Cu^(2+)、Zn^(2+)诱导稀有鮈鲫应激蛋白质的研究

以稀有鲫为材料 ,研究了应激蛋白质作为生物学指标的敏感性。结果表明 ,在无可观察效应浓度下 ,经 5d亚慢性胁迫暴露 ,以Cu2 +为胁迫因子 ,稀有鲫被诱导出约 54KDa的应激蛋白质 ;以Zn2 +为胁迫因子 ,稀有鲫被诱导出约 94KDa ,67KDa和 40KDa的应激蛋白质。应激蛋白质有可能成为一种生物学指标运用于生态风险性早期预警。

蛋白质组学技术筛选与鉴定鲤鱼嗅脑急性创伤后的应激蛋白质

优化建立鲤鱼(cyprinus carpio,CC)嗅觉端脑(嗅脑)全蛋白提取技术。联用低渗透裂解和液氮冻溶法破碎鲤鱼嗅脑组织(rhinencephalon tissue of cyprinus carpio,CCRT)、低速离心提取CCRT全蛋白,并采用双向凝胶电泳(2D-PAGE)技术进行有效分离。经分析与统计,每张CCRT的2D-PAGE图谱中的蛋白质斑点数目约为1200个。分别分离CCRT的脂溶性和水溶性全蛋白,并获得高分辨率的2D-PAGE图谱。选用差异蛋白质组学技术筛选经10%冰醋酸创伤后的CC,其端脑组织所表达出的6种应激蛋白质,并用肽质量指纹谱(peptide mass fingerprinting,PMF)和数据库检索技术给予鉴定。其中3种蛋白质为70S热休克蛋白、β微管蛋白和DNA链接酶IV,有望作为研究大脑急性创伤后的应激修复途径和机理的指示蛋白质。

干酪乳杆菌对数生长期与稳定期蛋白质组比较分析

研究建立干酪乳杆菌Lactobacillus casei XM2-1的蛋白质组双向电泳表达图谱,比较其在对数生长中期与稳定期初期的图谱差异。结果表明:有16个蛋白质点表达发生明显变化,经过基质辅助激光解吸/电离飞行时间质谱鉴定,其中有7个蛋白质点鉴定为应激蛋白质,分别为GroEL、DnaK、Hsp 20-1、Hsp 20-2和UspA蛋白质。可见,应激蛋白质可能在L.casei XM2-1稳定期对外界环境的抵御能力增强起到重要作用。

高温联合辐射对胚胎神经发育及脑热应激蛋白的影响

目的探讨高温联合辐射致胚胎神经发育与早期胚胎脑蛋白质和HSPs表达的关系。方法将130只孕鼠按随机抽签分为13组,每组10只。在鼠妊娠第8d,第9d分别接受60Co及γ-射线全身照射各1次,照射剂量1.0Gy;第10d各置温箱中,分别使其肛温保持在37±0.5℃,41±0.5℃,42±0.5℃,并持续3,4,5min。移出温箱2h后,每组每时段随机取5只脱颈法处死,采用Westerndotblot和lowry方法定量分析胚胎脑HSP70和蛋白质含量与胚胎神经系统畸形。结果高温联合辐射对胚胎神经毒性明显增加,并与高温强度及作用时间呈正相关;高温联合辐射作用于胚胎早期对胚胎脑蛋白质降低有复合作用(P<0.01),并可诱导脑HSP70合成。结论高温与辐射联合致神经发育与胚胎脑蛋白质合成及热应激蛋白的诱导有一定的相关性。

值得重视的禽类应激问题(一)

在现代化养禽生产中,养禽业的集约化本身就是增加应激反应的过程。由于应激因素的作用,使禽类机体内的生理环境和稳定性改变,产生了新的适应,以维持生命和恢复功能。亦即当禽类的行为发生变化时,禽类可通过应激进行适应。 自从Selye氏发现应激这一生理反应以来,对应激的研究日益深入。在禽类的应激方面,特别是对鸡的应激研究,则着重于应激对鸡的生产、生理。

Association of high expression in rat gastric mucosal heat shock protein 70 induced by moxibustion pretreatment with protection against stress injury

AIM:To study the effect of moxibustion on Zusanli or Liangmeng point on gastric mucosa injury in stress-induced ulcer rats and its correlation with the expression of heat shock protein 70 (HSP70). METHODS:Sixty healthy SD rats (30 males,30 females) were divided into control group,injury model group,Zushanli point group,Liangmeng point group. Stress gastric ulcer model was induced by binding cold stress method. Gastric mucosa ulcer injury (UI) index was calculated by Guth method. Gastric mucosa blood flow (GMBF) was recorded with a biological signal analyzer. Protein content and gene expression in gastric mucosal HSP70 were detected by immunohistochemistry (IHC) and reverse transcription polymerase chain reaction (RT-PCR). Thiobarbital method was used to determine malondialdehyde (MDA) content. Gastric mucosal endothelin (ET) and prostaglandin E2 (PGE2) were analyzed by radioimmunoassay. RESULTS:High gastric mucosal UI index,high HSP70 expression,low GMBF and PGF2,elevated MDA and ET were observed in gastric mucosa of rats subjected to cold stress. Moxibustion on Zusanli or Liangmeng point decreased rat gastric mucosal UI index,MDA and ET. Conversely,the expression of HSP70,GMBF,and PGE2 was elevated in gastric mucosa after pretreatment with moxibustion on Zusanli or Liangmeng point. The observed parameters were significantly different between Zusanli and Liangmeng points. CONCLUSION:Pretreatment with moxibustion on Zusanli or Liangmeng point protects gastric mucosa against stress injury. This protection is associated with the higher expression of HSP70 mRNA and protein,leading to release of PGE2 and inhibition of MDA and ET,impairment of gastric mucosal index.

Iron increases HMOX1 and decreases hepatitis C viral expression in HCV-expressing cells

AIM： To investigate effects of iron on oxidative stress, heme oxygenase-1 （HMOX1） and hepatitis C viral （HCV） expression in human hepatoma ceils stably expressing HCV proteins. METHODS： Effects of iron on oxidative stress, HMOX1, and HCV expression were assessed in CON1 cells. Measurements included mRNA by quantitative reverse transcription-polymerase chain reaction, and protein levels by Western blots. RESULTS： Iron, in the form of ferric nitrilotriacetate,increased oxidative stress and upegulated HMOX1 gene expression. Iron did not affect mRNA or protein levels of Bach1, a repressor of HMOXl. Silencing the up-regulation of HMOXl nuclear factor-erythroid 2-related factor 2 （Nrf2） by Nrf2-siRNA decreased FeNTA-mediated up-regulation of HMOXl mRNA levels. These iron effects were completely blocked by deferoxamine （DFO）. Iron also significantly decreased levels of HCV core mRNA and protein by 80%-90%, nonstructural 5A mRNA by 90% and protein by about 50% in the Con1 full length HCV replicon cells, whereas DFO increased them. CONCLUSION： Excess iron up-regulates HMOXl and down-regulates HCV gene expression in hepatoma cells. This probably mitigates liver injury caused by combined iron overload and HCV infection.

Wuling powder alleviates depressive-like behavior by attenuating endoplasmic reticulum stress in the mouse hippocampus

The response to endoplasmic reticulum(ER)stress has been noted in both human depression cases and depression models in rodents.Wuling powder,derived from the mycelium of the esteemed fungus Xylaria Nigripes(Kl.)Sacc,has demonstrated efficacy in alleviating depressive symptoms.The purpose of this research was to explore the antidepressant properties of Wuling powder and its basic molecular effects,particularly regarding alterations in ER stress.A model of social defeat stress was created by introducing a mouse to the cage of an unfamiliar,hostile mouse for intervals of 5–10 min daily over a span of 10 d.Subsequently,the mice received oral doses of Wuling powder for 2 weeks.The social approach-avoidance assay was employed to evaluate signs of depression-like behaviors.Moreover,protein and gene expressions linked to ER stress triggered by social defeat were analyzed through Western blotting analysis and quantitative real-time PCR.The behavioral tests indicated that Wuling powder ameliorated behaviors associated with depression due to social defeat stress.Treatment with Wuling powder markedly reduced the increased levels of the 78-k Da glucose-regulated protein and protein disulfide isomerase caused by social defeat stress.It also diminished the expression of inositol-requiring enzyme 1α(IRE1α)and spliced X box-binding protein-1(s XBP1)at the protein and m RNA levels.Furthermore,Wuling treatment notably decreased the levels of phosphorylated eukaryotic initiation factor 2 alpha kinase(P-e IF2α),activating transcription factor 4(ATF4)and C/EBP homologous protein(CHOP),simultaneously enhancing the ratio of B-cell lymphoma 2(Bcl-2)to Bcl-2-associated X protein(Bax).These results suggested that Wuling powder could alleviate ER stress and inhibit cell apoptosis in the hippocampus by inhibiting protein translation and synthesis,thereby attenuating depressive-like behavior.

Proteomic analysis of human epithelial ovarian cancer xenografts in immunodeficient mice exposed to chronic psychological stress

This work aims at comparing alterations in the proteomes of human epithelial ovarian cancer xenografts between stressed and non-stressed immunodeficient mice as well as exploring the molecular mechanisms linking chronic psychological stress to ovarian cancer oncogenesis and progression.SK-OV-3 cells were injected subcutaneously into nude mice.The stress group was subjected to a chronic physical restraint protocol for 6 h on 35 consecutive days,while the control group was unrestrained.All mice were sacrificed on day 36 after SK-OV-3 cell injection,and tumors were excised.Tumor tissues were processed for 2D gel electrophoresis,mass spectrometry(nanoUPLC-ESI-MS/MS) and Western blotting.The expression of 20 proteins was found to be significantly altered between the stress and control groups,of which 14 were up-regulated,five were down-regulated,and one protein was found only in the stress group.All proteins were identified by UPLC-ESI-MS/MS,and Western blotting results were consistent with those of proteomic methods.The present results provide new evidence relating to the molecular mechanism underlying the relationship between psychological stress and tumor progression.

Molecular signal networks and regulating mechanisms of the unfolded protein response

Within the cell, several mechanisms exist to maintain homeostasis of the endoplasmic reticulum （ER）. One of the primary mechanisms is the unfolded protein response （UPR）. In this review, we primarily focus on the latest signal webs and regulation mechanisms of the UPR. The relationships among ER stress, apoptosis, and cancer are also discussed. Under the normal state, binding immunoglobulin protein （BiP） interacts with the three sensors （protein kinase RNA-like ER kinase （PERK）, activating transcription factor 6 （ATF6）, and inositol-requiring enzyme la （IREla）） Under ER stress, misfolded proteins interact with BiP, resulting in the release of BiP from the sensors. Subsequently, the three sensors dimerize and autophosphorylate to promote the signal cascades of ER stress. ER stress includes a series of positive and negative feedback signals, such as those regulating the stabilization of the sensors/BiP complex, activating and inactivating the sensors by autophosphorylation and dephosphorylation, activating specific transcription factors to enable selective transcription, and augmenting the ability to refold and export. Apart from the three basic pathways, vascular endothelial growth factor （VEGF）-VEGF receptor （VEGFR）-phospholipase C-~ （PLCy）-mammalian target of rapamycin complex 1 （mTORC1） pathway, induced only in solid tumors, can also activate ATF6 and PERK signal cascades, and IREla also can be activated by activated RAC-alpha serine/threonine-protein kinase （AKT）. A moderate UPR functions as a pro-survival signal to return the cell to its state of homeostasis. However, persistent ER stress will induce cells to undergo apoptosis in response to increasing reactive oxygen species （ROS）, Ca2＋ in the cytoplasmic matrix, and other apoptosis signal cascades, such as c-Jun N-terminal kinase （JNK）, signal transducer and activator of transcription 3 （STAT3）, and P38, when cellular damage exceeds the capacity of this adaptive response.

Altered expression of micro RNAs in the response to ER stress

MicroRNAs, a class of small noncoding RNAs, play key roles in diverse biological and pathological processes. ER stress, resulting from the accumulation of unfolded or misfolded proteins in the ER lumen, is triggered by various physiological events and pathological insults. Here, using RNA deep sequencing analysis, we found that the expression of some microRNAs was altered in HeLa and HEK293 cells under ER stress. Protein and RNA levels of DGCR8, Drosha, Exportin-5, Dicer, and Ago2 showed no significant alteration in ER-stressed cells, which suggested that the change in microRNA expression might not be caused by the microRNA biogenesis pathway but by other, unknown factors. Real-time PCR assays confirmed that hsa-miR-423-5p was up-regulated, whereas hsa-miR-221-3p and hsa-miR-452-5p were down-regulated, in both HeLa and HEK293 cells under ER stress. Luciferase activity and Western blot assays verified that CDKN1A was a direct target of hsa-miR-423-5p and that CDKN1B was a direct target of hsa-miR-221-3p and hsamiR-452-5p. We speculated that by regulating their targets, microRNAs might function cooperatively as regulators in the adaptive response to ER stress.