小麂、黑麂、赤麂精母细胞联会复合体的比较研究

本工作以界面铺张——硝酸银染色技术,对小麂(Muntiacus reeuesi)、黑麂(M.crinifrons)和赤麂(M.muntjak)的精母细胞联会复合体(Syna ptonemal complex,SC)进行亚显微结构的比较研究。结果表明: 1.SC的平均相对长度和臂比指数同有丝分裂细胞相应染色体的数值有很好的一致性。根据SC的相对长度和臂比指数绘制了三种麂的SC组型图。雄性黑麂减数分裂前期形成一个复杂的易位多价体,意味着其核型的演化过程涉及两次染色体易位和一次臂间倒位。 2.在减数分裂前期,性染色体的形态和行为同常染色体的有明显差异,如性染色体嗜银性较强,配对延迟等。XY的配对起始于早粗线期,在中粗线期,Y的全长均同X配对;XY-SC开始解体于晚粗线期。 3.在粗线期,X染色体未配对区域出现自身折叠,形成“发夹”状结构。这种“发夹”结构的形成,可能是在性染色体的进化过程中,X染色体通过不对称易位得到的重复片段在减数分裂前期同源配对的一种细胞学表现。     

白腹锦鸡,红腹锦鸡,中国雉鸡SC组型的比较研究

以微铺展—硝酸银染色技术制备三种鸡的SC标本,进行电镜观察。结果表明:三种鸡的SC组非常相似,即2n=82,ZZ/ZW型性别决定,雄性为ZZ。除1号SC和Z-SC为中着丝粒外,其余均为端着丝粒。Z-SC的相对长度有明显的细胞间差异;平均相对长度度介于第3和第4号SC之间。三者SC组型上的差异主要表现在相应SC长度上的不同。并对其亲缘关系及在鸟类进化中的可能地位进行了讨论。此外,在微铺展法制备的锦鸡精母细胞SC标本中还发现了巨大中心粒,这在高等动物尚属首次。     

Polypeptide domains of ADP-ribosyltransferase obtained by digestion with plasmin

Proteolysis by plasmin inactivates bovine ADP-ribosyltransferase; therefore, enzymatic activity depends exclusively on the intact enzyme molecule. The transferase was hydrolyzed by plasmin to four major polypeptides, which were characterized by affinity chromatography and N-terminal sequencing. Based on the cDNA sequence for human ADP-ribosyltransferase enzyme [Uchida, K., Morita, T., Sato, T., Ogura, T., Yamashita, R., Noguchi, S., Suzuki, H., Nyunoya, H., Miwa, M., & Sugimura, T. (1987) Biochem. Biophys. Res. Commun. 148, 617-622], a polypeptide map of the bovine enzyme was constructed by superposing the experimentally determined N-terminal sequences of the isolated polypeptides on the human sequence deduced from its cDNA. Two polypeptides, the N-terminal peptide (Mr 29,000) and the polypeptide adjacent to it (Mr 36,000), exhibited binding affinities toward DNA, whereas the C-terminal peptide (Mr 56,000), which accounts for the rest of the transferase protein, bound to the benzamide-Sepharose affinity matrix, indicating that it contains the NAD+-binding site. The fourth polypeptide (Mr 42,000) represents the C-terminal end of the larger C-terminal fragment (Mr 56,000) and was formed by a single enzymatic cut by plasmin of the polypeptide of Mr 56,000. The polypeptide of Mr 42,000 still retained the NAD+-binding site. The plasmin-catalyzed cleavage of the polypeptide of Mr 56,000-42,000 was greatly accelerated by the specific ligand NAD+. Out of a total of 96 amino acid residues sequenced here, there were only 6 conservative replacements between human and bovine ADP-ribosyltransferase.     

Molecular interactions between DNA, poly(ADP-ribose) polymerase, and histones

Molecular interactions between purified poly(ADP-ribose) polymerase, whole thymus histones, histone H1, rat fibroblast genomic DNA, and closed circular and linearized SV40 DNA were determined by the nitrocellulose filter binding technique. Binding of the polymerase protein or histones to DNA was augmented greatly when both the enzyme protein and histones were present simultaneously. The polymerase protein also associated with histones in the absence of DNA. The cooperative or promoted binding of histones and the enzyme to relaxed covalently closed circular SV40 DNA was greater than the binding to the linearized form. Binding of the polymerase to SV40 DNA fragments in the presence of increasing concentrations of NaCl indicated a preferential binding to two restriction fragments as compared to the others. Polymerase binding to covalently closed relaxed SV40 DNA resulted in the induction of superhelicity. The simultaneous influence of the polymerase and histones on DNA topology were more than additive. Topological constraints on DNA induced by poly(ADP-ribose) polymerase were abolished by auto ADP-ribosylation of the enzyme. Benzamide, by inhibiting poly(ADP-ribosylation), reestablished the effect of the polymerase protein on DNA topology. Polymerase binding to in vitro-assembled core particle-like nucleosomes was also demonstrated.     

Inhibition of carcinogen-induced cellular transformation of human fibroblasts by drugs that interact with the poly(ADP-ribose) polymerase system. Initial evidence for the development of transformation resistance

Two types of interactions of 13 drugs with human fibroblasts were determined: I50 of nuclear poly(ADP-ribose) polymerase, as assayed with isolated nuclei in vitro, and the non-toxic concentration of drugs that prevented carcinogen-induced cell transformation of intact fibroblasts (RCF1). In general, RCF1 was much lower than I50, and one antitransformer did not inhibit the enzyme in vitro, indicating that low-affinity enzyme inhibitory sites appear to play no role in the mechanism of prevention of cell transformation. Two enzyme inhibitors, caffeine and 1-methylnicotinamide, exhibited no antitransforming activity. Benzamide when applied in population doubling 1 induced resistance to cell transformation in population doubling 6 by carcinogens added at this stage.     

苯胺嘧啶衍生物X-9通过下调整合素β1表达抑制肝细胞癌迁移侵袭

整合素在许多肿瘤细胞中高表达,并且参与肿瘤细胞的侵袭转移。在肝细胞癌中,整合素β1被报导高表达,并促进肿瘤细胞的侵袭。目前,对于整合素的表达调控癌细胞机制以及干预其表达进而抑制肿瘤细胞转移的研究较少。本研究探讨利用小分子化合物抑制整合素表达来抑制肿瘤细胞迁移和侵袭的可能。首先,对临床肝癌细胞患者癌组织和癌旁组织中的整合素β1的表达进行检测,发现其在癌组织中的表达显著高于癌旁组织（P<0.05）。对TCGA肿瘤数据库的生物信息学分析结果同样显示,整合素β1的高表达与肝癌的分期（P=0.019）和预后（P=0.013）相关。通过筛选发现,苯胺嘧啶衍生物X09可以抑制肝癌细胞中整合素β1的mRNA和蛋白质的表达（P<0.01）。细胞划痕愈合实验和细胞穿孔实验结果显示,苯胺嘧啶衍生物X-9能够抑制肝癌细胞的迁移和侵袭（P<0.01）。进一步的研究证实,在肝癌细胞中外源表达整合素β1可以逆转X-9对肝癌细胞迁移和侵袭的抑制;而在敲低整合素β1的细胞中,X-9对细胞的迁移和侵袭的抑制被消除。因此,鉴定出苯胺嘧啶衍生物X-9可以通过下调整合素β1表达,进而抑制肝癌细胞的迁移和侵袭。     

电针引起脊髓P物质释放的频率依赖性

我室以往的研究表明,不同频率的电针可引起脊髓释放不同种类的阿片肽。本工作观察Ｐ物质（ＳＰ）释放的频率依赖性,电针频率选择２,４,８,１５,３０和１００Ｈｚ,分别收集电针期间及电针前后各３０ｍｉｎ的脊髓灌流液,通过放射免疫方法测定大鼠电针有效组和电针无效组Ｐ物质免疫活性（ＳＰ－ｉｒ）．结果如下：（１）电针有效组：２Ｈｚ引起ＳＰ－ｉｒ降低,与电针前相比,Ｐ＜０．０１;４Ｈｚ电针前后ＳＰ－ｉｒ比较,无统计学意义;８,１５,３０,１００Ｈｚ时ＳＰ－ｉｒ均增加（Ｐ＜０．０１）,其中１５Ｈｚ时ＳＰ增加最多（Ｐ＜０．００１）,表明刺激引起ＳＰ释放有频率依赖性。（２）电针无效组：不论应用何种频率,电针前后脊髓灌流液中ＳＰ－ｉｒ变化不大（均Ｐ＞０．０５）。提示,电针时脊髓液中ＳＰ含量变化与镇痛效果有密切关系。