Optimization of culture conditions for short-term maintenance, proliferation, and colony formation of porcine gonocytes

Background: Gonocytes give rise to spermatogonial stem cells, and thereby play an essential role in establishing spermatogenesis. Optimized culture conditions for gonocytes provide an opportunity for their study and in vitro manipulation for potential application in reproductive technologies. Using six experiments in a step-wise design, we examined the effects of several culture conditions on the maintenance, proliferation, and colony formation of porcine gonocytes. Testis cells from neonatal piglets were cultured for 7 d in DMEM supplemented with 10% fetal bovine serum. The examined culture conditions included using different cell seeding densities, gonocyte proportions, incubation temperatures, sampling strategies, and medium changing regimens.Results: Confluency of cel s was optimal(>90% by ~6 d) when 3.0 × 104 testis cel s/cm2 containing ~40% gonocytes were used. Incubating the cel s at 35 °C or 37 °C resulted in similar cel number and viability at confluency, but incubation at 35 °C resulted in a delayed confluency. In the first 2 d of culture, gonocytes remained mostly floating in the medium and gradual y settled over the next 5 d. Consequently, not changing the medium for 7 d(as opposed to changing it every 2 d) led to a significant increase in the number of gonocyte colonies by reducing the loss of "floating gonocytes".Conclusion: We found that gonocytes require the presence of a critical minimum number of somatic cel s for settlement, and can proliferate and form growing colonies even in a basic medium. Large numbers of viable gonocytes remain floating in the medium for several days. The optimized culture conditions in the present study included seeding with 3.0 × 104 testis cel s/cm2 containing ~40% gonocytes, incubating at 37 °C, and without changing the medium in the first week, which can result in improved colony formation of porcine gonocytes.

Functional recovery of sciatic nerve through inside-out vein graft in rats

客观现在的学习瞄准了进一步全面功能，对用老鼠的外部神经新生的 histomorphometrical 和 immunohistochemical 评价臀部的神经横断模型。方法 10 公里老鼠臀部的神经差距在老鼠被创造。在控制组神经树桩是到邻近的肌肉的 sutured，在治疗组，差距被衔接用一在内外面静脉接枝。在假冒操作组，神经被操作并且留给未经触动。所有动物经历了在外科以后走轨道分析测试 4， 8，和 12 星期。随后，肌肉团测量被执行估计虚弱，词法上观察臀部的神经新生的组织学的考试和用反 S-100 检测 Schwann 房间的 Immunohistochemistry。结果与二个在题目之间因素用因素的 ANOVA 被分析。为 pairwise 比较的 Bonferroni 测试被用来检验处理的效果。myelinated 神经纤维的功能的分析显示出的结果那神经功能在处理在时间功课显著地改善了组。然而， myelinated 神经纤维的量的 morphometrical 分析证明在处理组的 8 和 12 个星期之间没有重要差别。肌肉重量比率更大， gastrocnemius 肌肉的重量损失被改善由在内外面静脉 grafting。积极 immunohistochemical 反应的位置进一步暗示那改革了轴突，在静脉 grafting 被执行以后， Schwann 像房间的细胞存在，并且被 myelination 的过程和改革神经的结构的恢复伴随。外部神经修理的结论功能的分析比量的 morphometrical

Effect of local administration of cyclosporine A on pe- ripheral nerve regeneration in a rat sciatic nerve transec- tion model

ObjectiveTo 估计在在老鼠 model.MethodsA 衔接臀部的神经缺点上在 chitosan 水管装载的 cyclosporine A (CsA ) 的效果 10 &#xA0；公里臀部的神经缺点用充满 10 &#xA0 的一根 chitosan 水管被衔接； l 搬运人药冲淡(10 &#xA0； g/L CsA ) 。在控制组，水管独自充满搬运人冲淡的一样的卷。改革纤维被学习 4， 8 和 12 &#xA0；在功能的学习更快证实了的 surgery.ResultsThe 以后的星期在处理组的改革轴突的恢复比控制组(P&#xA0； <&#xA0； 0.05 ) 。有在处理和控制组之间的 gastrocnemius 肌肉重量比率的统计上重要的差别(P&#xA0； <&#xA0； 0.05 ) 。改革纤维的 Morphometric 索引证明在对待 CsA 的动物的 myelinated 纤维的数字和直径比在控制组的那些显著地高。在 immunohistochemistry，对在 CsA 组的 S-100 的反应的地点清楚地比控制更积极在一根 chitosan 水管装载的 group.ConclusionCsA 导致功能的恢复的改进和臀部的神经的量的 morphometric 索引。没有任何复杂并发症，它与它的全身的管理相比是容易可得到的。

Kisspeptin stimulates oocyte maturation,and food deprivation modulates the abundance of kisspeptin system in zebrafish gonads

Kisspeptin is a hormone involved in the neuroendocrine control of reproduction in fish.We hypothesized that kisspeptin stimulates oocyte maturation and modulates other reproductive hormones in zebrafish;and the gonadal kisspeptin system in zebrafish is affected during energy unavailability.The main goals of this research were to test in vitro effects of kisspeptin on oocyte maturation and mRNA abundance in zebrafish ovarian follicles and determine how short-term feed restriction affects kisspeptin and its receptors in zebrafish testis and ovary.This study demonstrates the presence of kisspeptin and its receptors in zebrafish(Danio rerio)gonads and its direct action on ovarian follicles in vitro.Kisspeptin(10 ng/mL)induced oocyte maturation,as indicated by germinal vesicle break down at 18-and 24-hours post-treatment.Kisspeptin significantly increased the abundance of mRNAs encoding reproductive hormones and its receptors in zebrafish oocytes.This suggests that kisspeptin-10 affects ovarian functions by modulating other hormones.Reproduction is a process that requires energy.Therefore,whether energy availability affects the kisspeptin system in zebrafish gonads was determined.Food deprivation modulated kisspeptin expression differently in zebrafish testis and ovary.Kiss2 and kiss1ra were upregulated while kiss1rb was downregulated in the testis post-food deprivation.Meanwhile,no changes in kiss in the ovary were found after food deprivation.However,kiss1rb was downregulated in unfed fish at 3-and 7-days post-food deprivation.Overall,our results suggest sex-and tissue-specific changes in the gonadal abundance of the kisspeptin system in zebrafish.The fine tuning of reproduction during energy fluctuations in fish is likely mediated via changes in hormones,including kisspeptin as shown in this research.

Proteomic Analysis of Bovine Nucleolus

Nucleolus is the most prominent subnuclear structure, which performs a wide variety of functions in the eukaryotic cellular processes. In order to understand the structural and functional role of the nucleoli in bovine cells, we analyzed the proteomic composition of the bovine nucleoli. The nucleoli were isolated from Madin Darby bovine kidney cells and subjected to proteomic analysis by LC-MS/MS after fractionation by SDS-PAGE and strong cation exchange chromatography. Analysis of the data using the Mascot database search and the GPM database search identified 311 proteins in the bovine nucleoli, which contained 22 proteins previously not identified in the proteomic analysis of human nucleoli. Analysis of the identified proteins using the GoMiner software suggested that the bovine nucleoli contained proteins involved in ribosomal biogenesis, cell cycle control, transcriptional, translational and post-translational regulation, transport, and structural organization.

The ghrelinergic system in zebrafish gonads is suppressed during food unavailability

Ghrelin is an appetite stimulatory peptide that regulates reproduction in vertebrates.The acylation of ghrelin by ghrelin-O-acyltransferase(Goat)is essential for most of its functions.The ghrelinergic system(ghrelin,ghrelin receptor[Ghsr]and Goat)is present in teleost fish ovary and testis.In this research,we determined the abundance of the ghrelinergic system mRNAs in different stages of follicle(oocyte)development in zebrafish ovary.The highest levels of preproghrelin mRNA were observed in the first stage of follicular development(previtellogenesis).Significantly low levels of preproghrelin mRNA were observed in stages 3II,4 and 5 of the follicle development.Ghsr mRNA abundance remained unchanged at early stages of follicle development and decreased in stage 5 compared to other developmental stages.A significant decrease in preproghrelin and Ghsr mRNAs in the testis,and ghsr mRNA in the ovary of zebrafish deprived of food for 3 days were found.At 7 days food deprivation,preproghrelin,ghsr and goat mRNA in the ovary and preproghrelin and goat in the testis of zebrafish were significantly reduced.These results show that metabolic status modulates gonadal ghrelinergic system in zebrafish and provide support for a role for ghrelin in fish reproduction.

Canada’s inter-agency wild bird influenza survey

Canada’s Inter-agency Wild Bird Influenza Survey,which started in 2005,and is a collaborative effort among federal,provincial and territorial government agencies as well as non-governmental organizations and academic institutions,has two components:(i)a seasonal survey of live wild waterfowl species from selected geographic regions across Canada(to expand our understanding of the avian influenza viruses circulating in wild bird populations);and(ii)an ongoing survey of birds found dead that are submitted to a regional diagnostic laboratory(to enhance detection of highly pathogenic avian influenza strains).Combined swabs from the cloaca and oro-pharynx collected from each bird are screened using a real-time reverse transcriptase-polymerase chain reaction(RRT-PCR)that targets a unique segment of the influenza A M1 gene.If the M1 result is positive or inconclusive,RRT-PCR for gene segments of the H5 and H7 hemagglutinin subtypes are performed.All samples that are RRT-PCR positive for H5 or H7 are sent immediately for test confirmation and further characterization.All field and laboratory data are entered into a database developed and maintained by the Canadian Cooperative Wildlife Health Centre.Since the survey commenced in 2005,on average,30%of all live ducks sampled,5%of other species of live birds and 3%of birds found dead have tested positive for avian influenza,all of North American lineage and of low pathogenicity.

Editorial Reproduction and gonadal development in bony fishes

Fish-is an important source of dietary protein around the globe.Aquaculture aims to fulfill the supply gaps created by insufficient number of fish captured from the wild to meet the ever-increasing demand.One approach to address this renewed yield goal is by enhancing reproduction in fish used in aquaculture.However,enhancing reproduction necessitates a deeper understanding of our fundamental knowledge on reproductive endocrinology and physiology of fishes.There are many teleost fishes used for human consumption and the species choice largely depends on regional availability and geographical factors.This creates a need to study individual species to better understand the species-specific biology of reproduction in fishes.In line with this,research by fish reproductive biologists is generating significant new knowledge on fishes of interest in aquaculture,many of which are endemic to specific regions where humans inhabit.

猪Ⅱ型圆环病毒对现代养猪生产的重大影响

由于感染了猪Ⅱ型圆环病毒的疾病的范围已有所扩大，那些能加重疾病发展的感染辅助因子现已成为研究热点。本文综述了猪Ⅱ型圆环病毒对现代养猪生产的重大影响。