水通道蛋白4在大鼠坐骨神经损伤导致的神经病理性疼痛中的作用及机制

目的 :对坐骨神经损伤大鼠的脊髓后角神经元中水通道蛋白4(AQP4)和细胞外调节蛋白激酶(ERK)信号通路的激活之间的关联性做系统的探讨。方法 :制作大鼠坐骨神经慢性压迫性损伤(CCI)模型,经腹腔给予AQP4抑制剂TGN-020,应用DMSO组作对照,应用Von Frey纤毛进行机械痛检测,以免疫荧光(双重染色)检测脊髓ERK、NeuN的表达。结果 :脊髓后角p-ERK和NeuN共定位的细胞在神经损伤后可见增加,而给予TGN-020后,同时表达p-ERK和NeuN的细胞明显减少。TGN-020抑制AQP4的表达可以减轻神经损伤产生的痛觉敏感性异常。结论 :抑制AQP4可以通过抑制脊髓后角神经元中ERK通路的活化来改善坐骨神经损伤导致的神经病理性疼痛。     

抑制AQP4降低大鼠脊神经节ERK表达,减轻坐骨神经结扎导致神经病理性疼痛

目的探讨AQP4抑制剂对坐骨神经结扎导致的神经病理性疼痛的作用及其可能机制。方法制作大鼠坐骨神经慢性压迫性损伤(CCI)模型,采用热痛刺激仪测量热痛感受性潜伏期,Western blot和免疫荧光(双重染色)方法检测ERK, JNK, p38表达。结果神经损伤可诱导ERK, JNK, p38信号分子表达及卫星胶质细胞活化,AQP4抑制剂TGN-020则削弱ERK,JNK和p38信号分子及卫星胶质细胞的活化;p-ERK和GFAP共表达的细胞在损伤后明显增多,TGN-020则显著降低这一表达。结论抑制AQP4减轻坐骨神经结扎导致的神经病理性疼痛与抑制神经节卫星胶质细胞活化和MAPK信号通路活化相关。     

神经病理性疼痛中星形胶质细胞被激活后的p38丝裂原活化蛋白激酶信号转导通路

目的:探索神经性病理痛中星形胶质细胞被激活后的p38丝裂原活化蛋白激酶(p38MAPK)信号转导通路.方法:SD大鼠分为坐骨神经慢性结扎模型组(CCI组)和假手术组(Sham组),并于术前ld和术后1、3、7、14d取第4～5腰段脊髓做石蜡切片,免疫荧光组织化学标记p38MAPK的表达,免疫荧光双标技术检测其与脊髓神经细胞之间的关系.结果:CCI组术后术侧脊髓背角p38MAPK免疫阳性细胞数量增多;p38MAPK平均荧光强度明显增高并在术后第7天显示为最高.p38MAPK和小胶质细胞在CCI组脊髓背角术侧的分布有较好的一致性.结论:在神经病理性疼痛巾,p38MAPK信号转导通路被激活但未参与星形胶质细胞的痛觉信号转导.     

细胞DNA损伤应激反应中丝裂原活化蛋白激酶的信号通路

丝裂原活化蛋白激酶 (MAPKs)途径是细胞遗传毒性应激反应中主要的信号转导途径之一 ,它主要包括ERK ,JNK/SAPK ,p38等通路。各条通路都通过特异的MAPK信号级联放大反应使细胞形成应对DNA损伤的应激反应 ,从而保证细胞的正常生长和DNA复制的保真度。综述DNA损伤应激反应中的各条MAPK信号通路的激活机制     

c-Jun氨基末端激酶与caspase在细胞凋亡中的作用及相互关系

丝裂原活化蛋白激酶(mitogen-activated protein kinase,MAPK)是信号从细胞表面转导到细胞核内部的重要传递者.在真核细胞中,已确定出4条MAPK信号转导通路[1],即细胞外调节蛋白激酶(extracellular regulated protein kinases,ERK) 通路、c-Jun 氨基末端激酶(c-Jun N-terminal kinase,JNK)通路、P38通路及ERK5通路.JNK也称为应激蛋白激酶,参与应激反应和细胞死亡[2].     

c-Jun氨基末端激酶在脑缺血再灌注损伤中的作用

在生物体内,丝裂原活化蛋白激酶(mitogen-activated protein kinase,MAPK)是信号从细胞表面转导至细胞核内部的重要传递者,是一类丝/苏氨酸残基的蛋白激酶。目前在真核生物细胞中,已明确了4条MAPK信号转导通路,即细胞外信号调节蛋白激酶(extracellular signal-regulated protein kinases,ERK)通路、c-Jun氨基末端激酶(c-Jun N-terminal kinase,JNK)通路、p38通路及ERK5通路。JNK信号通路作为MAPK信号通路中重要的通路之一,参与了多种生理、病理过程,目前诸多研究表明其在脑缺血/再灌注损伤过程中尤其是程序性细胞死亡过程中起着重要的调控作用,本文就近年来对JNK通路的研究及JNK在脑缺血再灌注诱导的细胞凋亡中的作用及机制作一综述。     

缺血——再灌注损伤与丝裂素活化蛋白激酶信号传导

信号传导是细胞外剌激信号在细胞膜及细胞内的传递过程。丝裂素活化蛋白激酶(MitogenactivatedProteinkinase,MAPK)是一族胞浆内广泛分布具有丝氨酸双重磷酸化能力的蛋白激酶。凡是兴奋G蛋白偶联受体的物质等损伤因素剌激细胞时,都要通过MAPK激活。因此,MAPK途径是细胞外信号引起细胞反应的共同通路。MAPK家族主要有三个成员,即细胞外信号蛋白激酶(Extracellularsignal-regulatedproteinkinase,ERK),ERK又分离出2个亚型P44ERK/ERK1,P42ERK/ERK2;应激活化蛋白激酶(stress-activatedproteinkinase,SAPK),又称JNK(C-JunN-t…     

MAPK信号通路与神经系统损伤的研究进展

丝裂酶原活化蛋白激酶(Mitogen-activated protein kinase,MAPK)是生物体内重要的信号转导通路之一,主要有ERK、p38和JNK三条途径,参与调控多种细胞应答和生理病理过程。ERK和p38在神经损伤的区域迅速激活,减少神经损伤对机体带来的影响;磷酸化JNK在神经损伤部位聚集,促进神经细胞的凋亡,加重神经损伤发生的过程。MAPK信号通路与神经系统损伤的发生、修复存在着紧密的联系     

多发性骨髓瘤中MAPK信号通路对BLyS表达的调控研究

目的 研究多发性骨髓瘤(MM)细胞中丝裂原活化蛋白激酶(MAPK)信号通路的表达及活化情况,探讨MAPK信号通路对MM细胞B淋巴细胞刺激因子(BLyS)表达变化的影响及对MM细胞增殖与存活的影响,并初步探讨MAPK信号通路在IFN-γ(MM重要的促生长因子)上调MM细胞BLyS表达过程中的作用.方法 应用Western blot方法检测MM细胞中蛋白ERK、p-ERK、JNK、p-JNK、p38及p-p38的表达情况;应用RT-PCR及Western blot检测MAPK信号通路对BLyS表达的影响;应用WST-1法检测靶向JNK的MAPK信号通路抑制剂SP600125对MM细胞增殖与存活的影响.结果 MM细胞株中,除了ERK、JNK及p38的表达外,还有活化蛋白p-JNK的表达;靶向JNK的MAPK信号通路抑制剂SP600125可下调MM细胞BLyS的表达,其激动剂茴香霉素(anisomycin)可上调BLyS的表达;IFN-γ可上调MM细胞BLyS的表达,SP600125可部分抵消IFN-γ对BLyS的上调作用;SP600125可抑制MM细胞的增殖与存活.结论 MM细胞中有JNK/SAPK信号通路的活化;JNK/SAPK信号通路的活化程度与BLyS的表达高低呈正相关;JNK/SAPK信号通路在IFN-γ上调MM细胞BLyS表达过程中发挥重要作用.     

促性腺激素释放激素类似物对人乳腺癌细胞的影响

目的:探讨促性腺激素释放激素(gonadotropin-releasing hormone,GnRH)类似物曲普瑞林(triptorelin)对人乳腺癌细胞株MCF-7、MDA-MB-231细胞生长及细胞外信号调节蛋白激酶(extracellular signal-regulated kinase,ERK)/丝裂原活化蛋白激酶(mitogen-activated protein kinases,MAPK)和磷脂酰肌醇-3-激酶(phosphatidylinositol-3-kinase,PI3K)/蛋白激酶B(protein kinase B,Akt)信号通路中重要信号分子ERK1/2和Akt活化的影响。方法:使用不同浓度、不同时间的曲普瑞林刺激人乳腺癌MCF-7及MDA-MB-231细胞株,MTT法检测细胞增殖,流式细胞术检测细胞周期的分布,Western blotting检测Akt和ERK1/2的磷酸化程度。结果:曲普瑞林(10-5mol/L)作用人乳腺癌MCF-7细胞192 h、曲普瑞林(10-4mol/L)作用人乳腺癌MCF-7细胞168 h、192 h或曲普瑞林(10-4mol/L)作用人乳腺癌MDA-MB-231细胞192 h可明显抑制细胞生长(P0.05);曲普瑞林(10-4mol/L)作用人乳腺癌MCF-7和MDA-MB-231细胞192 h,ERK1/2的磷酸化程度均较正常对照组低,Akt的磷酸化程度较正常对照组高,但差异无统计学意义(P0.05)。结论:GnRH类似物曲普瑞林对人乳腺癌细胞的抑制作用,不仅仅是通过对垂体激素的降调节机制,还可能产生直接抑制作用。但该抑制作用未涉及ERK/MAPK和PI3K/Akt信号通路。     

Multiple kinase pathways mediate the early sciatic ligation-associated activation of CREB in the rat spinal dorsal horn

Phosphorylation of the cyclic AMP response element-binding protein (CREB) in the spinal dorsal horn may critically contribute to chronic pain following peripheral nerve injury. We employed inhibitors and activators of protein kinase A (PKA), protein kinase C (PKC), extracellular signal-regulated kinase 1 and 2 (ERK1/2) and calcium/calmodulin-dependent kinase II (CaMKII) to examine whether these kinases individually or in concert mediate the increase in CREB phosphorylation that is evident as early as 2 h after loose ligation of the sciatic nerve. Specific inhibitors of each kinase significantly attenuated the ligation-associated CREB phosphorylation when compared to saline-treated animals. Combined application of the ERK1/2 and CaMKII inhibitors also attenuated the ligation-associated CREB activation but not to a greater extent than either inhibitor alone. Specific activators of PKA, PKC and ERK1/2 elicited significant increases in CREB phosphorylation 2 h after drug application in the spinal dorsal horn of control, peripherally uninjured animals. Pre-treatment of animals with the ERK1/2 inhibitor abolished the increases elicited by either the PKA or the PKC activator. Significant increases in ERK1/2 phosphorylation were also detected 2 h after sciatic ligation confirming a role for the ERK pathway in injury-related responses in the dorsal horn. Each kinase inhibitor significantly attenuated the ligation-associated activation of ERK1/2 as well. These data suggest that early, sciatic ligation-elicited phosphorylation of CREB in the spinal dorsal horn is mediated by multiple kinase pathways, and that PKA, PKC and CaMKII activate CREB at least in part by way of the ERK pathway.     

Expression of aquaporin 5 increases proliferation and metastasis potential of lung cancer

Water channel aquaporin 5 (AQP5) is highly expressed at the apical membrane of alveolar type I epithelial cells and confers high osmotic water permeability. AQP5 is also expressed in lung cancer tissue. Previous studies showed there was an up‐regulation of AQP5 expression in cancer tissue compared to surrounding normal tissue. In addition, expression of AQP5 in lung cancer tissue was associated with poor prognosis. Herein, we tested the role of AQP5 in lung cancer oncogenesis and development. Lung cancer cells with different expression of AQP5 were used to study cell proliferation and migration, two important parameters for tumour cell biology. We found enhanced proliferation and migration potential in cancer cells with high AQP5 expression, while reduced proliferation and metastasis potential in cancer cells with low AQP5 expression. Oncogene analysis showed significantly increased PCNA and c‐myc expression in AQP5 transfected cells. AQP5 transfected cells also showed significant increased MUC5AC mucin expression, which might contribute to the enhanced metastasis potential of lung cancer. AQP5 overexpression resulted in enhanced activation of the epidermal growth factor receptor (EGFR), extracellular receptor kinase (ERK1/2), and p38 mitogen‐activated protein kinase (p38 MAPK) pathway in cancer cells. Moreover, deletion of AQP5 demonstrated decreased activation of the EGFR/ERK/p38 MAPK pathway in AQP5 knockout mice lungs, while deletion of AQP1 or AQP3 did not exhibit significant changes on activation of the EGFR/ERK/p38 MAPK pathway in lung tissue. In conclusion, our results provide evidence for AQP5‐facilitated lung cancer cell proliferation and migration, possibly through activation of the EGFR/ERK/p38 MAPK signalling pathway, but why AQP5 but not other aquaporin expression affects the EGFR/ERK/p38 MAPK pathway still needs further exploration. Copyright © 2010 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

氧化应激性肠上皮细胞损伤与MAPK信号通路的关系研究

目的观察氧化应激引起肠上皮细胞（IEC-6）损伤后丝裂原活化蛋白激酶（MAPK）信号通路活化的情况,从而探讨氧化应激损伤的机制。方法采用MTT法检测不同浓度（100、200、300、400、500、800、1000、2000μmol/L）H2O2对IEC-6生存率的影响;用WesternBlot法检测H2O2作用不同时间（0.25、0.5、1、2、3、4h）下ERK、JNK以及p38磷酸化的激活情况。结果与正常组相比,随着H2O2刺激浓度的增加,IEC-6的生存率逐渐降低,呈浓度依赖性。当H2O2刺激浓度为200μmol/L时细胞的生存率约为50%。ERK1/2、JNK1/2、p38在H2O2刺激0.25h开始磷酸化,在刺激0.5h达到最高,随后磷酸化水平恢复到基础值。结论氧化应激早期可通过激活ERK、JNK以及p38的磷酸化引起IEC-6的损伤。     

Activation of microglia and p38 mitogen-activated protein kinase in the dorsal column nucleus contributes to tactile allodynia following peripheral nerve injury

The activation of glial cells in the CNS has been suggested to be involved in abnormal pain sensation after peripheral nerve injury. Previous studies demonstrated phosphorylation of p38 mitogen-activated protein kinase (MAPK) in spinal cord glial cells after peripheral nerve injury, and such phosphorylation has been suggested to be involved in the development of neuropathic pain. The aim of this study was to examine the dorsal column nuclei for phosphorylation of p38 MAPK following peripheral nerve injury and to explore a possibility of its contribution to neuropathic pain. Immunohistochemical labeling for phosphorylated p38 (p-p38) MAPK was performed in histological sections of the rat spinal cord and medulla oblongata after the fifth lumbar (L5) spinal nerve ligation (SNL). The number of p-p38 MAPK-immunoreactive (IR) cells was significantly increased in the L5 dorsal horn and the gracile nucleus ipsilateral to the injury at days 3-21 after SNL. Double immunofluorescence labeling with cell-specific markers revealed that p-p38 MAPK-IR cells co-expressed OX-42, suggesting their microglial identity. Increased immunofluorescence labeling for OX-42 indicated that microglial cells were activated by SNL in the L5 dorsal horn and the gracile nucleus ipsilateral to the injury. Continuous infusion of a p38 MAPK inhibitor into the cisterna magna for 14 days beginning on the day of SNL suppressed the development of tactile allodynia, but not thermal hyperalgesia induced by nerve injury. These results demonstrate that SNL activates p38 MAPK pathway in microglia in the gracile nucleus as well as in the spinal cord dorsal horn. Activation of p38 MAPK in medullary microglia may contribute to the pathogenesis of neuropathic pain.     

Activation of JNK pathway in persistent pain

The c-Jun N-terminal kinase (JNK) is a stress-activated member of MAP kinase family. JNK activation has been strongly implicated in inflammatory responses, neurodegeneration, and apoptosis. Recent evidence shows that JNK pathway is also transiently activated in primary sensory neurons after tissue or nerve injury, which is required for the development of hyperalgesia and allodynia. In particular, JNK is persistently activated in astrocytes of the spinal cord after nerve injury, and this activation can maintain central sensitization and mechanical allodynia. In this mini-review, we will provide evidence for the involvement of JNK pathway in regulating persistent pain sensitization. We will also discuss possible upstream signaling mechanisms that cause JNK activation and downstream signaling mechanisms by which JNK modulates pain sensitivity. Thus, targeting JNK pathway might be a useful strategy to treat both neurodegeneration and chronic pain.     

Minocycline prevents impaired glial glutamate uptake in the spinal sensory synapses of neuropathic rats

Activation of glutamate receptors and glial cells in the spinal dorsal horn are two fundamental processes involved in the pathogenesis of various pain conditions, including neuropathic pain induced by injury to the peripheral or central nervous systems. Numerous studies have demonstrated that minocycline treatment attenuates allodynic and hyperalgesic behaviors induced by tissue inflammation or nerve injury. However, the synaptic mechanisms by which minocycline prevents hyperalgesia are not fully understood. We recently reported that deficient glutamate uptake by glial glutamate transporters (GTs) is key for the enhanced activation of N-methyl-d-aspartate (NMDA) receptors in the spinal sensory synapses of rats receiving partial sciatic nerve ligation (pSNL). In this study, we investigated how minocycline affects activation of NMDA receptors in the spinal sensory synapses in rats with pSNL by whole cell recordings of NMDA currents in spinal laminea I and II neurons from spinal slices. The effects of minocycline treatments on the dorsal horn expression of glial GTs and astrocyte marker glial fibrillary acidic protein (GFAP) were analyzed by immunohistochemistry. We demonstrated that normalized activation of NMDA receptors in synapses activated by both weak and strong peripheral input in the spinal dorsal horn is temporally associated with attenuated mechanical allodynia in rats with pSNL receiving intraperitoneal injection of minocycline. Minocycline ameliorated both the downregulation of glial GT expression and the activation of astrocytes induced by pSNL in the spinal dorsal horn. We further revealed that preventing deficient glial glutamate uptake at the synapse is crucial for preserving the normalized activation of NMDA receptors in the spinal sensory synapses in pSNL rats treated with minocycline. Our studies suggest that glial GTs may be a potential target for the development of analgesics.     

Neuropathic pain in aged rats: behavioral responses and astrocytic activation

We used the Bennett and Xie (1988) model of chronic neuropathic pain to study the effect of age on thermal and tactile sensitivity and on astrocytic activation in the dorsal horn of the spinal cord after nerve injury. Fischer 344 FBNF1 hybrid rats in three age groups, 4-6, 14-16, and 24-26 months, were studied. Rats were either unligated (day 0, control) or the left sciatic nerve was loosely ligated to cause a chronic constriction injury (CCI). CCI causes a neuropathic pain condition characterized by tactile allodynia and thermal hyperalgesia. Rats were behaviorally assessed for tactile and thermal sensitivity of their ligated and unligated hind paws up to 35 days postligation. Rats were sacrificed before or at various days postligation, and activated astrocytes were identified at the L4-L5 levels of their spinal cords by use of an antibody to glial fibrillary acid protein (GFAP). The number of GFAP-ir astrocytes in the dorsal horn of the spinal cord in the control, uninjured condition decreased with age (P < or = 0.001) but increased after CCI in all three age groups. After CCI, astrocytic activation in the cord was less robust in aged rats than in younger ones (P < or = 0.01). Not all the CCI rats displayed hyperalgesia to touch and to heat. Rats with an increased sensitivity to heat had increased levels of GFAP-ir in their cords; however, rats with decreased thermal sensitivity also displayed increased GFAP-ir. Thus the presence of activated astrocytes was not correlated with a single behavioral manifestation of neuropathic pain.     

Differential activation of mitogen-activated protein kinases and glial cells in the trigeminal sensory nuclear complex following lingual nerve injury

Mitogen-activated protein kinases (MAPKs) play a pivotal role in the mediation of cellular responses to a variety of signaling molecules. The current study demonstrates phosphorylation of extracellular signal-regulated kinase (ERK) and p38 MAPK in each subdivision of the trigeminal sensory nuclear complex (TSNC) following lingual nerve injury. Immunohistochemical labeling for phosphorylated ERK (p-ERK) or phosphorylated p38 (p-p38) MAPK was performed in histological sections of the brainstem. A transient increase in the immunoreactivity for p-ERK was found in each subdivision of the TSNC followed by a prolonged increase in the immunoreactivity for p-p38 MAPK after nerve injury. Double immunofluorescence labeling with cell-specific markers revealed that ERK and p38 MAPK were phosphorylated predominantly by OX-42-positive microglia or GFAP-positive astrocytes. Increased immunofluorescence labeling for OX-42 and GFAP indicated that microglia and astrocytes were activated by nerve injury in the TSNC. Activation of MAPKs and glial cells in the rostral subdivisions of the TSNC was comparable with that in the subnucleus caudalis of the trigeminal spinal tract nucleus (Vc). We conclude that differential activation of MAPKs and glial cells in the rostral subdivisions of the TSNC as well as the Vc may have a substantial role in the pathogenesis of neuropathic pain following trigeminal nerve injury.     

Ethanol-induced oxidative stress is mediated by p38 MAPK pathway in mouse hippocampal cells

It has been known that ethanol causes neuronal cell death through oxidative stress. Ethanol itself and reactive oxygen species (ROS) produced by ethanol modulate intracellular signaling pathways including mitogen-activated protein kinase (MAPK) cascades. This study was conducted to examine the impact of ethanol on MAPK signaling in HT22 cells. Ethanol (100 and 400 mM) caused activation of ERK, p38 MAPK, and JNK. ERK activation occurred in early time and p38 MAPK activation was evident when ERK activation was diminished. Specific inhibitor of p38 MAPK (SB203580) protected HT22 cells against ethanol, which was accompanied by an inhibition of ROS accumulation. However, inhibitors of ERK (U0126) and JNK (SP600125) had no effects on ethanol-induced neuronal cell death when they are treated with ethanol for 24 h. These results suggest that p38 MAPK may have important roles in ROS accumulation during ethanol-induced oxidative stress in HT22 cells.