Biophysical mechanism-mediated time-dependent effect on sperm of human and monkey vas implanted polyelectrolyte contraceptive

Aim： To determine the short and long-term morphological effects on sperm as induced by intra-vas alteration of pH and electrical charge. Methods： Desired biophysical influences were obtained by injection of reversible inhibition of sperm under guidance （RISUG） into the lumen of the vas deferens of human subjects and the monkey. RISUG is a polyelectrolyte hydrogel complex of styrene maleic anhydride （SMA） and dimethyl sulfoxide （DMSO） which generates an electrostatic charge and also lowers in a near space of pH domain. The morphology of sperm was examined by light microscopy, scanning and transmission electron microscopy. Human study enabled semen collection by masturbation as early as 3 h after injection and studies extended up to 6 months, in the monkey, on vas excision after RISUG implantation, sperm characteristics were examined in serial sections. Results： Semenology in clinical studies and histological data of the monkey showed a time-sequenced sperm plasma membrane, tail mitochondria and nuclear decondensation alterations in sperm structural components, which beared marked similarity to changes in the sperm head and tail during capacitation and entry into the ovum. Conclusion： The findings provide a means of causing such changes in the sperm that inhibit the fertilizing ability before the nucleus is affected. Therefore achieving non-obstructive vas-based contraception, without genotoxic or teratogenic effects caused by infertile sperm passing into the semen, is feasible.

miR-148b-3p promotes migration of Schwann cells by targeting cullin-associated and neddylationdissociated 1

MicroRNAs（miRNAs） are small,non-coding RNAs that negatively adjust gene expression in multifarious biological processes.However,the regulatory effects of miRNAs on Schwann cells remain poorly understood.Previous microarray analysis results have shown that miRNA expression is altered following sciatic nerve transaction,thereby affecting proliferation and migration of Schwann cells.This study investigated whether miR-148b-3p could regulate migration of Schwann cells by directly targeting cullin-associated and neddylation-dissociated 1（Cand1）.Up-regulated expression of miR-148b-3p promoted Schwann cell migration,whereas silencing of miR-148b-3p inhibited Schwann cell migration in vitro.Further experiments confirmed that Candl was a direct target of miR-148b-3p,and Candl knockdown reversed suppression of the miR-148b-3p inhibitor on Schwann cell migration.These results suggested that miR-148b-3p promoted migration of Schwann cells by directly targeting Candl in vitro.

In vitro screening of reversible and time-dependent inhibition on CYP3A by TM208 and TM209 in rat liver microsomes

TM208 and TM209,dithiocarbamate derivatives with potential anti-cancer effects,were evaluated in reversible and time-dependent cytochrome P450(CYP)3A inhibition assays in rat liver microsomes using testosterone as probe substrate.Both compounds were found to be weak reversible inhibitors and moderate mechanism-based inhibitors of rat CYP3A.For reversible inhibition on rat CYP3A,the Ki values of competitive inhibition model were 12.10±1.75 and 13.94±1.31 μM,respectively.For time-dependent inhibition,the inactivation constants(K_(l))were 31.93±12.64 and 32.91±15.58 μM,respectively,and the maximum inactivation rates(kinact)were 0.0349770.0069 and 0.0725970.0172 min^(-1) respectively.These findings would provide useful in vitro information for future in vivo DDI studies on TM208 or TM209。

Thermokinetic Study on the Reversible Competitive Inhibition of Bovine Liver Arginase

A new thermokinetic reduced extent method for studying of the reversible competitive inhibition of single sub-strate enzyme-catalyzed reactions was proposed in this paper. The reaction that arginase-catalyzed hydrolysis of L-arginine to L-ornithine and urea and the inhibition of this reaction by the product, L-ornithine, and exogenous L-lysine were studied at 37 ℃ in 40 mmolL-1 sodium barbiturate-HCl buffer solution (pH=9.4). Michealis con-stant Km for arginine and maximum velocity Vm of the reaction were determined to be 5.14 mmolL-1 and 1.13× 10-2 mmolL-1s-1, respectively. The product inhibition constant KP and inhibitory constant KI of L-lysine were de-termined to be 1.18 and 5.6 mmolL-1, respectively. All the results have better repeatability and self-consistency and are in agreement with literature values. This new method using more direct thermal information from the proc-ess would give more reliable kinetic information than the traditional initial rate method.

Contraception with RISUG and functional reversal through DMSO and NaHCO3 in male rabbits

The study aimed to evaluate reversal of short- and long-term vas occlusion with reversible inhibition of sperm under guidance （RISUG） using dimethyl sulfoxide （DMSO） and sodium bicarbonate （NaHCO3） in male rabbits （Oryctolagus cuniculus）. Animals were divided into seven groups containing five animals each. Fortnightly, semen analysis revealed that sperm concentration and output steadily declined after vas occlusion and complete azoospermia was attained at 30-60 days postinjection. Spermatozoa reappeared at 60-75 days of reversal and normozoospermia was noticed between 135 days and 150 days in the reversal groups. All spermatozoa were found nonmotile prior to azoospermia and a gradual recovery in sperm motility was observed between 105 days and 135 days of reversal. A significant decline in viability of sperms was noticed during vas occlusion up to 30-60 days which recovered at 60-75 days postreversal and normalized by 75-105 days in the reversal groups. A significant enhancement in the sperm abnormalities was recorded in all vas occluded animals as well as those in initial periods of reversal. Other parameters, namely, semen volume, ejaculation time, pH, color, and consistency, remained unaltered during all phases of the study. Fertility test, at the intervals of 15 days, demonstrated that animals exhibited complete sterility during the entire period of vas occlusion. A gradual recovery in fertility was observed with the appearance of spermatozoa following vas occlusion reversal and 100% fertility was observed following 135-150 days of reversal. F, progeny of reversed animals was found normal. The results suggest that reversal with DMSO or NaHCO3 is feasible, with normal progeny, following short- and long-term contraception.