布鲁氏菌对RAW264.7细胞内质网应激与细胞因子分泌的影响

【目的】布鲁氏菌与宿主相互作用的分子机制是目前的研究热点之一,布鲁氏菌通过形成来自于内质网的布氏小体而在巨噬细胞内生存和增殖,其机制目前尚不清楚,宿主细胞内质网应激反应对病原感染和炎症的调控密切相关。揭示内质网应激反应在布鲁氏菌感染巨噬细胞中的作用以及布鲁氏菌感染对巨噬细胞分泌免疫因子的影响。【方法】构建布鲁氏菌感染RAW264.7模型,在感染后不同时间收集细胞,通过实时荧光定量PCR检测细胞内质网应激反应标志分子GRP78和CHOP,以及TNF-α、IL-1β和IL-6在m RNA水平的变化;通过Western blot和ELISA分别检测其蛋白水平的变化。【结果】布鲁氏菌感染RAW264.7细胞的最佳感染复数MOI为100?1;证明在布鲁氏菌感染4-6 h,巨噬细胞可杀伤侵入的布鲁氏菌,之后存活的细菌可在细胞内增殖;感染后24 h出现细胞凋亡,48 h出现大量细胞坏死。布鲁氏菌感染可激活RAW264.7细胞的内质网应激反应,促进GRP78的表达,同时,抑制免疫因子的分泌。【结论】内质网应激反应参与了RAW264.7对布鲁氏菌感染的调节。     

香菇多糖注射液联合CHOP化疗治疗弥漫性大B细胞淋巴瘤的疗效及对Bcl-6、Ki-67、VEGF、β2-MG表达的影响

目的:探讨香菇多糖注射液联合CHOP化疗治疗弥漫性大B细胞淋巴瘤的疗效及对B细胞淋巴瘤因子-6(Bcl-6)、Ki-67抗原(Ki-67)、内皮细胞生长因子(VEGF)、β2微球蛋白(β2-MG)表达的影响。方法:选取2010年1月~2011年12月我院收治的64例弥漫性大B细胞淋巴瘤患者作为研究对象,按治疗方式进行分组,采用香菇多糖注射液联合CHOP化疗治疗的患者为观察组,单纯采用CHOP化疗治疗的患者为对照组。比较两组患者短期疗效、治疗前后Bcl-6、Ki-67、VEGF、β2-MG的表达情况、不良反应的发生情况及生存情况。结果:治疗后,观察组治疗总有效率显著高于对照组[84.38%(27/32)vs.53.13(17/32)](P0.05);肿瘤组织Bcl-6阳性表达率高于对照组[84.38%(27/32)vs.62.50%(20/32)](P0.05),Ki-67、VEGF、β2-MG阳性表达率均低于对照组[31.25%(10/32)vs.59.38%(19/32),28.13%(9/32)vs.56.25%(18/32),18.75%(6/32)vs.40.63%(13/32)](P0.05);不良反应总发生率低于对照组[21.88%(7/32)vs.46.88%(15/32)](P0.05);第5年随访时存活率高于对照组[59.38%(19/32)vs.34.38%(11/32)](P0.05)。结论:香菇多糖注射液联合CHOP化疗方案治疗弥漫性大B细胞淋巴瘤可有效提高近期疗效、生存率及生存质量,且安全性高,可能与其改善机体肿瘤组织中Bcl-6、Ki-67、VEGF、β2-MG的表达有关。     

Identification of 1,2,3-triazole derivatives that protect pancreatic β cells against endoplasmic reticulum stress-mediated dysfunction and death through the inhibition of C/EBP-homologous protein expression

The C/EBP-homologous protein (CHOP) acts as a mediator of endoplasmic reticulum (ER) stress-induced pancreatic insulin-producing β cell death, a key element in the pathogenesis of diabetes. Chemicals that inhibit the expression of CHOP might therefore protect β cells from ER stress-induced apoptosis and prevent or ameliorate diabetes. Here, we used high-throughput screening to identify a series of 1,2,3-triazole amide derivatives that inhibit ER stress-induced CHOP-luciferase reporter activity. Our SAR studies indicate that compounds with an N,1-diphenyl-5-methyl-1H-1,2,3-triazole-4-carboxamide backbone potently protect β cell against ER stress. Several representative compounds inhibit ER stress-induced up-regulation of CHOP mRNA and protein, without affecting the basal level of CHOP expression. We further show that a 1,2,3-triazole derivative 4e protects β cell function and survival against ER stress in a CHOP-dependent fashion, as it is inactive in CHOP-deficient β cells. Finally, we show that 4e significantly lowers blood glucose levels and increases concomitant β cell survival and number in a streptozotocin-induced diabetic mouse model. Identification of small molecule inhibitors of CHOP expression that prevent ER stress-induced β cell dysfunction and death may provide a new modality for the treatment of diabetes.     

Rab7a modulates ER stress and ER morphology

The Endoplasmic Reticulum (ER) is a membranous organelle with diverse structural and functional domains. Peripheral ER includes interconnected tubules, and dense tubular arrays called “ER matrices” together with bona fide flat cisternae. Transitions between these states are regulated by membrane-associated proteins and cytosolic factors. Recently, the small GTPases Rab10 and Rab18 were reported to control ER shape by regulating ER dynamics and fusion. Here, we present evidence that another Rab protein, Rab7a, modulates the ER morphology by controlling the ER homeostasis and ER stress. Indeed, inhibition of Rab7a expression by siRNA or expression of the dominant negative mutant Rab7aT22?N, leads to enlargement of sheet-like ER structures and spreading towards the cell periphery. Notably, such alterations are ascribable neither to a direct modulation of the ER shaping proteins Reticulon-4b and CLIMP63, nor to interactions with Protrudin, a Rab7a-binding protein known to affect the ER organization. Conversely, depletion of Rab7a leads to basal ER stress, in turn causing ER membrane expansion. Both ER enlargement and basal ER stress are reverted in rescue experiments by Rab7a re-expression, as well as by the ER chemical chaperone tauroursodeoxycholic acid (TUDCA). Collectively, these findings reveal a new role of Rab7a in ER homeostasis, and indicate that genetic and pharmacological ER stress manipulation may restore ER morphology in Rab7a silenced cells.