棉酚对体外人肥大前列腺成纤维细胞的作用

棉酚对体外人肥大前列腺成纤维细胞的生长、ＤＮＡ合成及形态学的作用结果表明：棉酚可以明显地抑制细胞的生长，且抑制作用与时间－剂量相关；用＾３Ｈ－胸腺嘧啶掺入法测定细胞的ＤＮＡ合成，结果也相同；细胞超微结构的改变主要为细胞质空泡化，内质网、线粒体的损害和核固缩，其中线粒体对棉酚最为敏感，损害最明显。作者还对棉酚抑制体外人肥大前列腺成纤维细胞增殖的作用机理及棉酚用于治疗前列腺肥大症的可能性进行了初步的探     

β—L—2‘,3’—双脱氧—5—氟胞苷在体外对乙型肝炎病毒复制的抑制作用

目的：研究β－Ｌ－２＇，３＇－双脱氧－５－氟胞苷的抗乙型肝炎病毒作用。方法：从２．２．１５细胞培养液中提取乙肝病毒ＤＮＡ，用ＤＮＡ印迹法分别药物对乙肝病毒ＤＮＡ复制的影响。观察药物对人Ｔ－成淋巴样细胞的生长抑制作用，用狭线印迹法检测人Ｔ－成淋巴样细胞的线粒体ＤＮＡ。结果：β－Ｌ－２＇，３＇－双脱氧－５－氟胞苷对乙肝病毒ＤＮＡ合成的半数抑制浓度为０．０５μｍｏｌ／Ｌ，但其抗乙肝病毒作用是可逆的。对人     

苏拉明对体外培养瘢痕疙瘩成纤维细胞增殖的影响

目的：探讨苏拉明对瘢痕疙瘩成纤维细胞生长及代谢的作用。方法：利用细胞培养技术将成纤维细胞从瘢痕疙瘩组织中分离出来进行培养,分别利用绘制细胞生长曲线,测定细胞ＤＮＡ合成代谢等手段来观察ｓｕｒａｍｉｎ以细胞生长及代谢的影响。结果：ｓｕｒａｍｉｎ对体外培养条件下的瘢痕疙瘩成纤维细胞的生长及ＤＮＡ的合成均有明显的抑制作用,但对细胞的形态没有明显影响。结论：在体外培养条件下ｓｕｒａｍｉｎ可以抑制瘢痕疙瘩成纤     

胸腺细胞体外培养对细胞凋亡和细胞周期进程的影响

本文采用流式的细胞仪研究了小鼠胸腺细胞体外培养对细胞内ＤＮＡ合成、细胞周期以及细胞凋亡的影响。结果表明：胸腺细胞体外培养引起的ＤＮＡ合成抑制，细胞有丝分裂延迟，体外培养后４小时即表现出明显的ＤＮＡ合成抑制。８－１２ｈ抑制深，２４ｈ蛋白质的合成开始恢复，细胞凋亡则表现时间依赖性增高。     

重组人肿瘤坏死因子对人结肠癌SW－480细胞在体外的杀伤作用研究

文中报道了重组人肿瘤坏死因子（ｒｈＴＮＦ）对人结肠癌细胞株ＳＷ－４８０的克隆抑制和细胞毒作用。改良噻唑兰（ＭＴＴ）法表明ＴＮＦ在６００００ｕ／ｍｌ时才有明显的细胞毒作用说明ＳＷ－４８０对ＴＮＦ低度敏感，而且无剂量依赖关系．但克隆抑制试验表明１０ｕ／ｍｌ就可明显抑制ＳＷ－４８０集落的生长，并有剂量依赖性，且在剂量１５０ｕ／ｍｌ以上时抑制作用随时间而增强．３Ｈ－ＴｄＲ掺入测定的ＴＮＦ抑制ＳＷ－４８０细胞ＤＮＡ合成和ＭＴＴ法测定的细胞毒曲线一致说明ＴＮＦ对ＳＷ－４８０的主要作用是抑制ＤＮＡ合成．ＴＮＦ对ＳＷ－４８０抑制后超微结构变化：最初是胞膜出现微孔，然后细胞内线粒体变性，溶解，核固缩，溶解，最后细胞溶解．这些结果与ＭＴＴ法细胞毒试验及３Ｈ－ＴｄＲ掺入试验相结合说明ＴＮＦ对ＳＷ－４８０杀伤作用主要针对细胞核内外染色体的ＤＮＡ合成上．     

维拉帕米对人单核巨噬细胞蛋白质、DNA、RNA合成的影响

目的：观察维拉帕米对人单核巨噬细胞ＤＮＡ、ＲＮＡ及蛋白质合成过程的影响。方法：维拉帕米与人单核巨噬细胞孵育２４ｈ或４ｄ，测定细胞３Ｈ－ＴｄＲ、３Ｈ－ＵＲ、３Ｈ－Ｌｅｕ掺入人单核巨噬细胞状态及细胞内蛋白质含量。结果：维拉帕米在低浓度时对ＤＮＡ、ＲＮＡ合成有抑制作用，呈浓度依赖性；高浓度时作用明显，对ＲＮＡ作用较ＤＮＡ显著（Ｐ＜０．０５）；对蛋白质合成在高浓度时有抑制作用。维拉帕米与成熟过程中人单核巨噬细胞作用４ｄ，细胞蛋白含量显著降低（Ｐ＜０．０５），并呈浓度依赖性。结论：此作用可能为维拉帕米抑制细胞生长及动脉粥样斑块形成逆转心肌肥厚的机制之一。     

沙棘汁对白血病细胞的杀伤作用

不同浓度的沙棘汁分别于Ｋ５６２、ＨＬ－６０和ＹＡ－１肿瘤细胞体外培养２４ｈ，可使ＹＡＣ－１、ＨＬ－６０细胞的ＤＮＡ合成有抑制作用，并似有剂量效应关系；对Ｋ５６２细胞ＤＮＡ中的ＵｄＲ有释放作用，亦即沙棘汁对瘤细胞的毒性作用。沙棘汁对白血病细胞生长抑制作用的机理可能与沙棘汁对其ＤＮＡ合成的解聚作用和抑制合成作用有关。     

沙棘汁对白血病细胞的杀伤作用

不同浓度的沙刺分别于Ｋ５６２、ＨＬ－６０和ＹＡＣ－１肿瘤细胞体外培养２４ｈ，可使ＹＡＣ－１、ＨＬ－６０细胞的ＤＮＡ合成抑制作用，并似有剂量效应关系；对Ｋ５６２细胞ＤＮＡ中的ＵｄＲ有释放作用，亦即沙棘汁对瘤细胞的毒性作用。沙棘汁对白血病细胞生长抑制作用的机理可能与沙刺汁对ＤＮＡ合成的解聚作用和抑制合成作用有关。     

金属蛋白酶MMP—9表达与人黑色素瘤细胞系转移表型的相关性

目的：探讨金属蛋白酶ＭＭＰ－９与肿瘤转移的相关性，方法：利用基因重组技术构建反义ＭＭＰ－９ｃＤＮＡ真核表达载体，用脂质体法转染ＭＭＰ－９高表达的转移性人黑色素瘤细胞株ＷＭ４５１，检测转染后细胞ＭＭＰ－９表达水平的改变以及体外生长、侵袭、裸鼠体内成瘤及自发转移能力的变化。结果：转染细胞ＭＭＰ－９的表达及活性明显下降，同时ＭＭＰ－２的表达也受到一定抑制，细胞生长速度、体外侵袭能力及裸鼠体内成瘤性及自发     

二甲亚砜和维甲酸对人卵巢癌细胞COC-1的逆转作用研究

以人卵巢癌细胞株ＣＯＣ－１为模型，观察了诱导分化剂二甲亚砜（ＤＭＳＯ）与维甲酸（ＲＡ）对该细胞生长增殖、ＤＮＡ合成和超微结构的影响。结果显示：ＤＭＳＯ和ＲＡ可明显抑制ＣＯＣ－１细胞的生长和ＤＮＡ合成，并改变其恶性细胞的结构特征，使之向正常细胞的方向逆转，从而提示ＤＭＳＯ和ＲＡ对人卵巢癌细胞有一定的诱导分化作用。     

HUMAN ANTIFE.RTILITY EFFECT OF GOSSYPOL CYTOLOGIC OBSERVATION OF SEMEN

Monthly semen samples of 28 fertile men treated with 15-20 mg/day enteric-coated gossy- pol tablets, formic acid gossypol or acetic acid gossypol were examined by light microscope and electron microscope until antifertility was reached. Some cases were also studied ultra- histochemically for adenosine triphosphatase activity. The controls were the subjects' own samples before treatment. By counting the morphologic changes of the spermatozoa, it was found that gossypol may cause marked 30% t0 69% increases in abnorma- lities l.5 months after the start of administration. Electron microscope showed that the heads of aberrant spermatozoa often presented with large vacuoles with membranous structures in the nu- clei and the acrosomes of some cells were abnor- mal or even completely lacking. The middle- pieces had irregularly arranged mitochondria and the cristae were decreased or even absent and the matrix became paler. The mitochondria were vacuolated. Control ATPase positive reac- tions on the outer and inner mitochondrial mem- branes were in general stronger than in the gossypol treated and the vacuolated mitochond- ria always were negative. As the drug took effect, the spermatogenic cells, mainly spermatids, were markedly increas- ed t0 10-100% and some spermatids had 2 t0 4 nuclei. Sertoli's cells were seen in the semen. In general, the higher the cell differentia- tion, the more gossypol sensitive it is.     

Effects of gossypol acetate, progesterone, danazol and GnRH-A (gonadotropin release hormone-A) on the DNA-synthesis of human endometrial cell in vitro

Both aberrant endometrial cell and endometrial cell in situ, showed cyclic changes under the control of ovarian hormones. In this study, in order to observe a direct effect of medical therapy on the endometriosis, endometrial cells were cultured in vitro. Sensitivity test to 10(-6) M gossypol acetate, progesterone, danazol and GnRH-A were performed using determination of DNA synthesis by incorporation of tritiated thymidine. The results showed these drugs (but not GnRH-A) decreased the amount of attached cells, and lowered the DNA synthesis of endometrial cell in vitro (P less than 0.01). The inhibition rates of gossypol acetate was higher than that of progesterone and danazol, while that of progesterone was higher than that of danazol (P less than 0.01). The relationship between the growth inhibitory effect of progesterone and danazol showed a significant correlation for all points (r = 0.89, P less than 0.05). It has been suggested that some drugs might act a direct inhibitory effect on endometrial cell, such effect might be one of the modes in the treatment of endometriosis. GnRH-A had no direct suppression on endometrial cell in vitro, it has got clinical effect through the central effect at the hypothalamo-pituitary-ovary axis in treatment of endometriosis.     

醋酸棉酚对人舌鳞癌Cal-27细胞侵袭性作用的实验研究

  目的  研究醋酸棉酚（gossypol acetic acid，GAA）对体外培养的人舌鳞癌Cal-27细胞侵袭影响及其作用机制。  方法  制备0 μmol/L、2.5 μmol/L、5.0 μmol/L、10.0 μmol/L的GAA作用于人舌鳞癌Cal-27 细胞，通过Transwell实验检测GAA对人舌鳞癌Cal-27 细胞侵袭能力的影响；通过qRT-PCR及Western blot实验检测GAA对人舌鳞癌Cal-27 细胞基质金属蛋白酶-2（Matrix Metalloproteinase-2，MMP-2）、基质金属蛋白酶-9（Matrix Metalloproteinase-9，MMP-9）mRNA及蛋白的表达变化。  结果  Transwell实验显示:GAA对人舌鳞癌Cal-27细胞细胞侵袭产生抑制作用，实验组和对照组比较差异有统计学意义（P < 0.05）；qRT-PCR实验及Western blot显示，实验组人舌鳞癌Cal-27细胞中MMP-2、MMP-9 mRNA及蛋白的表达降低，与对照组相比差异有统计学意义（P < 0.05）。  结论  GAA能抑制人舌鳞癌Cal-27细胞的侵袭性，并降低人舌鳞癌Cal-27细胞MMP-2、MMP-9的mRNA及蛋白表达水平，这可能是GAA抑制肿瘤侵袭作用的机制之一。     

醋酸棉酚诱导鼠前列腺癌细胞凋亡的研究

目的研究醋酸棉酚对鼠前列腺癌RM-1细胞的抑制作用和诱导凋亡作用,探讨其作用机制。方法将不同浓度的醋酸棉酚作用于鼠前列腺癌RM-1细胞,利用MTT法检测其有效作用浓度,分别采用电子显微镜及流式细胞仪观察和检测RM-1细胞凋亡和bax蛋白表达情况,RT-PCR检测mRNA表达水平变化。结果MTT法测得其IC50值为34.56mg／L,浓度为30mg／L的醋酸棉酚具有诱导鼠前列腺癌RM-1细胞凋亡的作用,且随药物作用时间的延长,凋亡率逐渐增加,同时bax蛋白表达也逐渐增加;RT-PCR显示bax-mRNA水平逐渐升高。结论醋酸棉酚能诱导鼠前列腺癌细胞凋亡,其作用机制之一是上调bax-mRNA水平、增加bax蛋白表达,且具有时间依赖性。     

棉酚对纯化LDH-X作用的分析

用纯化的大鼠LDH-X和LDH 1～5,以定量测定酶活性和凝胶电泳相配合的分析方法,发现棉酚对LDH-X的抑制作用显著大于LDH1～5,与在人体内的试验结果基本一致,再次证实了棉酚对LDH-X的抑制有特异性,进一步明确通过抑制精子特异的LDH-X酶,中断 精子的能量供给,损害线粒体功能,阻断生精过程是棉酚抗生育作用机理的一个重要方面。     

卵巢储备功能下降患者颗粒细胞mtDNA拷贝数 及4 977 bp缺失的研究

目的：研究卵巢储备功能下降（DOR）患者颗粒细胞线粒体DNA(mtDNA)拷贝数和4 977 bp缺失情况,初步探讨DOR患者颗粒细胞mtDNA结构的完整性。方法：选取DOR组及对照组各50例,分离提纯卵泡液中颗粒细胞,提取DNA,RT-PCR相对定量颗粒细胞mtDNA拷贝数及PCR检测其4 977 bp缺失情况。结果：DOR组及对照组均未检测到mtDNA 4 977 bp缺失,其拷贝数相对量差异亦无统计学意义（P＞0.05）。结论：DOR患者颗粒细胞mtDNA结构基本完整,颗粒细胞可作为DOR患者卵母细胞浆内线粒体自体移植的供体细胞。     

HSP70抑制H2O2所致C2C12肌原细胞Smac从线粒体的释放及细胞凋亡

目的：阐明HSP70对H2O2所致C2C12肌原细胞凋亡的影响及其作用机制。方法：采用基因瞬间转染技术使细胞HSP70或/和Smac高表达，采用Hoechst 33258染色观察H2O2 (0.5mmol/L)处理转基因C2C12肌原细胞不同时间后细胞核形态学改变并计算凋亡核百分率。DNA抽提及琼脂糖电泳观察凋亡特征性梯带。采用caspase活性定量检测试剂盒及蛋白质印迹分析caspase-9和caspase-3的活化。利用免疫荧光分析HSP70对H2O2所致Smac从线粒体释放的影响。结果：HSP70对H2O2及Smac所致C2C12肌原细胞凋亡具有明显的抑制作用,凋亡核百分率明显降低, DNA电泳未出现明显“梯状”条带;caspase-9和caspase-3的活化明显受到抑制;并进一步发现HSP70可抑制H2O2所致C2C12肌原细胞Smac从线粒体释放。结论：HSP70可通过抑制Smac从线粒体释放来抑制H2O2所致C2C12肌原细胞凋亡。     

Role of mitochondria in neuron apoptosis during ischemia-reperfusion injury

To investigate the role of mitochondria in neuronal apoptosis, ischemia-reperfusion mediated neuronal cell injury model was established by depriving of glucose, serum and oxygen in media. DNA fragmentation, cell viability, cytochrome C releasing, caspase3 activity and mitochondrial transmembrane potential were observed after N2a cells suffered the insults. The results showed that N2a cells in ischemic territory exhibited survival damage, classical cell apoptosis change, DNA ladder and activation of caspase3. Apoptosis-related alterations in mitochondrial functions, including release of cytochrome C and depression of mitochondrial transmembrane potential (△ψm) were testified in N2a cells after mimic ischemia-reperfusion. Moreover, activation of caspase3 occurred following the release of cytochrome C. However, the inhibitor of caspase3, Ac-DEVDCHO, couldn‘t completely rescue N2a cells from apoptosis. Administration of cyclosporine A, an inhibitor of mitochondria permeability transition pore only partly inhibited caspase3 activity and reduced DNA damage. Interestingly, treatment of Z-IETD-FMK, an inhibitor of caspase8 could completely reverse DNA fragmentation, but can‘t completely inhibit caspase3 activity. It was conclud edthat there were caspase3 dependent and independent cellular apoptosis pathways in N2a cells suffering ischemia-reperfusion insults. Mitochondria dysfunction may early trigger apoptosis and amplify apoptosis signal.     

醋酸棉酚对Raji细胞增殖和凋亡的影响

目的 观察醋酸棉酚对人Burkitt淋巴瘤细胞株Raji细胞增殖和凋亡的影响.方法 采用台盼蓝染色和噻唑蓝(MTT)法观察醋酸棉酚对Raji细胞生长存活的影响,瑞氏染色法观察药物处理后细胞形态学变化,琼脂糖凝胶电泳和Annexin V-FITC标记流式细胞仪检测细胞凋亡,流式细胞仪分析细胞周期、检测细胞凋亡率及Bcl-2蛋白表达水平变化,比色法测定Raji细胞Caspase-3样蛋白酶活性.结果 5 μmol/L以上浓度醋酸棉酚能抑制Raji细胞生长增殖并诱导其凋亡,效应与药物浓度及给药时间呈正相关.醋酸棉酚作用后,Raji细胞被阻滞于G_0/G_1期.比色法显示Caspase-3样蛋白酶活化.流式细胞仪检测到Bcl-2蛋白表达下调,且与Caspase-3活化在时间上存在同步趋势.结论 醋酸棉酚能抑制aaji细胞增殖并诱导凋亡,其原因可能与调节细胞周期及下调Bcl-2蛋白表达有关.     

经输精管向附睾内注射甲酸棉酚的抗生育研究

对72只Wistar大鼠经输精管向附睾内注射甲酸棉酚,观察附睾中精子数量、形态、移动距离和附睾、睾丸的组织学改变。结果:注射12h后精子移动距离缩短,24h后精子计数下降,组织学观察见附睾管呈无精状态,睾丸曲细精管各级生精细胞出现变性等损伤性改变,精母细胞脱落入曲细精管管腔,注射甲酸棉酚后2～40d内交配的雌鼠全部不孕。