脂多糖对肺动脉内皮细胞Ⅰ型Na+/H+交换器表达的影响

目的 :探讨脂多糖 （ LPS）对培养的牛肺动脉内皮细胞 （ PAEC） 型 Na+ /H+交换器 （ NHE-1）表达的影响及其与内皮细胞死亡的关系。方法 :体外培养牛 PAEC,MTT比色检测细胞存活率 ,并提取总 RNA,应用反转录聚合酶链式反应 （ RT-PCR）及 DNA电泳技术 ,以 NHE-1与β-actin电泳条带光密度值之比 （ e/c）作数据分析 ,以其比值反映 NHE-1m RNA的表达程度。应用 Fluo3 /Amy荧光探针标记 ,激光共聚焦观察细胞内钙离子的浓度。结果 :在L PS（ 0 .0 1～ 1.0 μg/ml）作用 48h后 ,MTT比色吸光度值分别为 0 .3 5± 0 .0 7,0 .3 1± 0 .0 7,0 .2 6± 0 .0 4,低于对照组 0 .44± 0 .0 6（ P<0 .0 1）。e/c分别为 0 .43± 0 .11,0 .83± 0 .14 ,1.2 2± 0 .19,高于对照组 0 .2 8± 0 .0 8（ P<0 .0 1,n=4）。激光共聚焦结果显示 L PS引起细胞内钙离子增多 ,呈剂量依赖型。结论 :LPS可以引起肺动脉内皮细胞NHE-1表达增加 ,抑制细胞增殖 ,导致细胞死亡 ,钙离子可能参与 NHE-1的激活及表达。     

Immunostimulatory DNA ameliorates experimental and spontaneous murine colitis

BACKGROUND & AIMS: Impaired mucosal barrier, cytokine imbalance, and dysregulated CD4(+) T cells play important roles in the pathogenesis of experimental colitis and human inflammatory bowel disease. Immunostimulatory DNA sequences (ISS-DNA) and their synthetic oligonucleotide analogs (ISS-ODNs) are derived from bacterial DNA, are potent activators of innate immunity at systemic and mucosal sites, and can rescue cells from death inflicted by different agents. We hypothesized that these combined effects of ISS-DNA could inhibit the damage to the colonic mucosa in chemically induced colitis and thereby limit subsequent intestinal inflammation. METHODS: The protective and the anti-inflammatory effect of ISS-ODN administration were assessed in dextran sodium sulfate-induced colitis and in 2 models of hapten-induced colitis in Balb/c mice. Similarly, these effects of ISS-ODN were assessed in spontaneous colitis occurring in IL-10 knockout mice. RESULTS: In all models of experimental and spontaneous colitis examined, ISS-ODN administration ameliorated clinical, biochemical, and histologic scores of colonic inflammation. ISS-ODN administration inhibited the induction of colonic proinflammatory cytokines and chemokines and suppressed the induction of colonic matrix metalloproteinases in both dextran sodium sulfate- and hapten-induced colitis. CONCLUSIONS: As the colon is continuously exposed to bacterial DNA, these findings suggest a physiologic, anti-inflammatory role for immunostimulatory DNA in the GI tract. Immunostimulatory DNA deserves further evaluation for the treatment of human inflammatory bowel disease.     

Bacterial cell wall components regulate adipokine secretion from visceral adipocytes

Recent studies suggest a relationship between intestinal microbiota and metabolic syndromes; however, the underlying mechanism remains unclear. To clarify this issue, we assessed the effects of bacterial cell wall components on adiponectin, leptin and resistin secretion from rat visceral adipocytes in vitro. We also measured the relative population of Firmicutes and Bacteroidetes in fecal microbiota and the amount of fecal mucin as an intestinal barrier function, when mice were fed a high-fat diet. In the present study, we demonstrated that bacterial cell wall components affect the secretion of adipokines, depending on the presence of antigens from gram-positive or gram-negative bacteria. Lipopolysaccharide markedly inhibited adiponectin, leptin, and resistin secretion, whereas peptidoglycan increased adiponectin secretion and decreased resistin secretion in vitro. In vivo experiments showed that the high-fat diet increased the population of Firmicutes and decreased that of Bacteroidetes. In contrast, the high-fat diet downregulated the stool output and fecal mucin content. These results demonstrate that bacterial cell wall components affect the onset of metabolic syndromes by mediating the secretion of adipokines from visceral adipose tissue. Furthermore, we believe that metabolic endotoxemia is not due to the increasing dominance of gram-negative bacteria, Bacteroidetes, but due to the depression of intestinal barrier function.     

LPS 对肝癌细胞株 HepG2增殖、凋亡及分泌炎症因子的影响

目的：体外观察不同浓度细菌内毒素脂多糖(LPS)对肝癌 HepG2细胞增殖、凋亡以及分泌炎症因子的影响,并探讨可能的机制。方法按照随机数字法将细胞分为对照组及1、10、50、100μg/ ml LPS 处理组。用 MTT 检测刺激24、48、72和96 h 后细胞的增殖能力;用流式细胞术检测刺激24 h 后细胞的凋亡情况;用 RT-PCR 法检测刺激24 h 后白介素-6(IL-6)和白介素-8(IL-8) mRNA 的表达含量;用化学发光法检测刺激24 h 后 IL-6和 IL-8的表达量,最后用 SPSS 19．0对数据进行统计学分析。结果与对照组比较,各处理组刺激24 h 和48 h 后细胞增殖活性明显升高(P <0．05),而在72、96 h 差异无统计学意义(P >0．05);刺激24 h 后,1、10、50和100μg/ ml LPS 处理组以及对照组之间的细胞凋亡率差异无统计学意义(P >0．05);刺激24 h 后,与对照组比较,随着刺激浓度的升高,IL-6和 IL-8的表达量明显升高(P <0．05)。结论 LPS 可以在48 h 内促进 HepG2的增殖,对HepG2细胞的凋亡没有影响,并且 LPS 作用可以上调 IL-6和 IL-8水平。     

ShcC proteins: Brain aging and beyond

To date, most studies of Shc family of signaling adaptor proteins have been focused on the near-ubiquitously expressed ShcA, indicating its relevance to age-related diseases and longevity. Although the role of the neuronal ShcC protein is much less investigated, accumulated evidence suggests its importance for neuroprotection against such aging-associated conditions as brain ischemia and oxidative stress. Here, we summarize more than decade of studies on the ShcC expression and function in normal brain, age-related brain pathologies and immune disorders with a focus on the interactions of ShcC with signaling proteins/pathways, and the possible implications of these interactions for changes associated with aging.     

脂多糖上调人牙髓细胞B细胞淋巴瘤-2蛋白及其相关X蛋白的表达

目的：探讨脂多糖（LPS）作用于人牙髓细胞后,B细胞淋巴瘤-2（Bc1-2）蛋白、Bcl-2相关X蛋白（Bax）在正常人牙髓和炎症牙髓中的表达,探讨其在牙髓炎症过程中的作用机制。方法在体外用不同质量浓度LPS刺激人牙髓细胞后,采用免疫细胞化学染色方法在不同的时间点检测Bcl-2、Bax的表达,用图像分析系统Simple?PCI?version?5.1分析。结果正常人牙髓细胞中Bcl-2、Bax表达量不高,而在牙髓炎症过程中两者均增强,但是随着LPS质量浓度的增加,Bcl-2的表达量开始下降,而Bax的表达则持续增强。结论内毒素能使人牙髓细胞膜上的Bcl-2、Bax表达增强,但Bax的表达强于Bcl-2,内毒素可能通过两者介导的信号传导途径引起细胞凋亡。     

Spironolactone inhibits production of proinflammatory mediators in response to lipopolysaccharide via inactivation of nuclear factor-κB

The effect of spironolactone (SPIR) on lipopolysaccharide (LPS)-induced production of proinflammatory mediators was examined using RAW 264.7 macrophage-like cells and mouse peritoneal macrophages. SPIR significantly inhibited LPS-induced production of nitric oxide (NO), tumor necrosis factor-α and prostaglandin E2. The inhibition was not mediated by cell death. SPIR reduced the expression of an inducible NO synthase mRNA in response to LPS. SPIR significantly inhibited phosphorylation of p65 nuclear factor (NF)-κB in response to LPS. Furthermore, SPIR inhibited phosphorylation of IκB kinase (IKK) as an upstream molecule of NF-κB in response to LPS. LPS did not induce the production of aldosterone in RAW 264.7 cells. Taken together, SPIR is suggested to inhibit LPS-induced proinflammatory mediators via inactivation of IKK/NF-κB in LPS signaling.     

The kiwi fruit peptide kissper displays anti‐inflammatory and anti‐oxidant effects in in‐vitro and ex‐vivo human intestinal models

Literature reports describe kiwi fruit as a food with significant effects on human health, including anti‐oxidant and anti‐inflammatory activity. Fresh fruit or raw kiwi fruit extracts have been used so far to investigate these effects, but the molecule(s) responsible for these health‐promoting activities have not yet been identified. Kissper is a kiwi fruit peptide displaying pore‐forming activity in synthetic lipid bilayers, the composition of which is similar to that found in intestinal cells. The objective of this study was to investigate the kissper influence on intestinal inflammation using cultured cells and ex‐vivo tissues from healthy subjects and Crohn's disease (CD) patients. The anti‐oxidant and anti‐inflammatory properties of kissper were tested on Caco‐2 cells and on the colonic mucosa from 23 patients with CD, by challenging with the lipopolysaccharide from Escherichia coli (EC‐LPS) and monitoring the appropriate markers by Western blot and immunofluorescence. EC‐LPS challenge determined an increase in the intracellular concentration of calcium and reactive oxygen species (ROS). The peptide kissper was highly effective in preventing the increase of LPS‐induced ROS levels in both the Caco‐2 cells and CD colonic mucosa. Moreover, it controls the calcium increase, p65‐nuclear factor (NF)‐kB induction and transglutaminase 2 (TG2) activation inflammatory response in Caco‐2 cells and CD colonic mucosa. Kissper efficiently counteracts the oxidative stress and inflammatory response in valuable model systems consisting of intestinal cells and CD colonic mucosa. This study reports the first evidence supporting a possible correlation between some beneficial effects of kiwi fruit and a specific protein molecule rather than generic nutrients.