Effects of Multiglycoside of Tripterygium Wilfordii(GTW)on Spermatogenic Ce

This study was undertaken to observe consecutively the morphology of testis andepididymis and the changes of enzymatic activities of AKP, ACP,3β-HSD andLDH-C4 in male rats treated with GTW 30 to 80 days.In addition,male antifertility effect and its possible reversibility were also observed.The results showed that GTW is apotential testicular toxicant in the animals.It can cause damage of the sperm cells withclose relation to the treated time and dosage.Sloughing of sperm cells and manymultinucleated giant cells in the seminiferous tubules were found on day 40 and 50 afterthe treatment.The lubules were hyperatrophic and the sperm cells were almost absent atthe end of the study(80days).No obvious morphological alteration was observed in theopididymal epithelial tissue.But changes of quality and number of the spermatozoa inepididymides were found prior to those of lestes.Reduced number and abnormal caudasperm were observed 30 days after the treatment.The ACP activity of Serloli cells increased slightly,whereas foe activities of ACP and LDH-C4 decreased gradually as thetreated lime prolonged.The 3β-HSD Of Leydik.cells was changed or subtle by m-Phaseor treatment and dramatically decreased at the end of treatment.The infertility caused byGTW was reversible 8 weeks after cessation of the treatment.

Selection of High-quality Sperms by the Nanotechnological Method of Magnetic Activation in Brazilian Cervids

Cervids show a high degree of abnormalities in their sperm cells.Thus,this study aimed to select high-quality spermatozoa using magnetic-activated sperm sorting(MASS)compared to density gradient centrifugation(DGC)by assessing the post-selection cell quality.Semen from six Mazama deer was collected by electroejaculation after chemical restraint.The semen was analyzed in four samples:Fresh,DGC,SEMgood-non-apoptotic fraction,and SEMpoor-apoptotic fraction.The material was analyzed for motility and vigor(light microscopy),concentration(Neubauer chamber),semen morphology(phase contrast),and supravital staining test(eosin/nigrosine).The DGC method used 20 x 106 cells in 90%and 45%percoll®gradient.The MASS used 10 x 106 cells with 20μl of iron nanoparticles attached to Annexin V and filtration in a magnetic separation column.Both processing methods(DGC and MASS)were effective in producing high-quality sperm samples,with a marked reduction in abnormalities from 41.83±10.25(fresh)to 14.83±3.17(DGC)and 12±3.01(SEMgood),with 80.3%±2.06 livings cells.These findings suggest that this nanotechnological method,using nanoparticles,effectively produces high-quality semen samples in cervids for use in assisted reproduction.

Mechanism of action of Wuzi Yanzong pill in the treatment of oligoasthenozoospermia in rats determined via serum metabolomics

Objective:To investigate the mechanism of action of Wuzi Yanzong pill(WYP)in rats with oligoasthenozoospermia(OAZ)via metabolomics and to provide a possible basis for improving this WYP-based treatment.Methods:A rat model of OAZ was established by treating male SpragueeDawley rats with glucosides from Tripterygium wilfordii Hook.F.Seventy-two rats were randomly divided into six groups:control,L-carnitine(positive control),model,and low-,medium-,and high-dose WYP groups.Rats in the experimental groups were treated with WYP for 4 weeks.At the end of the treatment period,sperm cell quality(density,motility,and viability)was assessed using a semen analysis system,mitochondrial membrane potential(MMP)was assessed using flow cytometry,and testicular injury was assessed using hematoxylin and eosin staining to validate the therapeutic effect of WYP in OAZ.Further,serum metabolomics-based analysis was performed using high-performance liquid chromatography-mass spectrometry to identify differential metabolic pathways and possible mechanisms of action of WYP in OAZ treatment.Results:A rat model of OAZ was considered successfully-established after comparing the quality of spermatozoa in the model group to that in the control group.WYP-M and WYP-H treatments significantly improved sperm cell density,motility,and viability compared with those in the model group(all P<.05).Compared with the model group,both WYP-M and WYP-H treatments increased MMP values(P=.006 and P=.021 respectively),while there was no significant difference in the L-carnitine group.L-carnitine and WYP administration reversed damage to the testes to varying degrees compared with that in the model group.Further,44 differential metabolites and four metabolic pathways,especially autophagy pathway,related to OAZ were identified via metabolomics.Conclusions:WYP improves sperm cell quality and MMP in OAZ primarily via autophagy regulation.These findings can be employed to improve the efficacy of WYP in humans.

Nitrosative stress in human spermatozoa causes cell death characterized by induction of mitochondrial permeability transition-driven necrosis

Peroxynitrite is a highly reactive nitrogen species and a potent inducer of apoptosis and necrosis in somatic cells. Peroxynitrite-induced nitrosative stress has emerged as a major cause of impaired sperm function; however, its ability to trigger cell death has not been described in human spermatozoa. The objective here was to characterize biochemical and morphological features of cell death induced by peroxynitrite-mediated nitrosative stress in human spermatozoa. For this, spermatozoa were incubated with and without （untreated control） 3-morpholinosydnonimine （SIN-l）, in order to generate peroxynitrite. Sperm viability, mitochondrial permeability transition （MPT）, externalization of phosphatidylserine, DNA oxidation and fragmentation, caspase activation, tyrosine nitration, and sperm ultrastructure were analyzed. The results showed that at 24 h of incubation with SIN-l, the sperm viability was significantly reduced compared to untreated control （P〈 0.001）. Furthermore, the MPT was induced （P〈 0.01） and increment in DNA oxidation （P 〈 0.01）, DNA fragmentation （P 〈 0.01）, tyrosine nitration （P 〈 0.0001） and ultrastructural damage were observed when compared to untreated control. Caspase activation was not evidenced, and although phosphatidylserine externalization increased compared to untreated control （P 〈 0.001）, this process was observed in 〈10% of the cells and the gradual loss of viability was not characterized by an important increase in this parameter. In conclusion, peroxynitrite-mediated nitrosative stress induces the regulated variant of cell death known as MPT-driven necrosis in human spermatozoa. This study provides a new insight into the pathophysiology of nitrosative stress in human spermatozoa and opens up a new focus for developing specific therapeutic strategies to better preserve sperm viability or to avoid cell death.

Osmoregulation determines sperm cell geometry and integrity for double fertilization in flowering plants

Distinct from the motile flagellated sperm of animals and early land plants,the non-motile sperm cells of flowering plants are carried in the pollen grain to the female pistil.After pollination,a pair of sperm cells are delivered into the embryo sac by pollen tube growth and rupture.Unlike other walled plant cells with an equilibrium between internal turgor pressure and mechanical constraints of the cell walls,sperm cells wrapped inside the cytoplasm of a pollen vegetative cell have only thin and discontinuous cell walls.The sperm cells are uniquely ellipsoid in shape,although it is unclear how they maintain this shape within the pollen tubes and after release.In this study,we found that genetic disruption of three endomembrane-associated cation/H+exchangers specifically causes sperm cells to become spheroidal in hydrated pollens of Arabidopsis.Moreover,the released mutant sperm cells are vulnerable and rupture before double fertilization,leading to failed seed set,which can be partially rescued by depletion of the sperm-expressed vacuolar water channel.These results suggest a critical role of cell-autonomous osmoregulation in adjusting the sperm cell shape for successful double fertilization in flowering plants.

Gamete Recognition in Higher Plants:An Abstruse but Charming Mystery

Although much effort has been made to uncover the mechanism underlying double fertilization, little knowledge has been acquired for understanding the molecular base of gamete recognition, mainly because of technical limitations. Still, progress has been made in terms of the mechanism, including the identification of candidate molecules that are involved in gamete recognition in angiosperms. New cues for gamete recognition have been found by the successful separation of the gametes and construction of gamete-specific cDNA libraries in several species, and the application of molecular approaches for studying this process by mutations. Thus, the topic is considered an abstruse but charming mystery.