牛蒡多糖对K562细胞增殖的抑制及其机制的探讨

目的研究牛蒡多糖对K562细胞增殖的抑制作用并初步探索其机制。方法 MTT法检测牛蒡多糖对K562细胞增殖的抑制作用,RT-PCR检测BCL-2mRNA、Bax mRNA的表达。结果牛蒡多糖能明显抑制K562细胞的增殖;BCL-2基因表达下调,Bax的基因表达增多。结论牛蒡多糖对K562细胞增殖有抑制作用,其机制可能与BCL-2基因表达下调,Bax的表达上调有关。     

MOMP in the absence of BH3-only proteins

The minimum requirement for mitochondrial apoptosis has been controversial ever since the discovery of BCL-2 as a cell death regulator. In this issue of Genes & Development, O''Neill and colleagues (pp. 973–988) end a long-standing debate by creating a cellular system free of BCL-2 family proteins, thereby identifying the outer mitochondrial membrane rather than BH3-only proteins as the only requirement for BAX/BAK activation and mitochondrial outer membrane permeabilization (MOMP).     

芍药苷对醋酸铅诱导海马神经元凋亡及Bcl-2/Bax蛋白表达的影响

目的 探讨芍药苷(paeoniflorin,PF)对醋酸铅诱导海马神经元凋亡及Bcl-2/Bax蛋白表达的影响。方法 分离培养胎鼠海马神经元细胞,细胞免疫荧光染色鉴定纯度。MTT测定海马神经元细胞活力以确定醋酸铅最适造模浓度及时间,同时筛选合适剂量PF干预海马神经元凋亡。依据MTT测定结果,分为空白组、模型组和20,40,80 μmol·L^-1 PF组干预海马神经元细胞,作用24 h后,加醋酸铅染毒,检测细胞色素C (cytochrome C,Cyt-C)含量、线粒体膜电位及细胞内Ca²⁺浓度。流式细胞仪检测细胞凋亡情况,Western blotting测定海马神经元细胞中caspase-3、cleaved-caspase-3、caspase-8、cleaved-caspase-8、caspase-9、cleaved-caspase-9、Bax、Bcl-2蛋白表达水平。结果 细胞免疫荧光染色鉴定结果显示分离培养的细胞为海马神经元细胞,且纯度较高。MTT测定结果显示醋酸铅最适造模浓度及时间为25 μmol·L^-1染毒24 h;PF剂量为20,40,80 μmol·L-1可显著改善海马神经元细胞活性,呈剂量依赖性。与空白组相比,模型组Cyt-C含量、凋亡率、细胞内Ca²⁺浓度显著升高(P<0.01),线粒体膜电位显著降低(P<0.01)。与模型组相比,40 μmol·L^-1 PF组和80 μmol·L^-1 PF组可降低Cyt-C含量、凋亡率、细胞内Ca²⁺浓度(P<0.05或P<0.01),升高线粒体膜电位(P<0.01),20 μmol·L^-1PF组可显著升高线粒体膜电位(P<0.05)。此外,一定剂量的PF可下调cleaved-caspase-3、cleaved-caspase-8、cleaved-caspase-9、Bax蛋白表达,上调Bcl-2蛋白表达。结论 PF可抑制醋酸铅诱导的海马神经元凋亡,可通过调控Bcl-2/Bax蛋白表达发挥神经保护作用。     

全反式维甲酸对大鼠肺气肿模型的早期干预作用

目的 研究全反式维甲酸(ATRA)对木瓜蛋白酶所致大鼠肺气肿模型的早期干预作用.方法 采用木瓜蛋白酶气管内滴注的方法,将体质量170～220 g 60只SPF级SD大鼠随机分为对照组(N),模型组(R),ATRA治疗组(R1),棉籽油组(R2)各15只.后3组向大鼠气管内滴注5％木瓜蛋白酶1 ml/kg,N组同样滴注等量生理盐水,14d后,分别向R1组和R2组大鼠腹腔内连续注入1 000 μg/kgATRA和棉籽油,第90天杀死全部大鼠.取出肺组织,行HE染色观察各组大鼠的肺脏的病理变化,和病理形态学图像分析了解肺气肿模型是否建立及ATRA对于木瓜蛋白酶复制的大鼠肺气肿模型的早期干预效果.行免疫组织化学染色,计算各组Ⅱ型肺泡上皮细胞(AECⅡ)凋亡率,及Bax蛋白、Bcl-2蛋白阳性细胞个数,比较各组之间AECⅡ凋亡率,及Bax蛋白、Bcl-2蛋白阳性细胞个数的差异.结果 肺体积结果显示:R、R1、R2组的肺体积与N组比较,明显增大；R、R2组之间差异无统计学意义.肺泡形态学结果显示:R、R1、R2与N组比较,每个视野内的肺泡数明显减少,平均肺泡面积增大,平均内衬间隔较大,R2组相对于R组,肺泡形态学各项指标差异无统计学意义.R、R1、R2组与N组比较,大鼠肺组织AECⅡ凋亡明显增多,R1与R组比较,AECⅡ凋亡率明显下降,R2与R组比较,AECⅡ凋亡率无明显变化,R、R1、R2组在造模后肺泡壁细胞Bcl-2阳性细胞百分比显著高于N组,R组肺泡壁细胞Bcl-2阳性细胞百分比显著低于R1组,R组肺泡壁细胞Bcl-2阳性细胞百分比与R2组差异无统计学意义.R、R1、R2组在造模后肺泡壁Bax蛋白阳性细胞百分比显著高于N组,R组肺泡壁细胞Bax蛋白阳性细胞百分比显著高于R1组,R组肺泡壁Bax蛋白阳性细胞百分比与R2组差异无统计学意义.R1组Bcl-2/Bax比例明显高于R、R2组,而与N组差异无统计学意义.结论 ATRA治疗可降低大鼠肺气肿模型AECⅡ凋亡率,调节Bax蛋白、Bcl-2蛋白表达,对大鼠肺气肿进程起到干预作用.     

Notch-activated signaling cascade interacts with mitochondrial remodeling proteins to regulate cell survival

Survival of differentiated cells is one of several processes regulated by Notch activity, although the general principles underlying this function remain to be characterized. Here, we probe the mechanism underlying Notch-mediated survival, building on emerging evidence that apoptotic responses coordinated by specialized intermediates converge on mitochondria, identifying a core event in death pathways. The Bcl-2 family protein Bax is one such intermediate, which in a unifying response to diverse apoptotic stimuli nucleates multiprotein assemblies on mitochondria, committing cells to irrevocable damage. Using Bax as the prototype stimulus, we analyze Notch signaling for potential interactions with mitochondria, probe intrinsic properties of the Notch receptor, and describe key intermediates in the Notch-activated signaling cascade. Ligand-dependent processing was necessary to generate the Notch intracellular domain (NIC) although signaling was independent of canonical interactions with nuclear factors. Notably, antiapoptotic activity was recapitulated by NIC recombinants, localized outside the nucleus, and compromised by enforced nuclear sequestration. NIC signaled via the kinase Akt to prevent the loss of mitochondrial function, contiguity, and consequent nuclear damage, outcomes critically depend on mitochondrial remodeling proteins Mitofusins-(Mfn)-1 and 2. Thus, the NIC-Akt-Mfn signaling cascade identifies a pathway regulating cell-survival, independent of canonical functions associated with NIC activity.     

高剂量灵芝孢子粉干预戊四氮致痫模型大鼠海马及皮质区神经细胞Bcl-2、Bax的表达

采用中药灵芝孢子粉低、中、高剂量（150, 300, 450 mg/kg）干预戊四氮致癫痫模型28 d，并设空白对照组。经免疫组织化学染色和TUNEL检测后发现，与模型相比，灵芝孢子粉高剂量组大鼠海马和皮质区神经细胞TUNEL阳性细胞数及Bax阳性细胞数均减少，而Bcl-2阳性表达细胞数增加。Morris水迷宫实验检测结果显示，灵芝孢子粉高剂量组大鼠逃避潜伏期缩短，跨过原平台次数增加。说明高剂量灵芝孢子粉上调癫痫模型大鼠海马和皮质区神经细胞Bcl-2蛋白的表达，抑制Bax免疫反应及癫痫所致的神经细胞凋亡，明显改善其学习记忆功能。     

Melatonin antagonizes the intrinsic pathway of apoptosis via mitochondrial targeting of Bcl-2

Abstract:  We have recently shown that melatonin antagonizes damage-induced apoptosis by interaction with the MT-1/MT-2 plasma membrane receptors. Here, we show that melatonin interferes with the intrinsic pathway of apoptosis at the mitochondrial level. In response to an apoptogenic stimulus, melatonin allows mitochondrial translocation of the pro-apoptotic protein Bax, but it impairs its activation/dimerization The downstream apoptotic events, i.e. cytochrome c release, caspase 9 and 3 activation and nuclear vesiculation are equally impaired, indicating that melatonin interferes with Bax activation within mitochondria. Interestingly, we found that melatonin induces a strong re-localization of Bcl-2, the main Bax antagonist to mitochondria, suggesting that Bax activation may in fact be antagonized by Bcl-2 at the mitochondrial level. Indeed, we inhibit the melatonin anti-apoptotic effect (i) by silencing Bcl-2 with small interfering RNAs, or with small-molecular inhibitors targeted at the BH3 binding pocket in Bcl-2 (i.e. the one interacting with Bax); and (ii) by inhibiting melatonin-induced Bcl-2 mitochondrial re-localization with the MT1/MT2 receptor antagonist luzindole. This evidence provides a mechanism that may explain how melatonin through interaction with the MT1/MT2 receptors, elicits a pathway that interferes with the Bcl-2 family, thus modulating the cell life/death balance.     

促甲状腺素受体抗体检测技术的回顾与展望

促甲状腺素受体抗体 (TRAb)与多种自身免疫性甲状腺疾病的发病密切相关 ,是临床诊断这些疾病的重要指标。目前已建立了两种较为成熟的TRAb检测技术 ,即根据TRAb可竞争抑制12 5I标记促甲状腺素 (TSH)与受体结合的原理设计的放射受体分析法 (RRA)和根据TRAb可模拟或阻断TSH作用影响体外细胞系cAMP产生的原理设计的生物分析法。RRA法简单易操作 ,灵敏度和特异度也比较高 ,在临床上应用广泛 ;而生物分析法则能够进一步区分甲状腺刺激性抗体和甲状腺刺激阻断性抗体 ,成为TRAb检测技术发展的重大突破。本文将对上述两种方法不同的诊断价值、局限性以及临床实用性作一综述。     

Apoptosis of human pancreatic cancer cells induced by Triptolide

AIM： To investigate apoptosis in human pancreatic cancer cells induced by Triptolide （TL）, and the relationship between this apoptosis and expression of caspase-3＇ bcl-2 and bax. METHODS： Human pancreatic cancer cell line SW1990 was cultured in DMEM media for this study. MTT assay was used to determine the cell growth inhibitory rate in vitro. Flow cytometry and TUNEL assay were used to detect the apoptosis of human pancreatic cancer cells before and after TL treatment. RT-PCR was used to detect the expression of apoptosis-associated gene caspase-3＇ bcl-2 and bax. RESULTS： TL inhibited the growth of human pancreatic cancer cells in a dose-and time-dependent manner. TL induced human pancreatic cancer cells to undergo apoptosis with typically apoptotic characteristics. TUNEL assay showed that after the treatment of human pancreatic cancer cells with 40 ng/mL TL for 12 h and 24 h, the apoptotic rates of human pancreatic cancer cells increased significantly. RT-PCR demonstrated that caspase-3 and bax were significantly up-regulated in SW1990 cells treated with TL while bcl-2 mRNA was not. CONCLUSION： TL is able to induce the apoptosis in human pancreatic cancer cells. This apoptosis may be mediated by up-regulating the expression of apoptosisassociated caspase-3 and bax gene.