Characterization of bladder and external urethral activity in mice with or without spinal cord injury--a comparison study with rats.

To clarify the lower urinary tract function in mice, we compared bladder and urethral activity between rats and mice with or without spinal cord injury (SCI). Female Sprague-Dawley rats and C57BL/6N mice were divided into five groups:1) spinal intact (SI) rats,2) SI mice,3) pudendal nerve transection (PNT) SI mice,4) spinal cord injury (SCI) rats, and 5) SCI mice. Continuous cystometry (CMG) and external urethral sphincter (EUS)-electromyogram (EMG) analyses were conducted under an awake, restrained condition. During voiding bladder contractions, SI animals exhibited EUS bursting with alternating active and silent periods, which, in rats but not mice, coincided with small-amplitude intravesical pressure oscillations in CMG recordings. In SI mice with bursting-like EUS activity, the duration of active periods was significantly shorter by 46% (32 ± 5 ms) compared with SI rats (59 ± 9 ms). In PNT-SI mice, there were no significant differences in any of cystometric parameters compared with SI mice. In SCI rats, fluid elimination from the urethra and the EUS bursting occurred during small-amplitude intravesical pressure oscillations. However, SCI mice did not exhibit clear EUS bursting activity or intravesical pressure oscillations but rather exhibited intermittent voiding with slow large-amplitude reductions in intravesical pressure, which occurred during periods of reduced EUS activity. These results indicate that EUS pumping activity is essential for generating efficient voiding in rats with or without spinal cord injury. However, EUS bursting activity is not required for efficient voiding in SI mice and does not reemerge in SCI mice in which inefficient voiding occurs during periods of reduced tonic EUS activity.

Autologous and heterotopic transplantation of adipose stromal vascular fraction ameliorates stress urinary incontinence in rats with simulated childbirth trauma.

INTRODUCTION: Autologous transplantation of adipose stromal vascular fraction (SVF) is a cost-effective and technically accessible option for cell therapy. Clinical study of SVF transplantation for male stress urinary incontinence (SUI) is underway, but the effectiveness remains unknown for female SUI, majority of which is caused by childbirth trauma. METHODS: Vaginal Distension (VD) rats were generated as in vivo model for female SUI. To quantitate the severity of SUI, leak point pressure (LPP) was measured by placing a bladder catheter. There was a characteristic waveform of LPP with two-peaks, and we counted the second peak as an LPP value. Adipose SVF was separated from inguinal fat and delivered into external urethral sphincter (EUS) through transperineal injection. LPP was measured 7 or 14 days after SVF transplantation. Tissue damage and collagen synthesis around the EUS were visualized by Masson's trichrome and eosin staining. Antibody against α-smooth muscle actin (α-SMA) was used to stain smooth muscle or activated stromal cells. Donor SVF cells were distinguished from recipient EUS tissue by tracking with GFP transgene. RESULTS: VD procedure decreased the frequency at which the normal LPP waveform appeared and lowered the LPP value. SVF injection normalized the waveform as well as the level of LPP. VD disrupted histological structure of EUS and SVF failed to differentiate into striatal muscles. Instead, SVF increased α-SMA positive cells and collagen synthesis but the phenomena depended on VD stimulus. GFP tracking indicated that the transplanted SVF cells persisted for four weeks and synthesized α-SMA protein simultaneously. CONCLUSIONS: Autologous transplantation of adipose SVF displayed bulking effects through collagen synthesis. However, such heterotopic activation was dependent on tissue damage.

Tissue engineered bulking agent with adipose-derived stem cells and silk fibroin microspheres for the treatment of intrinsic urethral sphincter deficiency.

In this study we developed a tissue engineered bulking agent that consisted of adipose-derived stem cells (ADSCs) and silk fibroin microspheres to treat stress urinary incontinence caused by severe intrinsic sphincter deficiency (ISD). ISD models were established by completely transection of the bilateral pudendal nerve (PNT) and confirmed by the decreased leak-point pressure (LPP) and increased lumen area of urethra. Injection of silk fibroin microspheres could recover LPP and lumen area at 4 weeks but its efficacy disappears at 8, 12 weeks. Moreover, it was exciting to find that tissue engineered bulking agent brought long-term efficacy (at 4, 8, 12 weeks post-injection) on the recovery of LPP and lumen area. Concomitantly with the function, tissue engineered bulking agent treated group also improved the urethral sphincter structure as exhibited by better tissue regeneration. The findings showed that silk fibroin microspheres alone could work effectively in short-term, while tissue engineered bulking agent that combined silk fibroin microspheres with ADSCs exhibited promising long-term efficacy. This study developed a new strategy of tissue engineered bulking agent for future ISD therapy.

Transplantation of muscle-derived stem cells plus biodegradable fibrin glue restores the urethral sphincter in a pudendal nerve-transected rat model

We investigated whether fibrin glue (FG) could promote urethral sphincter restoration in muscle-derived stem cell (MDSC)-based injection therapies in a pudendal nerve-transected (PNT) rat, which was used as a stress urinary incontinence (SUI) model. MDSCs were purified from the gastrocnemius muscles of 4-week-old inbred female SPF Wistar rats and labeled with green fluorescent protein. Animals were divided into five groups (N = 15): sham (S), PNT (D), PNT+FG injection (F), PNT+MDSC injection (M), and PNT+MDSC+FG injection (FM). Each group was subdivided into 1- and 4-week groups. One and 4 weeks after injection into the proximal urethra, leak point pressure (LPP) was measured to assess urethral resistance function. Histology and immunohistochemistry were performed 4 weeks after injection. LPP was increased significantly in FM and M animals after implantation compared to group D (P < 0.01), but was not different from group S. LPP was slightly higher in the FM group than in the M group but there was no significant difference between them at different times. Histological and immunohistochemical examination demonstrated increased numbers of surviving MDSCs (109 ± 19 vs 82 ± 11/hpf, P = 0.026), increased muscle/collagen ratio (0.40 ± 0.02 vs 0.34 ± 0.02, P = 0.044), as well as increased microvessel density (16.9 ± 0.6 vs 14.1 ± 0.4/hpf, P = 0.001) at the injection sites in FM compared to M animals. Fibrin glue may potentially improve the action of transplanted MDSCs to restore the histology and function of the urethral sphincter in a SUI rat model. Injection of MDSCs with fibrin glue may provide a novel cellular therapy method for SUI.