小鼠精子表面Con A结合糖复合物的形成与变化

用辣根过氧化物酶标记的ＣｏｎＡ（伴刀豆素Ａ）对小鼠睾丸与附睾切片,以及对取自附睾和子宫（交配后）内的精子涂片进行了标记,旨在认识精子在发生、成熟和获能过程中表面糖复合物的形成与变化。本研究表明,睾丸内的生精细胞和支持细胞均呈ＣｏｎＡ标记阳性。附睾的输出小管和附睾管上皮细胞,ＣｏｎＡ标记呈中度至强阳性,有部位的差别。附睾头和附睾尾内精子表面的标记无明显差别,标记位置均主要在顶体区和尾部。精子在子宫内存留１．５小时后,顶体后区出现中度阳性标记,但存留３小时和６小时后,顶体和顶体后区的标记均减弱或消失。这些结果提示,（１）精子发生期即可合成ＣｏｎＡ结合糖复合物,（２）精子在附睾成熟过程中表面的ＣｏｎＡ结合糖复合物无明显变化,（３）精子获能后顶体后区出现的ＣｏｎＡ结合糖复合物可能与受精能力有关。     

短额负蝗卵子发生过程中糖复合物的动态分布

以生物素标记的凝集素(UEA-I、SBA、PNA)为探针,利用凝集素组织化学方法对短额负蝗(Atracto-morphasinensis)卵子发生过程中滤泡细胞和卵母细胞内糖复合物的分布进行了定位研究。结果表明,在卵子发生的各期滤泡细胞和卵母细胞中没有UEA-I受体的表达,SBA和PNA受体以不同的分布模式呈阶段性表达。两者首次出现于卵母细胞生长期,随后PNA受体消失,SBA受体大量表达;在卵黄形成期前期SBA受体和重新出现的PNA受体表达于卵黄颗粒形成部位,卵黄形成期后期两者均为阴性表达;成熟卵子中两种受体又以不同程度重新出现于卵黄膜。两种受体在滤泡细胞内均大量表达。提示,N-乙酰半乳糖胺和半乳糖-β-(1,3)半乳糖胺复合物的修饰和变化与卵母细胞的发育、卵黄物质的形成及滤泡细胞的增殖分化密切相关,卵黄膜上的糖复合物可能与精卵识别有关。     

短额负蝗卵子发生过程中糖复合物的动态分布

以生物素标记的凝集素(UEA-I、SBA、PNA)为探针，利用凝集素组织化学方法对短额负蝗(Atractomorpha sinensis)卵子发生过程中滤泡细胞和卵母细胞内糖复合物的分布进行了定位研究。结果表明，在卵子发生的各期滤泡细胞和卵母细胞中没有UEA-I受体的表达，SBA和PNA受体以不同的分布模式呈阶段性表达。两者首次出现于卵母细胞生长期， 随后PNA受体消失，SBA受体大量表达；在卵黄形成期前期SBA受体和重新出现的PNA受体表达于卵黄颗粒形成部位，卵黄形成期后期两者均为阴性表达；成熟卵子中两种受体又以不同程度重新出现于卵黄膜。两种受体在滤泡细胞内均大量表达 提示，N-乙酰半乳糖胺和半乳糖-β-(1，3)半乳糖胺复合物的修饰和变化与卵母细胞的发育、卵黄物质的形成及滤泡细胞的增殖分化密切相关，卵黄膜上的糖复合物可能与精卵识别有关。     

小鼠精子表面SBA结合糖复合物的形成与变化

用ＨＲＰ标记的大豆凝集素（ＳＢＡ）对睾丸与附睾切片,以及取自附睾和子宫（交配后）内的精子进行了标记,旨在认识精子在发生、成熟和获能过程中表面糖复合物的形成与变化规律。在睾丸内,精母细胞和早期精子细胞胞质内有一强阳性颗粒,处于精子形成期的精子细胞呈弱阳性标记。附睾管内的精子团呈强阳性,附睾管上皮则仅在游离缘呈弱阳性。交配后１．５小时自子宫内洗出的精子,其顶体区的标记增至强阳性,但随着在子宫内存留时间的延长,标记强度逐渐减弱或消失。结果表明,１）精子表面的ＳＢＡ结合糖复合物出现于精子形成期;２）在成熟和获能过程中精子表面的ＳＢＡ结合糖复合物发生明显的变化     

短额负蝗精子发生过程中SBA受体的动态分布

用常规组织学方法观察短额负蝗Atractomorpha sinensis Bolivar精子发生过程中生精细胞的显微结构,并以大豆凝集素(soybean agglutinin,SBA)为探针利用凝集素细胞化学方法研究该过程中N-乙酰半乳糖复合物的分布变化。结果表明,短额负蝗精子发生经历了精原细胞增殖期、初级精母细胞期、次级精母细胞期、精子细胞形成期和精子成熟期5个时期,在这5个时期中各期生精细胞的大小、形态、核染色体等变化明显。在整个精子发生过程中,N-乙酰半乳糖复合物出现于精原细胞期,并于精母细胞期发生明显的修饰和变化,精子形成期和成熟期没有N-乙酰半乳糖复合物的表达。提示,N-乙酰半乳糖复合物的修饰和变化与短额负蝗生精细胞的生长和分化密切相关。     

类雄激素受体在尖唇散白蚁繁殖蚁和工蚁精子发生中的免疫细胞化学定位

为了比较和探讨类雄激素受体 (androgen receptor-like,AR-like)在白蚁生殖品级和非生殖品级精子发生过程中的作用,运用免疫细胞化学方法对尖唇散白蚁Reticulitermes aculabialis繁殖蚁和工蚁精子发生中的AR-like的定位进行了研究。结果显示：在繁殖蚁和工蚁精子发生过程中都有AR-like免疫阳性细胞的分布,均分布于初级精母细胞的细胞质和细胞核中;与繁殖蚁相比,AR-like在工蚁中的表达较弱。结果提示,AR-like的表达与精子发生过程中初级精母细胞的减数分裂有关,雄激素及其受体通过调控精母细胞的第一次减数分裂来影响白蚁精子的发生。虽然工蚁的精巢发育受抑制,但其精子发生和维持具有与繁殖蚁相同的激素调节机制,能形成精子。本研究为工蚁性腺退化不育而根据群体的变化又可以发育为补充繁殖蚁这一特殊的生理功能提供了组织学依据。     

几种植物生殖细胞质膜表面的凝集素受体荧光标记

为进一步探讨从生殖细胞到精子的发育过程中细胞质膜表面凝集素受体的可能变化,及其与两类对凝集素标记有不同结果的精子的关系,用异硫氰酸荧光素标记的伴刀豆凝集素(Con A)、麦芽凝集素(WGA)和大豆凝集素(SBA)对蚕豆(Vicia faba L．)、鸢尾(Iris tectorium Maxim．)和朱顶红(Hippeastrum vittatum Herb．)的生殖细胞质膜表面的凝集素受体进行标记。结果显示：在不同植物中均有部分生殖细胞不能被凝集素探针标记,且在保持尾状形态的生殖细胞的表面发现有凝集素受体的极性分布。这可能是导致部分精子表面不能被同种凝集素标记的重要原因。此外,同一种凝集素受体在不同物种的生殖细胞上分布不一致,不同的凝集素受体在同一种植物的生殖细胞上的分布模式亦有不同。在蚕豆和鸢尾的生殖细胞表面均有这三种凝集素的受体。在朱顶红生殖细胞的表面有前两种凝集素的受体,分布比较均一,但是没有大豆凝集素的受体。此外,在具尾生殖细胞表面发现有凝集素受体极性分布的现象,为探讨精细胞功能及其表面糖蛋白分布的可能差异提供了重要启示。     

糖蛋白物与发育

糖与蛋白质或脂类共价结合而成的糖蛋白、蛋白聚糖、糖脂以及脂多糖统称糖复合物 ,最近几年人们对糖复合物在生物发育过程中的作用进行了大量的研究 ,已经在许多方面取得了重大进展。1 .糖复合物与配子发生和受精配子发生和受精过程中有大量糖复合物的参与 ,他们在配子发生、精卵识别与受精以及受精完成后防止多精穿入的皮层反应等过程中发挥作用。哺乳动物精子表面有一层几百种糖蛋白组成的糖萼 ,其成熟是精子成熟的标志。射出的精子头部外表面的糖蛋白能阻止顶体酶的释放 ,在获能过程中该糖蛋白被雌性生殖管道分泌物中的酶降解后精子才获…     

精子发生过程中组蛋白甲基化和乙酰化

精子发生(Spermatogenesis)这一高度复杂的独特分化过程包括精原细胞发育为精母细胞、单倍体精细胞的形成和精子成熟,并以阶段特异性和睾丸特异性基因的表达、有丝分裂和减数分裂以及组蛋白向鱼精蛋白的转变为特征。表观遗传修饰在减数分裂重组、联会复合物的形成、姊妹染色体的结合、减数分裂后精子的变态、基因表达阻遏和异染色质形成过程中发挥着重要作用。其中具有一定组成形式、起抑制作用和/或激活作用的组蛋白甲基化和乙酰化标记,不仅保证了正确的染色体配对和二价染色体的成功分离,并且精确调节减数分裂特异性基因的适时表达。精子发生过程中组蛋白甲基化和/或乙酰化错误会直接影响表观遗传修饰的建立和维持,导致生精细胞异常甚至引发不育。文章旨在对精子发生过程中组蛋白甲基化和乙酰化表观遗传修饰的动态变化及其相关酶的调节机制进行综述,为进一步研究精子发生的表观遗传调控,预防男性不育疾病的发生提供基础资料。     

北方山溪鲵精子发生不同阶段的显微与超微结构

用光镜和电镜观察了北方山溪鲵(Batrachuperus tibetanus)精子发生过程中各种类型生精细胞的显微与超微结构变化。结果显示,北方山溪鲵在4~8月时处于精子发生期,精子形成在7~8月。成熟精子的结构具有小鲵科精子的一些共同特征,如顶体前端呈三叶草状,尾部由轴纤维、波动膜、轴丝及轴丝旁纤维构成,轴纤维粗大呈圆柱形,尾部无线粒体等。比较分析认为,在两栖类的系统发育中,轴纤维、波动膜和轴丝旁纤维的消失为近裔性状。     

中华蚱蜢卵子发生中花生凝集素受体的初步鉴定和发育表达

The distributions of PNA binding glycoconjugates in the plasma membrane of Acrida cinerea Thunberg germ cells were detected using biotin labeled PNA, for better understanding of the formation and changes of glycoconjugates during oogenesis. The ultrastructure of vitellogenesis also was observed by electron microscopy for detection of the origin and track of vitelline material. In the ovary, PNA receptors appeared in the oocyte cytoplasm of the second phases of oogenesis; positive granules gradually increased from the third phase to the fourth, and they exhibited a maximum expression before the vitellogennic stage in the cytoplasm of the oocyte. From the vitellogennic to chorionation stage, positive granules gradually declined. Binding sites on follicle cells were changed with their morphological variation in every stage of oogenesis. The vitelline of A. cinerea formed within the oocyte by degrees. The results suggest that PNA receptors and yolk materials are synthesized by the oocytc at an early period. With the development of the oocyte, some exogeous materials from two sources act as PNA receptors and others take part in vitelline synthesis. One is blood lymph that offers some useful materials to the oocyte directly through follicle cell gaps; the other are follicle cells that produce and transmit some materials to oocyte to support vitellogenesis. In addition, PNA receptors secreted by follicle cells participate in the formation of yolk membrane [ Acta Zoologica Sinica 5 l （5） ： 932 - 939, 2005 ].     

Flow cytometric characterization of viable meiotic and postmeiotic cells by Hoechst 33342 in mouse spermatogenesis.

BACKGROUND: Spermatogenesis in adult is a complex stepwise process leading to terminally differentiated spermatozoa. The cellular heterogeneity of testis renders complex the studies on molecular aspects of this differentiation process. Analysis of the regulation of adult spermatogenesis would undoubtedly benefit from the development of techniques to characterize each germinal differentiation step. METHODS: Hoechst 33342 staining of mouse testicular cells allows characterization of an enriched population in germinal stem cell and spermatogonia, called side population. In this study, we examined the definition of the various germinal populations stained by Hoechst 33342, notably meiotic and postmeiotic cells. RESULTS: Preleptotene spermatocytes, spermatocyte I, spermatocyte II, and round and elongated spermatids were discriminated by Hoechst 33342 staining. In addition, we associated differentiation of spermatocyte I through leptotene to diplotene with changes in Hoechst 33342 red fluorescence pattern. CONCLUSIONS: Hoechst 33342 staining of viable germinal cells constitutes a valuable tool to study normal and impaired mouse adult spermatogenesis or to isolate viable cells from various differentiation stages for studies of molecular mechanisms regulating spermatogenesis.     

Temporal expression of c-kit in spermatogenesis of two grasshopper species

Two species of grasshoppers, Calliptamus abbreviatus （Ikonn.） and Shirakiacris shirakii （I. Bol.）, were collected randomly in the Siping area of Jilin Province, China. By using immunohistochemical methods and statistical analysis, we observed and compared the temporal expression of c-kit protein in four representative stages of spermatogenesis of the two grasshoppers, namely： spermatogonia; primary spermatocyte; secondary spermatocyte; and mature sperm. Results showed that there was c-kit positive temporal expression at each stage of spermatogenesis, but there were different positive expression levels： （i） weak positive expression of c-kit protein appeared in spermatogonia and the positive granules were thinner; （ii） strong positive expression of c-kit protein existed in primary spermatocyte and positive granules became biggest among all developmental stages; （iii） c-kit positive expression stayed stronger in secondary spermatocyte while positive granules became thinner; （iv） there was a strong positive expression of c-kit and thinner positive granules in mature sperm, which distributed on head and tail; （v） the biggest c-kit positive granules had been found massing at the end of spermary; and （vi） significant differences of c-kit positive expression existed in spermatogenesis between two species of grasshoppers. The results indicated that c-kit protein may play a crucial role in spermatogenesis and even retain the physiological action of sperms and fertilization in grasshoppers.     

白肛海地瓜精子发生及形态的超微结构

通过透射和扫描电镜观察了白肛海地瓜(Acaudina leucoprocta)的精子发生过程及其形态结构,揭示了白肛海地瓜精子发生时期一系列变化,其精子发生分为精原细胞、初级精母细胞、次级精母细胞、精细胞、成熟精子5个时期。精原细胞体积最大。精母细胞染色质开始凝集。精细胞前顶体颗粒形成。白肛海地瓜成熟精子的超微结构为原生型,由头部、中部、尾部组成,头部圆形,最前端为顶体,核染色质凝集成团块状,中部是线粒体和中心粒复合体融合成1个超大结构,尾部长约60μm,尾部鞭毛横切面为典型的"9+2"型结构。     

三角帆蚌精子的发生

报道了光镜和透射电镜下三角帆蚌精子的发生过程及其一系列重要的形态变化。包括核延长,染色质浓缩,线粒体逐渐融合并后移;胞质减少及鞭毛形成,精原细胞是精巢中体积最大的细胞,细胞膜界限不明显,内质网发达,精母细胞开始出现中心粒,精细胞分化可分为3个阶段。成熟精子属原生型,由头部、中段和尾部三部分组成。     

Lectin binding pattern in normal human labial mucosa

Summary The pattern of lectin binding in normal human labial mucosa was examined by light and electron microscopy using eight different lectins (ConA, LCA, WGA, UEA-1, RCA-1, SBA, DBA and PNA) and compared with the patterns in normal human skin and oesophageal mucosa. As seen by light microscopy, ConA, LCA, and WGA stained cell membranes in all layers of the mucosae. RCA-1 stained the plasma membrane of cells in the basal and middle layers, whereas cells in the superficial layers showed little positive staining. UEA-1, SBA, and PNA stained the cells in the middle layers weakly in some cases. No positive staining for DBA was seen. By electron microscopy, reaction product indicating ConA-binding sites was observed in the plasma membrane, cisternae of the endoplasmic reticulum, nuclear envelope and the Golgi apparatus. Binding of LCA, WGA, and RCA-1 was observed in the plasma membrane. These results show that the binding pattern of PNA, SBA, and RCA-1 in labial mucosa is different from that in the normal skin or oesophageal mucosa, although the labial mucosal epithelium, epidermis, and oesophageal epithelium are all stratified squamous epithelia. These differences in the cell-surface sugar residues are likely to be related to the possible functional differences in these tissues.     

Capping and ultrastructural localization of sperm surface isoantigens during spermatogenesis.

Isoantisera from female rabbits injected with rabbit whole semen have been used to study the appearance of cell surface isoantigens during spermatogenesis. Using isoantiserum IgG and adjuvant control IgG the presence of surface isoantigens on separated pachytene spermatocyte populations and populations of cells at more advanced stages of differentiation was confirmed with fluorescein-labeled goat IgG anti-rabbit IgG. The label was uniformly dispersed over the cell surface on cells labeled at 4°C but occurred in caps on cells warmed to 37°C indicating isoantigen mobility within the plane of the membrane. Residual bodies and mature spermatozoa did not show cap formation. Spermatogonia, Leydig cells, and Sertoli cells were not labeled. These observations were confirmed at the ultrastructural level with peroxidase-conjugated goat IgG anti-rabbit IgG. The percentage of the cell surface labeled was determined on cells at specific stages of spermatogenesis by stereological analysis. No significant surface labeling was observed on spermatogonia, Leydig cells, or Sertoli cells. The percentage of label bound to the surface of spermatogenic cells increased from approximately 4% in the pachytene spermatocytes to greater than 96% in the most mature testicular spermatids.     

Evidence for the association of two cell surface glycoproteins of 13762 mammary ascites tumor cells : Concanavalin A-induced redistribution of peanut agglutinin-binding proteins

The behavior of the cell surface concanavalin A (conA) receptors and of peanut agglutinin (PNA) receptors on the MAT-B1 ascites subline of the 13762 rat mammary adenocarcinoma was examined using fluorescein-labeled conA and PNA. ASGP-1, the major glucosamine-containing glycoprotein of these ascites cells, is the only PNA-binding protein observed by dodecyl sulfate electrophoresis. ASGP-2, the second most prominent component after glucosamine labeling, is the most abundant conA-binding protein. These two glycoproteins were previously shown to be associated as a complex in detergent extracts of the cells [20]. ConA-binding proteins, upon incubation with fluorescein-labeled conA (FITC-conA), redistribute on the cell surface into small and large aggregates similar, but not identical, to those seen in ‘patching’ and ‘capping’ experiments with lymphocytes. PNA-binding proteins failed to redistribute during incubation with fluorescein-labeled PNA (FITC-PNA) and appeared in a diffusely stained pattern around the circumference of the cells. However, when cells were treated with unlabeled conA followed by FITC-PNA, or with FITC-PNA followed by unlabeled conA, there was marked redistribution of the FITC-PNA. These results indicate that ASGP-1 redistributes in response to the movement of conAbinding proteins and supports our hypothesis that ASGP-1 and ASGP-2 are associated on the plasma membrane at the cell surface as well as in detergent extracts.