Endoscopic implantation of autologous myoblasts for stress urinary incontinence and evaluation of its efficacy in sphincterotomized rabbits.

INTRODUCTION: Stress urinary incontinence (SUI) is one of the health problems with more impact on patients' lives. The aim of the present work was to develop a therapy for SUI using tissue engineering by isolation and culture of autologous myoblasts (CAM) followed by endoscopic implantation. We also evaluated the efficacy of this therapy in a rabbit model of incontinence after sphincterotomy. MATERIALS AND METHODS: We used healthy male New Zealand rabbits. The animals were first bled to obtain platelet-poor plasma (PPP) and biopsied for myoblast isolation. Post-sphincterotomy, they were divided into two groups: the treatment group (including animals that received CAM resuspended in PPP) and the control group (including animals receiving only PPP). The leak-point pressure (LPP) was used to measure continence in both groups at different time points. The results were evaluated with hierarchical linear regression models. Histological evaluation of the rabbits' sphincters was also performed at the end of follow-up. RESULTS: No statistically significant differences were observed between the baseline LPP values of each group. The post-sphincterotomy values of both groups were below 50% of the baseline value, which was a mandatory condition for incontinence. The post-implantation values of the treatment group were higher than 50% of the baseline value, which led us to assume continence recovery. A statistically significant difference was observed in the LPP values between the two treatment groups (p=0.003). Histological study revealed interconnected islands formed by muscle fibers in the treatment group, and connective tissue surrounding the urethral lumen and inflammatory infiltrate in the control group. DISCUSSION AND CONCLUSIONS: The implantation of CAM significantly improved LPP values in the treatment group, and the improvement remained throughout the evaluation period. It may be associated with the consistency of the implant and its stability at the injection site. Longer follow-up studies and human clinical investigations are required to consider CAM implantation as an alternative treatment for stress urinary incontinence.

Partial characterization of enterocin MR99 from a corn silage isolate of Enterococcus faecalis.

AIMS: To assess the inhibitory activity on Gram-positive and Gram-negative bacteria of several species of enterococci recovered from a natural corn silage. METHODS AND RESULTS: The inhibitory activity of strains of Enterococcus faecalis (58), Enterococcus faecium (35), Enterococcus gallinarum (3) and Enterococcus casseliflavus (4) were studied employing indicator strains from various sources (clinical, food and ATCC). Enterococcus faecalis MR99, the only strain with inhibitory activity, inhibited other enterococci, Listeria spp., Staphylococcus aureus, Clostridium spp., Bacillus spp., Escherichia coli, Shigella sonnei and Shigella flexneri. The bacterium contained only one conjugative pheromone-responsive plasmid. The partially chromatography-purified MR99 enterocin (PPE) had a molecular weight of approx. 5000 Da and a pI of 6.2, was sensitive to proteolytic enzymes and could be extracted in benzene and butanol. It appeared stable to adjustment of pH 4.0, 5.0, 6.0, 7.0 and 8.0 and was resistant to heat. Inactivation was at 15 min at 121 degrees C. Enterocin MR99 was bactericidal on strains of Listeria monocytogenes, Staph. aureus, and bovine mastitis agents, it was bacteriostatic on E. coli. Although enterocins MR99 and AS48 have inhibitory activity on Gram-negative bacilli, PCR studies demonstrated a lack of relationship between them. CONCLUSIONS: The active component had a protein nature, was resistant to heat and presented a wide inhibitory spectrum. SIGNIFICANCE AND IMPACT OF THE STUDY: The biological properties of Ent. faecalis MR99 suggest that this strain merits further investigations so it can be applied in human and veterinary health programmes.

Neurotransmitter-induced currents in retinal bipolar cells of the axolotl, Ambystoma mexicanum.

1. Whole-cell patch clamping was used to study the membrane properties of isolated bipolar cells and the currents evoked in them by putative retinal neurotransmitters. 2. Isolated bipolar cells show an approximately ohmic response to voltage steps over most of the physiological response range, with an average input resistance of 1.3 G omega and resting potential of -35 mV. These values are underestimates because of the shunting effect of the seal between the patch electrode and the cell membrane. Depolarization beyond -30 mV produces rapid activation (10-100 ms) of an outward current (carried largely by potassium ions), which then inactivates slowly (0.5-2 s). 3. Of five candidates for the photoreceptor transmitter, four (aspartate, N-acetylhistidine, cadaverine, putrescine) had no effect on bipolar cells. The fifth substance, L-glutamate, opened ionic channels with a mean reversal potential of -12 mV in some cells (presumed hyperpolarizing bipolar cells), and closed channels with a mean reversal potential of -13 mV in other cells (presumed depolarizing bipolar cells). 4. The conductance increase induced by glutamate in presumed hyperpolarizing bipolar cells was associated with an increase in membrane current noise. Noise analysis suggested a single-channel conductance for the glutamate-gated channel of 5.4 pS. The power spectrum of the noise increase required the sum of two Lorentzian curves to fit it, suggesting that the channel can exist in three states. 5. The conductance decrease induced by glutamate in presumed depolarizing bipolar cells was associated with a decrease in membrane current noise that could be described as the sum of two Lorentzian spectra, and which suggested a single-channel conductance of 11 pS. The noise decrease implies that the channels closed by glutamate are not all open in the absence of the transmitter. 6. GABA (gamma-aminobutyric acid) and glycine, transmitters believed to mediate lateral inhibition in the retina, open chloride channels in isolated bipolar cells, and increase the membrane current noise. Noise analysis suggested that the channels gated by GABA and glycine have conductances of 4.4 and 7.5 pS respectively. The noise spectra required the sum of two Lorentzian curves to fit them. 7. By whole-cell patch clamping cells in retinal slices, the synaptic transmitter released by photoreceptors was shown to close channels with an extrapolated reversal potential around -3 mV in depolarizing bipolar cells.(ABSTRACT TRUNCATED AT 400 WORDS)