Total saponins of Panax ginseng effects on proliferation and differentiation of human embryonic neural stem cells and in a Parkinson's disease mouse model

BACKGROUND： Total saponins of Panax ginseng （TSPG） exhibits neuroprotection against Parkinson＇s disease in the substantia nigra. OBJECTIVE： To investigate the effects of TSPG on human embryonic neural stem cells （NSCs） proliferation and differentiation into dopaminergic neurons using in vitro studies, and to observe NSC differentiation in a mouse model of Parkinson＇s disease, as well as behavioral changes before and after transplantation. DESIGN, TIME AND SETTING： In vitro neural cell biology trial and in vivo randomized, controlled animal trial were performed at the Institute of Basic Medical Sciences, Chongqing Medical University between September 2004 and December 2007. MATERIALS： TSPG （purity 〉 95%） was isolated, extracted, and identified by Chongqing Academy of Chinese Materia Medica. Recombinant human basic fibroblast growth factor （bFGF） and recombinant human epidermal growth factor （EGF） were purchased from PeproTech, USA. A total of 25 C57/BL6J mice, aged 18-20 weeks were included. Twenty were used to establish a Parkinson＇s disease model with i.p. injection of MPTP （1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine） and TSPG alone or combined with interleukin-1 （IL-1）-treated NSCs prior to transplantation into the corpus striatum. The remaining five mice were pretreated for 3 days with TSPG prior to MPTP injection, serving as the TSPG prevention group. METHODS： Primary NSCs were isolated, cultured and purified from embryonic cerebral cortex. Immunocytochemistry was employed to detect specific antigen expression in the NSCs. In vitro experiment： （1） to induce proliferation, NSCs were treated with TSPG, EGF＋bFGF, or TSPG＋EGF＋bFGF, respectively; （2） to induce dopaminergic neuronal differentiation, NSCs were treated with TSPG, IL-1, or TSPG＋IL-1, respectively. MAIN OUTCOME MEASURES： In vitro experiment： the effects of TSPG on NSCs proliferation were evaluated with flow cytometry and MTT assay. Tyrosine hydroxylase expression was determined by immunocytochemistry assay to observe effects of TSPG on dopaminergic neuronal differentiation. In vivo experiment： differentiation of grafted NSCs in the mouse brain was determined by immunohistochemical staining. Behavioral changes were evaluated by spontaneous activity frequency, memory function, and score of paralysis agitans. RESULTS： （1） NSCs were cultured and passaged for more than three passages. Immunocytochemistry revealed positive nestin staining, as well as neurofilament protein and glial fibrillary acidic protein. （2） TSPG significantly increased NSC proliferation, in particular when combined with EGF and bFGF, which was twice as effective as FGF or bFGF alone. TSPG also induced dopaminergic differentiation in NSCs, in particular when TSPG was added together with IL-1, resulting in an effect five times greater than that of IL-1 alone. （3） At day 30 following transplantation, most NSCs in the TSPG prevention group differentiated into dopaminergic neurons, and the scores of paralysis agitans, spontaneous activity, and memory function were significantly increased compared with TSPG alone or TSPG＋IL-1 groups （P 〈 0.05）. CONCLUSION： TSPG stimulated NSC proliferation, in particular when combined with FGF and bFGF. TSPG significantly induced dopaminergic neuronal differentiation of NSCs, and the effect was greater when combined with IL-1. In addition, TSPG greatly improved behavior in the Parkinson＇s disease mouse model following NSC transplantation. Following NSC transplantation, TSPG pretreatment exhibited superior efficacy over either TSPG alone or TSPG in combination with IL-1, in terms of behavioral improvements in the Parkinson＇s disease mouse model.

Transglutaminase 3 expression in C57BL/6J mouse embryo epidermis and the correlation with its differentiation

Epidermal-type transglutaminase 3 (TGM3) is involved in the cross-linking of structural proteins to form the cornifiedenvelope in the epidermis. In the present study, we detected the expression of TGM3 in the mouse embryo using RT-PCR.TGM3 mRNA is weakly presented from E11.5 to E14.5 and increases significantly from E15.5 to birth. Then wedetermined the spatial and temporal expression pattern of TGM3 in the skin and other organs by in situ hybridization. Wefound a deprivation of TGM3 in skin at E11.5, while a rich supply in periderm cells and a weak expression in basal cellsfrom E12.5 to E14.5. From the period of E15.5 to E16.5, after keratinization in the epidermis, TGM3 was expressed inthe granular and cornified layers. The electron microscopic observation of the C57BL/6J mouse limb bud skin develop-ment provided several morphological evidences for the epidermal differentiation. The above findings suggest that theexpression of TGM3 plays a important role in the epidermis differentiation in embryogenesis.

Lens Capsule HSPG-Perlecan Regulates Lens Fibre Differentiation during Chick Embryo Development

Lens fibre differentiation is a life-long process related with lens transparency, and is particularly intense during development, being related with an FGF-2 antero-posterior gradient at the equator level as the main growth factor involved which has been related with the basal membrane of the lens anlagen known as “Lens capsule”. However the lens fibre differentiation induced by FGF2 depends, as in other biological systems, on the local bioavailability of FGF-2 regulated by their relationship with extracellular matrix molecules as Heparan Sulphate Proteoglycans. Here, we try to clarify how Perlecan (a heparan sulphate proteoglycan specific from basement membranes) is involved in lens fibre differentiation at earliest stages of eye development. Our results show that Perlecan, is a major component in the lens capsule during the earliest stages of lens development in chick embryos being present during lens plate induction, lens vesicle stage and the onset of lens fibre differentiation. In order to demonstrate a direct involvement of HSPG-Perlecan in lens fibre differentiation, we generate depleted lenses by HSPG-Perlecan synthesis disruption and specific enzymatic digestion. The HSPG-Perlecan depleted lens show a significant delay or abolition in the lens fibre differentiation which remains in an immature cells displaying DNA synthesis in the posterior epithelium and a decrease in FGF2 lens expression. These data support the hypothesis that lens capsule HSPG-Perlecan is a key molecule involved in lens fibre differentiation during development, probably by involvement in FGF-2 biodisponibility.

DEC1 nuclear expression:A marker of differentiation grade in hepatocellular carcinoma

AIM: To investigate the expression patterns of human differentiated embryo chondrocyte 1 (DEC1) in hepatocellular carcinoma (HCC) and corresponding adjacent non-tumor and the normal liver tissues, the association between DEC1 expression and histopathological variables and the role of DEC1 in hepatocarcinogenesis. METHODS: The expression of DEC1 was detected immunohistochemically in 176 paraffin-embedded sections from 63 patients with HCC and 50 subjects with normal liver tissues. RESULTS: DEC1 protein was persistently expressed in the cytoplasm of hepatocytes in normal liver and HCC tissues. Compared with adjacent non-tumor liver tissues, HCC tissues showed high nuclear expression of DEC1 protein. However, high DEC1 nuclear expression was more frequently detected in well-differentiated (83.3%) than in moderately (27.3%) and poorly differentiated HCC (16.7%). Low DEC1 expression was associated with poor histological differentiation and malignancy progression. A correlation was found between the nuclear expression of DEC1 protein and histological differentiation (r = 0.376, P = 0.024). CONCLUSION: DEC1 is expressed in the cytoplasm of hepatocytes and because nuclear DEC1 expression is decreased with decreasing differentiation status of HCC, nuclear DEC1 might be a marker of HCC differentiation.

EFFICIENT CALLUS INDUCTION AND PLANT REGENERATION FROM IMMATURE EMBRYO CULTURE OF TRITICUM AESTIVUM L.- THINOPYRUM INTERMEDIUM ALIEN DISOMIC ADDITION LINES

An efficient plant regeneration system was developed from the immature embryos of Triticum aestivum L. - Thinopyrum intermedium alien disomic addition lines, which resistant to powdery mildew. The protocol was based on a series of experiments involving the callus induction and differentiation. The experiment studied the effects of embryo size on callus induction and differentiation of the immature embryos. We found that the embryo size is critical for the establishment of embryogenic callus. Immature embryos (0.8～1.5 mm) showed high ability to produce embryogenic callus capable of regenerating green plants. The medium Murashige and Skoog’s (MS) added with 2mg/L 2, 4-dichlorophenoxyacetic acid (2, 4-D) gave the best embryogenic callus induction, maintenance and regeneration. The embryogenic callus maintained high regeneration during six subcultures in the callus induction medium. Suitable time of partial desiccation could effectively improve the regeneration capacity of the callus cultured for 3～4 month.Bud green spot and root green spot were observed during the differentiation of callus and the difference between them was described. Regenerated shoots were rooted on half-strength MS medium containing 0.2 mg/L Naphthalene acetic acid (NAA). Plants were successfully transferred to soil and grew well. This efficient plant regeneration system provides a foundation for the study of somaclonal variation of Triticum aestivum L. - Thinopyrum intermedium alien disomic addition lines.

Study on Callus Induction from Immature Embryos and Plant Regeneration of Different Genotypes of Maize

The induction, subculture and differentiation of callus from immature embryos of maize （ Zea mays L. ） inbred line were studied and optimized. The re- suits revealed that, 2 mg/L 2,4-D was the optimal concentration to induce embryonic callus. Calli induced from inbred Qi319 and LY92 had good morphology and high regeneration. The embryos of the two inbred lines were selected as explants to establish efficient and stable tissue culture and transformation system.

Cocoa (<i>Theobroma cacao</i>L.) Somatic Embryos Tolerate Some Ice Crystallization during Cryopreservation

Secondary somatic embryos (SSEs) of cocoa, a recalcitrant tropical, seed-producing species, were cryopreserved using a vitrification approach and Differential Scanning Calorimetry (DSC) was employed to optimise sucrose preculture and Plant Vitrification Solution 2 (PVS2) incubation. The objective of the study was to evaluate the influence of sucrose preculture and PVS2 dehydration on water content of SSE that will enable it to survive cryostorage. SSEs were precultured for 3 or 5 days on media containing 0.5 M or 0.75 M sucrose and cryoprotected in loading solution (2 M glycerol and 0.4 M sucrose in medium) for 20 min before they were dehydrated with cold PVS2 for 0 - 90 min. Thermal analysis revealed the occurrence of ice crystallization in the SSEs with the extent declining with increasing PVS2 exposure. Maximal survival of SSEs was promoted by preculture on 0.5 M sucrose medium and dehydration with PVS2 for 45 - 60 min, which was characterised by small ice crystallization. Exposure of SSEs beyond 60 min leads to excessive dehydration as characterized by no change in the thermograms. Based on these findings, preculture of SSEs on 0.5 M sucrose medium and dehydration with cold PVS2 for 60 min has been adopted for the successful cryopreservation of cocoa germplasm.

‘玛瑙红’樱桃胚败育果实和正常果实比较转录组分析

为探索‘玛瑙红’樱桃在胚败育情况下果实仍正常发育的分子机理,本研究采用Illumina高通量测序技术,对不同发育时期的‘玛瑙红’樱桃胚败育果实和正常果实样品进行转录组测序及相关分析。结果共获得98.87%的Clean Reads,86.24%的Mapped Reads,Q30>94.21%;共筛选出43673个差异表达基因,其中所有样品组合共有的差异表达基因为8213个,大多分布在FPKM 0~0.01区间。富集分析显示,注释到GO的差异表达基因参与分子功能、细胞组分、生物过程等,其中参与生物过程的最多;注释到KEGG通路的差异表达基因分别参与环境信息处理、生物体系统、遗传信息处理、细胞过程和新陈代谢五大通路的18个分支,大多注释到新陈代谢通路;预测为转录因子的差异表达基因共2002个,共注释到53个转录因子家族,其中ERF家族最多。研究表明,‘玛瑙红’樱桃胚败育果实与正常发育果实在膨大期基因表达差异较大,果实转色期至果实成熟基因表达差异逐渐变小,这可能是胚败育果实正常成熟的原因。

水稻愈伤组织诱导及不定芽分化培养体系建立

目的:筛选高效诱导水稻愈伤组织及不定芽分化培养体系的培养基,为水稻基因工程育种奠定基础。方法:以优质水稻品种(润珠香占)成熟胚为外植体,研究成熟胚愈伤组织出愈情况、不同培养基以及植物生长物质浓度配比对愈伤组织诱导及不定芽分化的影响。结果:水稻愈伤诱导培养基出愈率大小为N6>B_(5)>MS>WPM,N_(6)与B_(5)、MS、WPM差异显著;2,4-D浓度愈伤组织诱导率大小为B_(4)>B_(3)>B_(5)>B_(6)>B_(2)>B_(1),B_(4)与各处理组之间差异显著;植物生长物质浓度配比对愈伤组织不定芽分化率大小为T_(5)>T_(7)>T_(6)>T_(1)>T_(8)>T_(3)>T_(2)>T_(4),T_(5)与各处理组之间差异显著。结论:水稻成熟胚愈伤诱导的最佳培养基及2,4-D质量浓度分别为N6、2.0 mg/L;不定芽分化最佳植物生长物质6-BA、KT、NAA质量浓度比为1∶1∶1,不定芽分化率最高。

草地早熟禾无融合生殖胚囊发育的转录组学分析

为探究草地早熟禾无融合生殖胚囊发育过程中基因表达谱的变化,本研究利用高通量Illu-mina Hiseq测序技术对‘清水’草地早熟禾无融合生殖子房发育的无孢子起始细胞发生时期(8DBA)和无孢子生殖胚囊发育时期(6DBA、4DBA和2DBA)共4个阶段的小穗进行转录组测序。结果显示,不同差异分组(2DBA vs 4DBA、2DBA vs6DBA和2DBA vs 8DBA)共有13575个DEGs,包括7191个上调基因和6384个下调基因。GO富集分析发现,3个差异分组的DEGs显著富集在过氧化氢的响应、蛋白质重折叠、营养水库活动、错误折叠蛋白结合和双糖生物合成等过程中。KEGG通路分析表明,3个差异分组的DEGs显著富集在代谢途径、次生代谢产物的生物合成、淀粉和蔗糖代谢及内质网中的蛋白加工等通路上,在2DBA vs 8DBA分组中发现植物激素信号转导途径的富集。4个时期占比最多的转录因子家族是NAC、AP2/ERF-ERF和MYB-related等。将在4个发育阶段特异性高表达的基因OPR1、ASN2、SAMS2、HSP81-2、PXG4和FPK2作为与‘清水’草地早熟禾无融合生殖胚囊发育相关的候选基因。qRTPCR分析表明,除WOX2在4DBA上调,与RNA-seq的表达存在差异外,其余16个DEGs的表达量变化趋势与高通量测序结果基本一致。

玉米胚发育过程蛋白组分析

高油玉米是一种籽粒含油量比普通玉米高50%以上的玉米类型,其籽粒中油分含量85%集中于胚中。本研究以高油玉米自交系GY220为材料,对授粉后15 d(G15)、25 d(G25)、35 d(G35)的胚蛋白质组进行双向凝胶电泳分离,利用串联质谱对分离蛋白质进行鉴定,并分析了差异蛋白功能。结果表明在授粉后3个发育时期共鉴定到41个差异表达蛋白,其中授粉后25 d相对于15 d G25/G15上调18个,下调19个;授粉后35 d相对于25 d G35/G25上调18个,下调25个;授粉后35 d相对于15 d G35/G15上调20个,下调22个。通过GO注释分析和KEGG富集分析显示差异表达蛋白主要在小分子代谢、氧化还原和碳水化合物代谢等过程富集。根据差异蛋白表达丰度及其功能注释,甘油-3-磷酸脱氢酶(Zm00001d041962)和果糖激酶(Zm00001d035037)在15 d、25 d时蛋白表达量较高,在35 d时蛋白表达量较低;RT-PCR分析表明在自交系GY220胚发育进程中Zm00001d041962和Zm00001d035037表达量呈逐渐增加趋势,两个基因均与大豆、花生、油菜中的相关基因高度同源。本研究为进一步提升玉米籽粒品质和挖掘胚发育功能基因奠定基础。

槲树雌花和种子生长发育动态研究

【目的】阐明槲树雌花发育的生殖周期和发育时序特征。【方法】以人工控制授粉后不同发育时期的槲树雌花为试验材料,通过形态学、生理指标测定和石蜡切片等检测方法,对授粉前后不同发育时期雌花及子房的外部形态、种子发育进程和败育动态进行观察。【结果】(1)在雌花整个生长发育过程,刺苞逐渐变硬,苞片由绿转黄,柱头不断萎缩,花柱不断缩短;子房体积不断膨大,外围壳斗逐渐增厚,于授粉后140 d左右形成棕褐色橡子。(2)授粉后17 d,胚珠发育完成;授粉后30 d,胚囊发育成熟;授粉后38 d,初生胚乳核出现,原胚形成;之后经过了大约13 d休眠后,依次经历球型胚、心型胚、鱼雷胚和子叶胚的分化;而胚乳作为滋养组织,一直处于高度分裂状态,在种子成熟期被子叶全部吸收利用,最终收获无胚乳的成熟种子。(3)初步明确了槲树种胚在整个生长周期内共存在2次败育高峰期,分别在授粉后69 d和99 d。(4)种子败育的表观形态主要表现为败育胚子房壁褐化、胚珠皱缩,显微结构上则主要表现为珠被发育异常、胚胎畸形或空腔胚囊。【结论】本研究建立了槲树雌花分化和种子形成过程外部形态变化与内部解剖特点之间的关系,可为今后栎属胚胎学研究、开花结实机理和胚败育机制研究提供理论依据。

孵化过程中鸡胚性别分化期的判定

【目的】研究鸡胚孵化过程中,通过不同方法判断其性别分化的时间。【方法】以黄羽肉鸡胚蛋为研究对象,孵化黄羽肉鸡胚蛋(温度37.8℃,相对湿度60%)。分别通过胚线观察法、生殖腺器官形态观察法、组织学形态学、分子生物学来判断鸡的性别分化期,试验重复3次。【结果】通过胚线观察法分析发现,在鸡胚孵化期第3天,可以通过血管分支的粗细和形态来辨别雌雄。通过生殖腺器官的观察,在孵化期第8天,能够明显关察到雄性性腺对称发育,雌性性腺则不对称发育。通过组织学形态鉴定,在孵化期第7~10天,观察到雄性性腺有类似于曲精细管的团索状结构出现,雌性性腺有皮髓结构出现,能够判定鸡胚性别分化期可能为胚胎孵化期第7天。通过分子生物学鉴定,孵化期第3~10天,观察到雄性个体有1条扩增条带,其大小是为550 bp左右,观察到雌性个体有2条扩增条带,其大小为550 bp和450 bp左右,因此判定性别分化期可能为孵化期第3天。【结论】综上判定鸡性别分化期约在胚胎孵化期第3~8天。

Hepatic differentiation from embryonic stem cells in vitro

OBJECTIVE: To investigate an method for hepatic differentiation from embryonic stem cells (ES cells) in vitro and the resulting differentiation ratio, in order to develop a procedure for producing a new type of hepatocyte for hepatocyte replacement therapy in the treatment of liver failure. METHODS: ES cells from Balb/C mice were cultured and maintained in an undifferentiated state in gelatin-coated dishes using Dulbecco's modified Eagle's medium (DMEM) containing 1000 U/ml leukemia inhibitory factor (LIF). Then, LIF was withdrawn from the DMEM to allow the ES cells to develop into embryonic bodies (EBs). EBs were plated onto tissue culture dishes, and growth factors such as acidicfibroblast growth factor (aFGF) and hepatocyte growth factor (HGF) were added to the medium to promote directional differentiation. The course of development and differentiation was observed dynamically using an inversion microscope. The expression of hepatic proteins, such as alpha-fetoprotein (AFP), albumin (ALB), cytokeratin 8 (CK8), cytokeratin 18 (CK18), in cytoplasm was analyzed by immunocytochemistry (ICC). The concentration of ALB in the medium was determined dynamically by radioimmunoassay (RIA). RESULTS: ES cells replicated as clones, without differentiating, in DMEM containing LIF. They developed into EBs in medium without LIF. Our ICC assay showed that differentiating cells did not express hepatic proteins, such as AFP, ALB, CK8, and CK18 until day 7, day 9, day 11, and day 11, respectively (up to 2 days later when growth factors are not present). The concentration of AFP in the medium was first detected on day 8, at a concentration of 3.4 ng/ml, and increased to 22.8 ng/ml by day 15. The concentration of ALB in the medium was 0.2 micro g/ml on day 11, and increased to 2.2 micro g/ml by day 15. ALB-positive cells under ICC manifest morphological structures were consistent with normal mouse hepatocytes. The differentiation ratio of hepatocytes in the ES cell differentiation system was 30% on day 15 (significantly lower without the presence of growth factors). CONCLUSIONS: ES cells can differentiate into mature hepatocytes. Growth factors, such as aFGF and HGF, can enhance this differentiation and produce sufficient numbers of functional hepatocytes. This method may be a reliable new way of differentiating ES cells into hepatocytes for use in replacement therapy in the treatment of liver failure.

宫腔镜联合CD138及子宫内膜微生态检测对慢性子宫内膜炎的诊断价值

目的探讨宫腔镜联合白细胞分化抗原138(CD138)及子宫内膜微生态检测对慢性子宫内膜炎(CE)的诊断价值。方法纳入2020-06—2023-05郑州大学第二附属医院生殖医学部收治的行体外受精-胚胎移植(IVF-ET)失败的541例患者,A组(101例)行宫腔镜联合CD138及子宫内膜微生物组群检查,B组(240例)行宫腔镜联合CD138,C组(200例)行宫腔镜检查。分析三种检查方法对CE的诊断价值。结果A组、B组、C组CE的检出率分别为74.26%、62.50%、51.50%,差异有统计学意义(P<0.05)。C组宫腔镜的漏诊率为16.67%,B组的漏诊率为9.90%。三联诊断的准确率高于三者单独诊断与二联诊断(P<0.05)。结论宫腔镜联合CD138与子宫内膜微生物组群检测,能提高IVF-ET失败患者的CE检出率,降低漏诊率,并指导临床精准治疗。

MUM-1、CD138、CD38对慢性子宫内膜炎的诊断价值及其与生殖预后的关系

目的分析多发性骨髓瘤癌基因-1(MUM-1)、白细胞分化抗原(CD)138、CD38在对慢性子宫内膜炎(CE)中的诊断价值及其与生殖预后的关系。方法选取2018年1月至2021年12月因不孕症于山东省济南市第二妇幼保健院生殖健康科就诊,并在山东大学齐鲁医院生殖医学中心接受体外受精-胚胎移植(IVF-ET)的194例患者作为研究对象。所有研究对象术前均接受宫腔镜检查,记录宫腔情况,取子宫内膜组织进行MUM-1、CD138、CD38标志物免疫组织化学染色及子宫内膜活检,并将子宫内膜苏木精-伊红染色法(HE)染色结果作为金标准,免疫组织化学染色结果与HE染色结果的一致性分析使用Kappa检验,分析免疫组织化学染色MUM-1、CD138、CD38单独及3项指标联合检测对CE的诊断效能。根据生殖预后将194例患者分为失败组和成功组,比较失败组和成功组的临床资料,采用多因素Logistic回归分析IVF-ET失败的危险因素。结果HE染色结果显示,194例接受IVF-ET的患者中90例确诊为CE;MUM-1、CD138、CD38单独及3项指标联合检测的免疫组织化学结果与HE染色结果的一致性分析显示,Kappa值分别为0.475、0.537、0.464、0.752(P<0.05);3项指标联合诊断CE的灵敏度、准确度、阳性预测值均高于MUM-1、CD138、CD38单独诊断,差异均有统计学意义(P<0.05);CD38单独诊断CE的灵敏度高于MUM-1、CD138单独诊断,特异度低于MUM-1、CD138单独诊断,差异均有统计学意义(P<0.05)。194例接受IVF-ET的患者中妊娠失败106例,妊娠成功88例。失败组与成功组年龄、体质量指数、不孕年限、月经经期、月经周期,以及基础内膜厚度、基础促卵泡刺激素、基础雌二醇、基础促黄体生成激素水平比较,差异均无统计学意义(P>0.05);失败组继发性不孕、人工流产次数≥1次、CE、MUM-1阳性、CD138阳性、CD38阳性患者比例均高于成功组,差异均有统计学意义(P<0.05)。多因素Logistic回归分析结果显示,继发性不孕、人工流产次数≥1次、患有CE、MUM-1阳性、CD138阳性、CD38阳性是IVF-ET失败的危险因素(P<0.05)。结论MUM-1、CD138、CD38联合检测可提升对CE的诊断效能,且患有CE、MUM-1阳性、CD138阳性、CD38阳性是IVF-ET失败的危险因素。

Effect of soothing liver therapy on oocyte quality and growth differentiation factor-9 in patients undergoing in vitro fertilization and embryo transfer

OBJECTIVE:To investigate the effect of Soothing liver therapy on infertile women undergoing in vitro fertilization and embryo transfer(IVF-ET)and to explore its mechanism.METHODS:Fifty-eight women with tubal infertility were randomized into two groups:30 in an experimental group treated with Xiaoyao powder(Shugan)plus gonadotropin-releasing hormone analog(GnRHa)/follicle-stimulating hormone(FSH)/human chorionic gonadotropin(hCG)and 28 in the control group who were treated with GnRHa/FSH/hCG only.The total gonadotropin(Gn)doses required,endometrial thickness,oocyte numbers,high quality embryo production rate and pregnancy rate of the two groups were compared.The concentration of growth differentiation factor-9(GDF-9)in follicular fluid was detected by western blotting and the expression of GDF-9 mRNA in granulosa cells was measured using reverse tran-scription-polymerase chain reaction amplification.RESULTS:In the experimental group,the Gn dose was significantly lower than that in the control group;the endometrial thickness,high quality embryo production and pregnancy rates were significantly higher and the expression of GDF-9 mRNA was also significantly higher than in the control group(all P<0.05).CONCLUSION:Shugan treatment can improve the pregnancy rate of women with tubal infertility;its mechanism is possibly related to the increased expression of GDF-9 in granulosa cells.

早期胚胎极性建立及对谱系分化的影响

极性建立是影响早期胚胎发育的关键因素之一。极性建立起始于8细胞胚胎的肌球蛋白磷酸化,磷酸化激活肌动蛋白导致其启动收缩力。随后,肌动蛋白发生重组在每个卵裂球的非接触表面形成富含微绒毛的顶端结构域,并在极性蛋白复合物等的共同作用下形成标志着顶端结构域成熟的肌动球蛋白环。从极性建立的过程可知,顶端结构域的形成受到肌动蛋白相关蛋白以及极性蛋白复合物的影响,并且部分合子基因组激活(zygote genome activation,ZGA)和谱系分化特异性基因也会调控极性建立。在早期胚胎发育过程中,极性建立是第一次细胞谱系分化的基础。它通过影响不对称细胞分裂、谱系分化因子的不对称定位以及Hippo信号通路的活性来调控谱系分离和形态发生。本文对哺乳动物早期胚胎极性建立及对谱系分化影响的相关研究进行了梳理和总结,并讨论了目前已有研究在调控机制和物种方面的不足,以期为阐明早期胚胎极性建立提供线索与系统性视角。

SPP1、DEC1、C1QTNF6蛋白与口腔鳞状细胞癌患者临床病理指标及预后的关系

目的:探讨口腔鳞状细胞癌患者血清重组人分泌型磷蛋白1(SPP1)、软骨分化的表达基因1(DEC1)和补体C1q/肿瘤坏死因子相关蛋白6(C1QTNF6)的表达水平与其临床病理指标及预后的关系。方法:免疫组织化学染色法和电化学发光免疫分析法检测88例口腔鳞状细胞癌患者癌组织和血清中SPP1、DEC1和C1QTNF6蛋白的表达水平;Pearson相关性分析和Kaplan-Meier生存分析法分析患者血清SPP1、DEC1和C1QTNF6蛋白表达水平与其临床病理指标的相关性和对患者预后的影响。多因素Cox回归法分析影响口腔鳞状细胞癌患者预后的危险因素。结果:免疫组织化学染色结果显示口腔鳞状细胞癌患者癌组织中SPP1、DEC1和C1QTNF6蛋白的阳性表达率较癌旁组织分别增加了1.94、2.98和2.35倍(P<0.05或P<0.01);电化学发光免疫分析法结果显示口腔鳞状细胞癌患者血清SPP1、DEC1和C1QTNF6蛋白表达水平较正常人分别上调了8.61、6.20和4.03倍(P<0.05或P<0.01);Pearson相关性分析结果显示患者血清SPP1、DEC1和C1QTNF6蛋白表达水平与患者发生多灶嗜神经侵犯、肿瘤浸润程度、淋巴结转移、较高的TNM分期呈正相关(P<0.05);Kaplan-Meier生存分析结果显示,血清SPP1、DEC1和C1QTNF6蛋白高表达水平的口腔鳞状细胞癌患者总生存率低;多因素Cox回归分析结果表明多灶嗜神经侵犯、肿瘤高浸润度、淋巴结转移和高的TNM分期是影响预后的危险因素(P<0.05)。结论:口腔鳞状细胞癌患者癌组织和血清中SPP1、DEC1和C1QTNF6蛋白表达水平升高,且与患者发生多灶嗜神经侵犯、肿瘤浸润和淋巴结转移、较高的TNM分期呈正相关,而与患者预后呈负相关。

单个植入前胚胎mRNA差异显示方法的建立

在已有的研究基础上 ,优化和建立了单个植入前胚胎的mRNA差异显示 (singlepreimplantationembryodifferentialdisplaypolymerasechainraction ,SPEDDRT PCR) .以小鼠单个成熟的卵母细胞和单个的 2细胞期胚胎作为起始材料 ,比较二者之间基因的表达差异 .选择在卵母细胞中不表达而在 2细胞期表达的差异片段 .进行GenBank检索和表达序列标签 (EST)库电子克隆 .与多胚研究方法一样 ,由线粒体编码的和 2细胞特异表达的NADH脱氢酶亚单位 2和ATPase 6证实了SPEDDRT PCR方法是可行而且可靠的 ,是一种需要材料极少。