both normal and BOO rats. Expression levels of P2X₁ and P2X₄ were correspondingly high in the bladders of normal of rats, while in the BOO rats, the expression levels of P2X₁, P2X₄, P2X₆ and P2X₇ were decreased and of P2X₂ and P2X₃ were increased. Expression level of P2X₆ in each group was very low and it had no distinct change. Compared with control group, P2X₁ decreased and P2X₂ increased significantly in two-week BOO group, but the rest of P2X receptor subtypes changed little. The differences of control group and BOO four-week group were all significant.DiscussionBladder outlet obstruction is the common diseases in clinic, and it is is one of the main reasons that lead to overactive bladder. Recent researches done abroad indicate that in addition to the cholinergic and adrenergic nerves, the non-adrenergic non-cholinergic nerve is another important factor which can participate in the functional accommodation of bladder. However, the relative researches are deficient now.P2X receptors are purinergic receptors discovered in recent years, belonging to ATP- gated ion channel receptors. At present, there are seven known P2X receptor subtypes, named P2X_1-7. These receptors are widely distributed in the tissues all over the body, and they are also expressed in the central and peripheral nervous system. They play important physiological and pathophysiological roles in a variety of biological processes, and chiefly participate in accommodating vesical contraction and relaxation.The study revealed that the functions of the bladder were damaged, and P2X receptor subtypes changed after bladder outlet obstruction. Besides, the P2X₂ and P2X₃ receptors displaced P2X₁ and P2X₄, and they became chief receptors accommodating vesical functions after bladder outlet obstruction. With the increase of the duration of obstruction, the damages of bladder functions and upper urinary tract aggravated, and the changes of P2X receptor subtypes were more evident. It was supposed that the changes of P2X receptor subtypes may be a critical factor in development of bladder dysfunction after bladder outlet obstruction.ConclusionChanges of mRNA of some P2X receptor subtypes in BOO rats may play an important role in the pathogenic mechanisms of bladder dysfunction after bladder outlet obstruction.