Identification Of Ryanodine Receptor Subtypes On Bladder Detrusor Muscle Cells Of Patients With Benign Prostatic Hyperplasia

ObjectiveBenign prostatic hyperplasia ( BPH) is common in aged men, which will make the outlet of bladder stricture and obstruction, and results in clinical symptom. With the increasing of urinary resistance and chronic urinary reten-sion, bladder detrusor may be injured to be dysfunction, and lose the ability of contract ultimately. Previous studies have showed that detrusor muscle was contracted by mechanism of Ca²⁺ - induced Ca²⁺ release ( CICR) , which showed that Ca²⁺ played an important role in excitation - contraction coupling on bladder detrusor muscle. Besides the Ca²⁺ channels existed in cells membrane which induced calcium influx from extracellular medium, there were two kinds of Ca²⁺ channels in intracellular stores which induced calcium released from intracellular stores, Ryanodine receptor( RyR) was one of them.RyR is one kind of Ca²⁺ channels existed in endoplasmic reticulum (ER) / sarcoplasmic reticulum ( SR) of cells, which is composed of four subunits, and its higher structure is tetraploid. At present, three kinds of RyR subtypes have been separated: skeletal muscle ( RyR1 ) , cardiac muscle ( RyR2 ) and brain (RyR3). With the progress of researches, it has been found that some tissue/ cells did not only express one subtype, but also expressed the other one or two subtypes. At the moment, it is not clear about the function and peculiarity of all kinds of RyR subtypes. However, overall kinds of RyR subtypes have a common character: CICR, which may enlarge Ca²⁺ signal to lead to relevant biologicaleffect by this kind of positive feed - back.Some studies showed that the change of RyR function and peculiarity took part in many pathological progresses, and the mechanism of CICR induced by RyR would change a lot. Therefore, defining RyR suvtypes of special tissue/ cells is important to research the progress of Ca²⁺ signal transduction and the relationship with disease. This study identified the RyR subtypes of bladder detru-sor muscle cells on normal human beings and the patients with BPH by RT -PCR and Western blot, explored the change of bladder detrusor which caused by BPH initially, confirmed the mechanism of BPH more, and provided some valuable data about treatment and judgement of prognosis for BPH.MethodsThis study identified the RyR subtypes of bladder detrusor muscle cells on normal human beings and the patients with BPH by RT - PCR and Western blot. The specimen was bladder detrusor muscle of the patients who received operations in Urology of China Medical University Affiliated No. 1 Hospital from Nov. 2003 to May.2004. It was divided into two groups; test group is composed of 20 cases, from 60 - 91yr ( mean 71. 6yr) who received suprapubic transvesical prostatectomy for BPH; control group is also composed of 20 cases, from 40 -79yr( mean 56. 3yr) who received partial cystectomy for bladder tumor.1. To identify the RyR subtypes that bladder detrusor muscle cells expressed on normal human beings and the patients with BPH by RT - PCR: extracting total RNA of specimen, synthesizing cDNA by reverse transcription ( RT) , amplifying cDNA by PCR, undertaking agar gel electrophoresis, staining and photographing.2. To identify the RyR subtypes that bladder detrusor muscle cells expressed on normal human beings and the patients with BPH by Western blot: extracting total protein of specimen, determining the density of protein by Lowrys method, resolving the protein by SDS - PAGE, transfering to nitrocellulose ( NC) membrane, incubating, staining and photographing.