花生凝集素受体在中华绒螯蟹生精细胞的分布
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摘要
以中华绒螯蟹为材料,应用凝集素印迹法检测中华绒螯蟹生精细胞中的花生凝集素受体.以凝集素组织化学技术为手段,采用生物素标记的花生凝集素对中华绒螯蟹精子发生过程中花生凝集素受体进行了标记,研究精子发生过程中花生凝集素受体的分布特征.结果表明,中华绒螯蟹生精细胞中的花生凝集素受体的分子质量为72 ku;精子发生过程中,不同的生精细胞膜有不同程度的阳性标记.精原细胞质膜上有较强的阳性标记,呈深棕色;精母细胞的质膜阳性标记减少;精细胞与精母细胞染色程度相似.PNA受体在生精细胞中分布,可能与生殖细胞发育以及生殖细胞之间或者生殖细胞与体细胞之间的信息传递有关.精子质膜、顶体区域也有阳性标记,这可能与精卵识别及结合等受精过程息息相关.
In order to detect the distribution characteristics of peanut agglutinin(PNA) receptors in spermatogenic germ cells of Eriocheir sinensis, lectin-blotting and lectin-histochemistry were used in this study. The results showed that the molecular weight of PNA receptors was 72 ku. Based on the binding sites of the biotin-labeled PNA, positively stained cells could be observed and recognized as surface receptors present in all types of spermatogenic cells. But, there were different degrees of PNA-postive singals emerged on these cells' membrane. On spermatogonial cell surface, positive staining was strong, in dark brown. Positive signals were similar and weakened on spermatocytes and spermatids. The distribution characteristics of PNA receptors on spermatogenic cells may be related to the development of germ cells as well as the information transmission between germ cells or between germ cells and somatic cells. There were also positive signals on sperm surface and in its acrosome. These PNA receptors may be closely related to the fertilization, such as sperm-oocyte recognition and binding.
In order to detect the distribution characteristics of peanut agglutinin(PNA) receptors in spermatogenic germ cells of Eriocheir sinensis, lectin-blotting and lectin-histochemistry were used in this study. The results showed that the molecular weight of PNA receptors was 72 ku. Based on the binding sites of the biotin-labeled PNA, positively stained cells could be observed and recognized as surface receptors present in all types of spermatogenic cells. But, there were different degrees of PNA-postive singals emerged on these cells' membrane. On spermatogonial cell surface, positive staining was strong, in dark brown. Positive signals were similar and weakened on spermatocytes and spermatids. The distribution characteristics of PNA receptors on spermatogenic cells may be related to the development of germ cells as well as the information transmission between germ cells or between germ cells and somatic cells. There were also positive signals on sperm surface and in its acrosome. These PNA receptors may be closely related to the fertilization, such as sperm-oocyte recognition and binding.
引文
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[2] 官继承,李倩,左福元.细胞表面糖复合物对生殖的影响[J].黑龙江畜牧兽医,2012(7):20-22.DOI :10.13881/j.cnki.hljxmsy.2012.07.006.
[3] PENG H M,LI C P,LIU H,et al.Extraction of agglutinin from Oncomelania hupensis and its haemagglutination activity[J].Chinese Journal of Parasitology & Parasitic Diseases,2011,29(2):130-133.DOI :10.1007/s10008-010-1224-4.
[4] BELLANDE K,BONO J J,SAVELLI B,et al.Plant lectins and lectin receptor-like kinases:how do they sense the outside[J].International Journal of Molecular Sciences,2017,18(6):1164-1188.DOI :10.3390/ijms18061164.
[5] 王静静.芸豆植物凝集素及其制剂的生殖生物学相关性质研究[D].昆明:云南师范大学,2017.
[6] 杨成焕.红芸豆植物凝集素(RKBL)终止小鼠中期妊娠作用机理的初步研究[D].昆明:云南师范大学,2015.
[7] NAND A,SINBH V,WANG P,et al.Glycoprotein profiling of stem cells using lectin microarray based on surface plasmon resonance imaging[J].Analytical Biochemistry,2014,465:114-120.DOI :10.1016/j.ab.2014.07.028.
[8] 欧阳霞辉,奚耕思.动物精子发生及成熟过程中凝集素受体的分布与变化[J].陕西师范大学学报(自然科学版),2003(S2):226-229.DOI :10.15983/j.cnki.jsnu.2003.s2.046.
[9] BENOFF S.Carbohydrates and fertilization:an overview[J].Molecular Human Reproduction,1997,3(7):599-637.DOI :10.1093/molehr/3.7.599.
[10] 潘建梅,奚耕思.动物卵子膜凝集素受体及其在生殖中的功能[J].陕西师范大学学报(自然科学版),2003(S2):234-236.DOI :10.15983/j.cnki.jsnu.2003.s2.047.
[11] 韩学敏.荆豆凝集素-1受体及花生凝集素受体在兔和犬子宫内的分布及其激素调节[D].长春:吉林大学,2007.
[12] SAKAMAKI K,SAWADA K,KOSHIMIZU U,et al.Identification of peanut agglutinin receptors on mouse testicular germ cells[J].Biology of Reproduction,1989,41(6):1097-1102.DOI :10.1095/biolreprod41.6.1097.
[13] DEY S K,LIM H,DAS S K.Molecular cues to implantation[J].Endocrine Reviews,2004,25(3):341-373.DOI :10.1210/er.2003-0020.
[14] 潘建梅.中华稻蝗配子发生中花生凝集素受体和原癌基因c-kit表达的研究[D].西安:陕西师范大学,2004.
[15] RAYCHOUDHURY S S.Multiple fucosyltransferases and their carbohydrate ligands are involved in spermatogenic cell-Sertoli cell adhesion in vitro in rats[J].Biology of Reproduction,1997,56(5):1268-1273.DOI :10.1095/biolreprod56.5.1268.
[16] CHANDLER K B,COSTELLO C E.Glycomics and glycoproteomics of membrane proteins and cell-surface receptors:Present trends and future opportunities[J].Electrophoresis,2016,37(11):1407-1419.DOI :10.1002/elps.201500552.
[17] 欧阳霞辉,奚耕思.中华蚱蜢精子发生过程中花生凝集素受体的分布与变化[J].动物学杂志,2006,41(1):7-11.DOI :10.13859/j.cjz.2006.01.002.
[18] HARAYAMA H,WATANABE S Y,MASUDA H,et al.Lectin-binding characteristics of extracts from epididymal boar spermatozoa[J].Journal of Reproduction and development,1998,44(1):21-27.DOI :10.1262/jrd.44.21.
[19] CHANDLER K B,COSTELLO C E.Glycomics and glycoproteomics of membrane proteins and cell-surface receptors:present trends and future opportunities[J].Electrophoresis,2016,37(11):1407-1419.DOI :10.1002/elps.201500552.
[20] TECLE E,GAGNEUX P.Sugar-coated sperm:Unraveling the functions of the mammalian sperm glycocalyx[J].Molecular Reproduction & Development,2015,82(9):635-650.DOI :10.1002/mrd.22500.
[21] 杨万喜,堵南山,赖伟.日本沼虾生精细胞与支持细胞之间的连接关系[J].动物学报,1999,45(2):178-186.DOI :10.3969/j.issn.1674-5507.1999.02.009.
[22] HEIMANN P,LAAGE S,JOCKUSCH H.Defect of sperm assembly in a neurological mutant of the mouse,wobbler (WR)[J].Differentiation,1991,47(2):77-83.DOI :10.1111/j.1432-0436.1991.tb00225.x.
[23] CROSS N L,MEIZEL S.Methods for evaluating the acrosomal status of mammalian sperm[J].Biology of Reproduction,1989,41(4):635-641.DOI :10.1095/biolreprod41.4.635.
[24] 章其琪,MURPHY R.彩色染色揭示精子的秘密[J].国外畜牧学(猪与禽),2003,23(4):42-43.DOI :10.3969/j.issn.1001-0769.2003.04.012.
[25] GUYONNET B,ZABETMOGHADDAM M,SANFRANCISCO S,et al.Isolation and proteomic characterization of the mouse sperm acrosomal matrix[J].Molecular & Cellular Proteomics Mcp,2012,11(9):758-774.DOI :10.1074/mcp.M112.020339.