Therapeutic effect of Low intensity Extracorporeal Shock Wave Therapy (Li-ESWT) on diabetic bladder dysfunction in a rat model

Abstract

Objectives: Low intensity extracorporeal shock wave therapy (Li-ESWT) has proven to be effective and safe for the treatment of various urological disorders including erectile dysfunction and chronic pelvic pain syndrome. In this study, we elucidated the therapeutic effect and possible mechanisms of Li-ESWT on diabetic bladder dysfunction (DBD) in a rat model. Materials and Methods: In all, thirty-two female Sprague-Dawley rats were divided into three groups: normal control (NC), diabetes mellitus (DM) control, and DM Li-ESWT. The two DM groups were given high fat diets for one month, followed by 2 intraperitoneal injections of streptozotocin (STZ) 30 mg/kg separated by one week. Body weight and fasting blood glucose were monitored every week. Only rats with fasting blood glucose 140 mg/dL or more were considered diabetic and used in the subsequent portions of the study. The Li-ESWTs were applied toward the pelvis of the rats twice a week for 4 weeks with energy flux density (EFD) 0.02 mJ/mm², 500 shocks, at 3Hz. All rats underwent plasma insulin tolerance test, conscious cystometry, leak-point pressure (LPP) assessment, and immunohistochemical studies. Results: DM groups had significantly lower insulin sensitivity and higher body weight. Conscious cystometry also revealed voiding dysfunctions. In the DM Li-ESWT group, the rats had significantly improved voiding functions that were reflected in longer micturition intervals and higher LPP compared to DM control. Immunofluorescence in DM control groups showed increased tyrosine hydroxylase (TH) expression and decreased neuronal nitric oxide synthase (nNOS) expression in the longitudinal urethral smooth muscles. Besides, rats had dilations and deformities of suburothelium capillary network of the bladder, revealing the deterioration of the nerve function of the urethra and destruction of the vascularization of the bladder. However, the DM Li-ESWT group exhibited recovery of the nerve expression of the urethra and vascularization of bladder. Conclusions: Li-ESWT ameliorates the bladder dysfunction and urinary continence in the DBD rat model, reflected in restoration of the nerve expression of the urethra and the vascularization of the bladder. Non-invasive Li-ESWT could be an alternative therapeutic option for DBD.

Keywords: bladder dysfunction; diabetes mellitus; low intensity extracorporeal shock wave.

Figure 1

Time flow chart. SW: shock wave. STZ: streptozotocin. ITT: insulin tolerance test.

Figure 2

Body weights (A) and insulin tolerance test (B). The body weights were recorded weekly. The insulin tolerance test was performed after 5 weeks of STZ injection. The data are mean ± SD. Normal control group: n = 10; diabetic group: n = 22.

Figure 3

Assessment of voiding function with conscious cystometry. (A) Normal voiding pattern was noted in 9 rats of the control group and 7 rats of the DM+Li-ESWT group. (B) Overactive bladder pattern was noted in one rat of the control group, 6 rats of the DM group and one rat of the DM+Li-ESWT group. (C) Underactive bladder pattern was noted in 2 rats of the DM group and 2 rats of the DM+Li-ESWT group. (D) Acontractile bladder pattern was noted in 2 rats of the DM group and 2 rats of DM+Li-ESWT group, which was also regarded as abnormal voiding pattern. (E) Pie charts of the voiding patterns. Normal voiding patterns were significantly more common (P < 0.05) in the normal control group compared with the DM group. A significant proportion of the DM rats that received Li-ESWT had normal voiding patterns. BP: bladder pressure (cmH2O). AP: abdomen pressure (cmH2O). NBP: net bladder pressure (cmH2O). UW: urine weight (g). DM: diabetes mellitus.

Figure 4

(A) Representative cross sectional images of the urethral smooth muscles stained with Alexa 488 conjugated phalloidin (Pha; green fluorescence) and the sympathetic nerves stained with anti-tyrosine hydroxylase (TH; red fluorescence) antibody. Compared to the control group, urethras in the DM group demonstrated higher TH expression in longitudinal smooth muscles, which was improved after Li-ESWT. (B) Representative cross sectional images of the urethral smooth muscles stained with Alexa 488 conjugated phalloidin (Pha; green fluorescence) and the nitric nerves stained with anti-neuronal nitric oxide synthase (nNOS; red fluorescence) antibody. Compared to the control group, urethras in the DM group showed lower nNOS expression in longitudinal smooth muscles, which was improved after Li-ESWT. (C) Representative images of the bladder suburothelium stained with anti-collagen IV (Col IV; red fluorescence) antibody. Compared to the normal controls, dilations and deformities of suburothelium capillary network were noted in the DM rats, which were improved after Li-ESWT. All the tissue sections were counterstained with 4',6-diamidino-2-phenylindole (DAPI; blue fluorescence) to identify the cell nuclei.