全反式维甲酸通过抑制ERK和激活p38 MAPK信号诱导KLF4基因表达

全反式维甲酸（all-trans retinoic acid, ATRA）诱导细胞分化与上调转录因子Krüppel样因子4 (KLF4)表达有关, 但目前对ATRA诱导KLF4表达的分子机制尚不清楚．为了研究ATRA在血管平滑肌细胞（VSMC）中诱导KLF4表达的分子机制,本 研究观察ATRA对视黄酸受体α (retinoic acid receptor α, RARα)和KLF4表达的影响及RARα介导ATRA诱导KLF4表达所依 赖的信号转导途径．实验结果显示,ATRA可显著诱导RARα和KLF4表达,用RARα拮抗剂Ro 41 5253阻断ATRA与受体相互作 用后,ATRA诱导的KLF4表达受到显著抑制．用p38 MAPK、ERK和Akt抑制剂阻断ATRA与RARα相互作用所激活的信号转导途径 后,发现阻断p38 MAPK信号途径显著抑制ATRA诱导的KLF4表达,抑制ERK信号途径使ATRA对KLF4表达的诱导作用明显增强, 抑制Akt信号途径不影响KLF4基因表达．表明RARα介导ATRA对KLF4表达的诱导作用,ATRA通过抑制ERK和激活p38 MAPK信号 途径发挥其对KLF4基因表达的诱导作用．     

Inhibition of invasion by N-trans-feruloyloctopamine via AKT,p38MAPK and EMT related signals in hepatocellular carcinoma cells

N-trans-feruloyloctopamine (FO) isolated from Garlic skin was identified as the primary antioxidant constituents, however, the effect of which on HCC invasion is still unclear. Herein, the FO was synthesized and its antitumor activities were evaluated in HCC cell lines. Cellular functional analyses have revealed that the reformed FO owns strong abilities of inhibiting cell proliferation and invasion in HCC cells. Molecular data have further showed that FO could significantly decrease the phosphorylation levels of Akt and p38 MAPK. In addition, the expression of Slug was inhibited and the level of E-cadherin increased. Molecular docking analysis indicates that the H-bond and hydrophobic interactions were critical for FO and E-cadherin binding, but FO did not seem to act directly on phosphorylated Akt and p38 MAPK. We have thus concluded that reformed FO inhibits cell invasion might be directly through EMT related signals (E-cadherin) and indirectly through PI3K/Akt, p38 MAPK signaling pathways. FO might be a promising drug in HCC treatment and prognosis.     

脑外伤后ERK信号通路调节AQP4及其锚定蛋白DG的表达

脑外伤是青年人最主要的致死与致残疾病。脑水肿是脑外伤的严重并发症,其形成与脑内最主要的水通道蛋白4（aquaporin4, AQP4）关系密切。AQP4对水的转运与其在星形胶质细胞胞膜上的极性分布有关。肌营养不良-肌萎缩蛋白复合物（dystrophin-dystroglycan complex, DDC）可能与AQP4的锚定及极性分布有关。肌萎缩蛋白（dystroglycan, DG）是该复合物的核心成员,但其对AQP4锚定及极性表达的作用目前并不清楚。脑外伤后,AQP4的表达改变是否与DG有关,其二者表达变化的调控机制均不清楚。为了揭示以上科学问题,为临床治疗脑外伤后脑水肿提供理论依据,分别进行在体、离体及离体干扰实验。研究发现脑外伤后,AQP4、α-DG、β-DG的表达,于6 h增至峰值,后逐渐减弱,于24 h降至最低,48 h再次表达上调。在此过程中,其表达变化规律虽基本一致,但确实存在不一致的现象。排除其他因素干扰,在星形胶质细胞划伤后,DG与AQP4及p-ERK的表达改变完全一致;抑制及激活ERK信号通路后,分别导致DG与AQP4的表达下调及上调。以上结果证实,脑外伤后,DG参与AQP4在星形胶质细胞的锚定,但并非AQP4极性表达的专属锚定蛋白质;机械损伤后,早期ERK信号通路激活,并上调DG及AQP4的表达。     

Involvement of p38 MAPK in haemozoin‐dependent MMP‐9 enhancement in human monocytes

The lipid moiety of natural haemozoin (nHZ, malarial pigment) was previously shown to enhance expression and release of human monocyte matrix metalloproteinase‐9 (MMP‐9), and a major role for 15‐(S,R)‐hydroxy‐6,8,11,13‐eicosatetraenoic acid (15‐HETE), a nHZ lipoperoxidation product, was proposed. Here, the underlying mechanisms were investigated, focusing on the involvement of mitogen‐activated protein kinases (MAPKs). Results showed that nHZ promoted either early or late p38 MAPK phosphorylation; however, nHZ did not modify basal phosphorylation/expression ratios of extracellular signal‐regulated kinase‐1/2 and c‐jun N‐terminal kinase‐1/2. 15‐HETE mimicked nHZ effects on p38 MAPK, whereas lipid‐free synthetic (s)HZ and delipidized (d)HZ did not. Consistently, both nHZ and 15‐HETE also promoted phosphorylation of MAPK‐activated protein kinase‐2, a known p38 MAPK substrate; such an effect was abolished by SB203580, a synthetic p38 MAPK inhibitor. SB203580 also abrogated nHZ‐dependent and 15‐HETE‐dependent enhancement of MMP‐9 mRNA and protein (latent and activated forms) levels in cell lysates and supernatants. Collectively, these data suggest that in human monocytes, nHZ and 15‐HETE upregulate MMP‐9 expression and secretion through activation of p38 MAPK pathway. The present work provides new evidence on mechanisms underlying MMP‐9 deregulation in malaria, which might be helpful to design new specific drugs for adjuvant therapy in complicated malaria. Copyright © 2013 John Wiley & Sons, Ltd.     

Diallyl sulfide ameliorates carbon tetrachloride‐induced hepatotoxicity in rats via suppressing stress‐activated MAPK signaling pathways

The underlined effects of diallyl sulfide (DAS) against CCL₄‐induced oxidative, inflammatory, and apoptotic acute hepatic damage were assessed. Administration of DAS (50, 100, and 200 mg/kg) along with CCL ₄ effectively mitigated serum aspartate aminotransferase, alanine aminotransferase activities, MDA, TNF‐α, IL‐1β, and MCP‐1 levels, as well as significantly restored HO‐1, GSH levels and SOD activity in liver tissues compared with those in rats treated with CCL ₄. Moreover, DAS inhibited CCL ₄‐induced increase of liver NF‐κB (p65), Bax, p38 MAPK, and JNK protein expression. In addition, DAS accelerated protein expression of Nrf2 and Bcl‐2. The hepatoprotective properties of DAS were further confirmed by the reduced severity of hepatic damage as demonstrated by histopathological findings. In conclusion, DAS achieved its protective potential against CCL4‐induced hepatotoxicity through antiapoptotic activity, as well as the synchronized modulation of NF‐κB and Nrf2 for the favor of antioxidant/anti‐inflammatory effects via suppression of the upstream stress‐activated MAPKs pathways.     

Inhibition of P38 MAPK Reduces Tumor Conditioned Medium-Induced Angiogenesis in Co-Cultured Human Umbilical Vein Endothelial Cells and Fibroblasts

Tumor conditioned medium (CM) has been widely used to stimulate endothelial cells to form capillary-like structures in in vitro angiogenesis models. We report herein the effect of HT1080 and A549 CM after they were mixed with microvascular endothelial cells medium-2 (EGM-2) on angiogenesis in human umbilical vein endothelial cells (HUVECs). Both HT1080 and A549 CM decreased HUVEC proliferation, to different extents. While A549 CM significantly increased capillary-like structure formation in a co-culture system, no effect of HT1080 was apparent. Inhibition of p38 mitogen-activated protein kinase (MAPK) blocked both basal and A549 CM induced capillary-like structure formation, but inhibition of extracellular signal-regulated kinases (ERK) and that of c-Jun N-terminal protein kinases (JNK) MAPK had no such effect. Activation of ERK MAPK was inhibited by both CMs, whereas p38 MAPK was inactivated by HT1080 and activated by A549 CM and a control. Neither CM had an effect on JNK MAPK. The results suggest that p38 MAPK played a critical role in capillary-like structure formation in the co-culture, partly via promotion of apoptosis in HUVECs.     

Lipopolysaccharide inhibits GPR120 expression in macrophages via Toll‐like receptor 4 and p38 MAPK activation

Free fatty acid receptor G protein‐coupled receptor 120 (GPR120) is highly expressed in macrophages and was reported to inhibit lipopolysaccharide (LPS)‐stimulated cytokine expression. Under inflammation, macrophages exhibit striking functional changes, but changes in GPR120 expression and signaling are not known. In this study, the effects of LPS treatment on macrophage GPR120 expression and activation were investigated. The results showed that LPS inhibited GPR120 expression in mouse macrophage cell line Ana‐1 cells. Moreover, LPS treatment inhibited GPR120 expression in mouse alveolar macrophages both in vitro and in vivo. The inhibitory effect of LPS on GPR120 expression was blocked by Toll‐like receptor 4 (TLR4) inhibitor TAK242 and p38 mitogen‐activated protein kinase inhibitor LY222820, but not by ERK1/2 inhibitor U0126 and c‐Jun N‐terminal kinase inhibitor SP600125. LPS‐induced inhibition of GPR120 expression was not attenuated by GPR120 agonists TUG891 and GW9508. TUG891 inhibited the phagocytosis of alveolar macrophages, and LPS treatment counteracted the effects of TUG891 on phagocytosis. These results indicate that pretreatment with LPS inhibits GPR120 expression and activation in macrophages. It is suggested that LPS‐induced inhibition of GPR120 expression is a reaction enhancing the LPS‐induced pro‐inflammatory response of macrophages.     

p38 MAPK在小鼠睾丸不同发育阶段的表达和定位

为探讨丝裂原活化蛋白激酶p38 MAPK在小鼠睾丸不同发育阶段的表达,应用蛋白质免疫印迹杂交技术和免疫组织化学SABC法检测1至7周龄小鼠睾丸p38 MAPK的表达、定位及发育变化,并通过图像分析技术对免疫组织化学结果进行统计学分析。免疫印迹杂交发现,p38 MAPK在2～7周龄小鼠睾丸中均有表达。免疫组织化学结果显示,在2周龄小鼠睾丸曲细精管上皮中即可观察到p38 MAPK免疫阳性反应,免疫反应阳性细胞为精原细胞;3、4、5周龄小鼠睾丸仅有个别曲细精管上皮可见p38 MAPK免疫阳性反应;6、7周龄小鼠睾丸中p38 MAPK表达较丰富,免疫反应阳性细胞为精原细胞和初级精母细胞,免疫阳性反应物均主要位于细胞核内。在7周龄小鼠睾丸中还可见到部分间质细胞的细胞质亦呈p38 MAPK阳性。这些结果提示,p38 MAPK可能对生精细胞的增殖分化具有调控作用。     

牦牛AQP4基因CDS序列的克隆及其生物信息学特征分析

为了研究高原动物对青藏高原高寒、低氧等极端生境的适应机理,进一步探讨高原动物对高原反应——高原脑水肿抗性的分子机理,运用基因克隆与生物信息学相关技术和方法,对牦牛脑AQP4(水通道蛋白4,AQP4)基因CDS全长序列进行克隆、基因序列比对及其生物信息学特征分析。结果表明,牦牛AQP4的CDS含有一个966 bp的开放阅读框,编码322个氨基酸;牦牛AQP4基因编码蛋白分子量34.69 k D,理论等电点(p I)7.59,其编码蛋白含有6次跨膜结构,属于疏水性蛋白;二级结构主要由α-螺旋、延伸及无规则卷曲构成;AQP4基因编码产物氨基酸同源性及系统进化分析发现,牦牛AQP4基因编码氨基酸序列与黄牛、绵羊等物种间同源性较高,系统进化情况与其亲缘关系远近一致。     

The binding of actin to p38 MAPK and inhibiting its kinase activity in vitro

Ineukaryocyte,themitogen-activatedproteinkinases(MAPKs)playcriticalrolesinmanysignaltransductionprocesses[1].p38signalpathwayisanimportantbranchoftheMAPKs[2].Oneoftheprimaryfunctionsofp38medicatestheinflammatorysignalbyphosphorylatingATF[3].Itisknownthatinhibitingthep38activitycanblockthesignaltransductionofinflammationandsub-sequentlyalleviateinflammatoryresponse[4].Inrecentyears,severalresearchgroupshavetriedtousesomespecificinhibitorsofp38forclinictrial[5—7].IthasbeendemonstratedthatP…     

Platycodin D induces anoikis and caspase‐mediated apoptosis via p38 MAPK in AGS human gastric cancer cells

Mitogen‐activated protein kinases (MAPKs) cascades play important roles in cell proliferation, death, and differentiation in response to external stimuli. However, the precise role of MAPKs in platycodin D (PD)‐induced cytotoxicity remains unclear. In this study, we investigated the anticancer effect of PD and its underlying mechanism on AGS human gastric cancer cells. PD significantly inhibited cell proliferation and induced anoikis, which is a form of apoptosis in which cells detach from the substrate. It showed phosphatidylserine externalization, DNA fragmentation, increase of sub‐G1 phase, and activation of caspases in a dose‐ and time‐dependent manner. This apoptosis has been associated with the extrinsic pathway via Fas‐L and the intrinsic pathway via mitochondrial Bcl‐2 family members. Moreover, PD led to the phosphorylation of stresses‐activated protein kinases such as JNK and p38, followed by the activation of AP‐1. However, pretreatment with SB203580 (a p38 specific inhibitor) suppressed PD‐induced p38 and AP‐1 activation, and subsequently attenuated the PD‐induced apoptosis in AGS cells. These results suggest that p38 activation is responsible for PD‐induced apoptosis in AGS cells and PD might be useful for the development as the anticancer agent of gastric cancer. J. Cell. Biochem. 114: 456–470, 2013. © 2012 Wiley Periodicals, Inc.     

Hydrogen sulfide alleviates uranium‐induced rat hepatocyte cytotoxicity via inhibiting Nox4/ROS/p38 MAPK pathway

As a gasotransmitter, hydrogen sulfide (H₂S) plays a crucial role in regulating the signaling pathway mediated by oxidative stress. The purpose of this study was to investigate the protective effects of H ₂S on uranium‐induced rat hepatocyte cytotoxicity. Primary hepatocytes were isolated and cultured from Sprague Dawley rat liver tissues. After pretreating with sodium hydrosulfide (an H ₂S donor) for 1 hour (or GKT‐136901 for 30 minutes), hepatocytes were treated by uranyl acetate for 24 hours. Cell viability, reactive oxygen species (ROS), malondialdehyde (MDA), NADPH oxidase 4 (Nox4), and p38 mitogen‐activated protein kinase (p38 MAPK) phosphorylation were respectively determined. The effects of direct inhibition of Nox4 expression by GKT‐136901 (a Nox4 inhibitor) on ROS and phospho‐p38 MAPK levels were examined in uranium‐treated hepatocytes. The results implicate that H ₂S can afford protection of rat hepatocytes against uranium‐induced adverse effects through attenuating oxidative stress via prohibiting Nox4/ROS/p38 MAPK signaling.     

抑制p38MAPK信号通路对Bpiv3复制的影响

由牛副流感病毒3型(Bovine parainfluenza virus type 3,Bpiv3)感染引起的牛副流感病已成为各国牛场最重要的传染病之一,每年都会给世界养牛业造成巨大的经济损失,但关于该病致病的分子机制研究较少。本研究通过观察Bpiv3感染对MDBK细胞中丝裂原活化蛋白激酶(MKK3)及其下游分子p38丝裂酶原活化的蛋白激酶(p38MAPK)的表达的影响,探讨相关的信号转导机制,对p38 MAPK通路在Bpiv3感染过程中的作用进行了初步研究。Bpiv3感染细胞后,采用Western Blot检测MKK3,p38 MAPK在蛋白水平的表达变化,并采用ELISA法检测细胞上清中IL-6,IL-8,IL-13和TNF-α的水平变化,采用SPSS 12软件进行统计学分析。结果表明,Bpiv3在感染后能够诱导MKK3的激活以及p38的磷酸化,激活了p38 MAPK信号通路。而且p38 MAPK信号通路参与了Bpiv3的复制过程。ELISA检测Bpiv3感染后以及使用抑制剂SB202190处理后的细胞上清中IL-6、IL-8、IL-13和TNF-α的水平发现,p38 MAPK信号通路参与了Bpiv3诱导的炎症反应。研究证实Bpiv3感染能够激活p38 MAPK通路,显著上调MKK3的表达并诱导p38发生磷酸化,进一步激活下游分子发挥生物学活性,促进Bpiv3的复制及诱导促炎细胞因子的产生。p38 MAPK信号通路的激活可能是Bpiv3感染诱发炎症反应的机制之一。