The Effect Of Bone Marrow Mesenchymal Stem Cells On Rat Chronic Prostatitis And Its Mechanisms

Backgrounds:About 90% of men have suffered recessive or dominant inflammatory diseases of urinary system in worldwide.In 1995,NIH divided prostatitis into four types.Only I(ABP)and II(CBP) type have clear etiology.Past research has shown that prostatitis is one of the most factors which induce benign prostatic hyperplasia and prostate cancer, therefore its prevention gets more and more attention. Because of the complexity of the clinical etiology of chronic prostatitis and the existing of prostate barrier,antibiotics,the core treatment of prostatitis,often can not obtain ideal effect.The prostatitis defers and repeats.Early we observed that there were abundant inflammatory cells infiltrating in the prostate gland of the rat model with chronic bacterial prostatitis and the inflammatory factors,such as TNF-α、IL-1β and IL-6,secreting by these cells,are the dynamic factors in the development of inflammation. After two to four weeks under antibiotic treatment, there is no bacterial in the prostate gland in part of rats, but inflammatory cells still exsiting. Antibiotics only play the role of microbial resistance, lack of immune inflammation curative effect. Therefore new drugs or new treatment are needed for prostatitis treatment.Bone marrow mesenchymal stem cells is a stem cell population with rapid proliferation,multi-directional differentiation, low immunogenicity, immune suppression and tissue repair ability. In the past ten years, BMSCs has been applied in clinical treatment. First BMSCs appeared in the treatment of osteogenesis, then in lysosomal storage disorder, graft versus host disease, Crohn and in the treatment of inflammatory bowel disease. Earlier studies have shown that MSCs play the role of inhibiting the inflammatory response through inhibiting T cells, B cells, antigens presented cell proliferation and inhibiting proinflammatory factor. In recent years, more new progress has been made in the study about the MSCs in immunosuppression. In the gram-negative model induced by bacteria, Elman found that MSCs can improve the survival rate of mice through enhancing the phagocytosis of monocytes activity. Intravenous injected to bronchial asthma rats, Nemeth found MSCs can be specificity gathered within the lung inflammation after 1 hour and reduce the immune inflammatory response by secreting TGF – beta,which can release Th2 related inflammation factors in the bronchial lavage flui,and the levels of Ig G1, IgE in serum. Recent study shows that MSCs can suppress and eliminate bacteria by secreting some cytokines and antibacterial peptides.In the treatment of pneumonia induced by E. coli, Gupta, Lee, Krasnodembakaya found MSCscan reduce pulmonary inflammatory response by dripping MSCs to the endotracheal. The mechanism is keratinocyte growth factor secreted by MSCs enhance the phagocytosis ability of monocytes,and antimicrobial peptides LL-37, apolipoprotein-2 secreted by MSCs directly remove the E. coli. TheSE studies show that BMSCs has good treatment effect on inflammatory disease.However, whether BMSCs can inhibit chronic prostatitis is unclear.Ultrasonic is widely used in clinical diagnosis. As a kind of energy, ultrasonic is also be used in the treatment with a "mechanical effect, thermal effect, cavitation" effect.When cavitation effects, Micro jet, shock wave and other mechanical energy can increased permeability of cell membrane, opening close connection between the cells. Injected into the veins, micro bubble can enhance the ultrasonic cavitation effect and biological effect through shocking cyclical and implosion under ultrasonic irradiation. Our team’s early experiment results have confirmed the blood- prostate barrier to hinder the drugs into the prostatic stroma. Close connection between capillary endothelial cells in blood participates in the barrier of the prostate. Blood- prostate barrier also hinders the BMSCs into the prostate. Ultrasonic composite micro bubble can open blood- prostate barrier,However, whether can promote BMSCs to prostate homing needs further research.Objectives:Bone marrow adherent culture method is using in BMSCs isolated, cultured and amplification. chronic prostatitis rat model is established by injecting e. coli into prostate under ultrasound-guided. After ultrasound- microbubble facilitate the BMSC homing to the site of inflammation, BMSCs play the role of inhibiting the inflammatory response. This study is expected to broaden the knowledge of BMSCs immune regulation mechanism, and provides new strategy for clinical prevention and control of chronic prostatitis.Methods:1. Adherent culture method is using in BMSCs isolated,cultured and amplification with 3 weeks male SD rats.Identify the cell surface markers and differentiation. GFP-LUC adenovirus transfects and mark BMSCs. Cell proliferation characteristics after transfecting will be observing.2. E.coli will be injected into prostate of SD rats,220-250 G, under ultrasound guided,to establish chronic prostatitis rat model. The rat’s survival and biological behavior will be observing.3. Ultrasound- microbubble will be using to facilitate the BMSC, tail intravenous, homing to the site of inflammation. Testing the level of related inflammatory factor and observing the improvement of the pathological morphology.Results:1. Rat bone marrow derived MSCs have been isolated,cultured and amplification by adherent culture method.BMSCs have proliferative and mulineage differentiation potential,The proliferation activity unchanged.2. Chronic prostatitis rat model has been established successfully by injection of bacterial under ultrasound-guided.There is e.coli in the prostate. Symptoms of frequent urination happen in rats and no death has found.3. Ultrasound- microbubble can facilitate the BMSC homing to the site of inflammation. Histopathological evalution showed that typical pathological inflammation in the prostates were observed in CBP groups, CBP+BMSCs group and CBP+MEUS group,including the change of gland structure,interstitial edema,inflammatory cells infiltration and fibrous hyperplasia,while pathological changes were relieved obviously in CBP+MEUS+BMSCs group(P<0.01). The mRNA expression of IL-1β、TNF-α in CBP group,CBP+BMSCs group,CBP+MEUS group were higher than those in control group and CBP+MEUS+BMSCs group(P<0.05).The protein content of IL-1β、TNF-α in CBP group,CBP+BMSCs group,CBP+MEUS group were higher than those in control group(P<0.01),while there is no statistical difference between CBP+MEUS+BMSCs group and control group(P>0.05)Conclusions:1. BMSCs have been isolated,cultured and amplification successfully by adherent culture method,which lays the foundation for the next experiment.2. The method of establishing model by injecting e.coli unde ultrasound results less trauma, complications, lower mortality than traditional method.3. BMSC can home to the site of inflammation facilitated by Ultrasoundmicrobubble,and can inhibit inflammation of prostate. Relevant mechanism needs further research.