加味丹参饮抑制p38MAPK表达保护缺氧/复氧乳鼠心肌细胞损伤的实验研究

目的 观察加味丹参饮含药血清对缺氧/复氧(hypoxia/reoxygenation,H/R)乳鼠心肌细胞p38 MAPK蛋白表达的影响, 以探讨其保护心肌的作用机制. 方法 将20只SD乳鼠的心肌细胞进行原代培养, 建立H/R模型并随机分为H/R组、SB203580 (p38MAPK阻断剂组)、加味丹参饮含药血清组,正常心肌细胞组作为对照组. 采用T淋巴细胞化学染色法鉴定心肌细胞,罗丹明B(Rhodamine B)染色法观察细胞形态,MTT 比色法检测含药血清对乳鼠心肌细胞的毒性,Western-blot 法检测MKK3 (促分裂原活化蛋白激酶-激酶3)、MKK6 (促分裂原活化蛋白激酶-激酶6)、p38MAPK、phospho-p38MAPK蛋白表达. 结果 与正常血清对照组比较,H/R组MKK3、MKK6的蛋白表达增多,p38MAPK通路阻断剂SB203580和加味丹参饮均不能减少其表达;p38MAPK、phospho-p38MAPK在H/R 时表达均增加(P<0.01),而SB203580 和加味丹参饮含药血清均能减少其表达(P<0.01). 结论 加味丹参饮含药血清预处理可通过抑制缺氧,复氧心肌细胞p38MAPK信号通路发挥保护作用,抑制p38MAPK表达及其磷酸化,减轻心肌细胞损伤,起到保护心肌细胞的作用.     

反义p38α基因转染对缺氧复合烧伤血清处理心肌细胞炎性因子的表达

目的探讨反义p38α基因转染对缺氧复合烧伤血清处理心肌细胞炎性因子的表达.方法建立缺氧复合烧伤血清培养心肌细胞模型,分对照组、缺氧复合烧伤血清组(非转染组)和反义p38α基因转染组(转染组).烧伤血清采自40%TBSA Ⅲ度烧伤Wistar大鼠;采用含1%O2的混合气体造成缺氧模型;采用基因重组技术构建反义p38α基因重组体;用免疫印迹法和RT-PCR法分别检测不同时相点p38α激酶蛋白及TNFα、IL-1β mRNA表达变化,并作统计学分析.结果成功构建反义p38α基因重组体;缺氧复合烧伤血清迅速、持续激活心肌细胞p38α激酶,心肌细胞TNFα、IL-1β表达增多;反义p38α基因显著抑制p38α激酶过度活化(P＜0.01),下调 TNFα、IL-1β表达(P＜0.01).结论 p38α激酶途径介导了缺氧和烧伤血清所致心肌细胞损伤,抑制p38α激酶的持续活化能有效下调炎性因子TNFα、IL-1β表达,减轻该条件下心肌细胞损害.     

钙敏感受体参与MAPK通路诱导细胞凋亡信号传导通路的机制研究

目的钙敏感受体(calcium-sensing recetor,CaR)具有七个跨膜区的G蛋白耦联受体,它通过激活PLC系统引起肌浆网释放钙从而引起细胞内钙的升高。CaR与心肌细胞凋亡的关系及相关信号传导途径。方法大鼠乳鼠心肌细胞培养复制缺血再灌注损伤,并分为五组,分别:1)对照组;2)模拟缺血/再灌注组;3)GdCl3 NiCl2 CdCl2组;4)SP600125组;5)SB203580组;观察加入CaR激动剂时CaR的表达情况;MTT法和流式细胞仪观察细胞凋亡;Western Blot检测心肌组织,同时还检测了培养心肌细胞中磷酸化JNK和p38的表达。结果心肌缺血/再灌注和加入CaR激动剂时CaR的表达明显高于对照组,细胞内钙升高幅度明显大于对照组。HE染色、TUNEL和电镜观察发现缺血/再灌注和激动剂组损伤明显,并且凋亡细胞广泛存在。同时,细胞浆中cyt c和Caspase3的表达明显增加,而线粒体中的cyt c和胞浆中的Bcl2的表达下降。磷酸化JNK和p38的表达增多,在加入JNK抑制剂SP600125和p38抑制剂SB203580心肌细胞凋亡率明显下降,磷酸化JNK和p38的表达减少。结论CaR参与大鼠心肌缺血/再灌注损伤介导细胞内钙超载。CaR激活后,通过诱发线粒体损伤而引起cytc-Caspase 3途径促进心肌细胞凋亡;同时亦可激活JNK和p38酪氨酸蛋白激酶途径参与细胞凋亡。     

机械牵张对烧伤血清复合缺氧培养大鼠心肌细胞p38、JNK、c-jun、ATF-2表达的影响

目的　构建严重烧伤后合并心肌负荷增加的体外心肌细胞模型 ,探讨p3 8、JNK、c jun、ATF 2的活化情况 ,明确高机械牵张刺激对严重烧伤后心肌细胞损伤信号的传导。方法　采用原代培养的乳鼠心肌细胞 ,分别给予正常血清(SN组 )、烧伤血清 缺氧 (1%O2 ) (SH组 )、烧伤血清 缺氧 (1%O2 ) 10 %轴向静态牵张 (SHS组 ) 3种处理 ,采用Westernbloting检测磷酸化p3 8、JNK、c jun、ATF 2水平。结果　施加缺氧复合烧伤血清刺激后 ,磷酸化p3 8、c jun、ATF 2水平升高。在施加高机械牵张后p3 8升高更加明显 ,而后两者与前者表现相反。JNK自始至终没有出现活化。结论　严重烧伤后 ,p3 8可能是介导心肌细胞损伤的重要信号传导分子 ,而JNK并未参与严重烧伤后心肌细胞损伤的信号传导 ,转录因子c jun、ATF2亦可能参与严重烧伤后心肌细胞的损伤 ,但在严重烧伤合并心脏负荷增加的情况下可能并不是主要的调控因子。     

丝裂原活化蛋白激酶信号转导通路在金黄色葡萄球菌α-毒素所致人外周血单核细胞凋亡中的作用

目的研究丝裂原活化蛋白激酶信号转导通路在金黄色葡萄球菌α-毒素所致人外周血单核细胞凋亡中的作用。方法以Annexin V异硫氰酸荧光素/碘化丙啶双染流式细胞仪检测人外周血单核细胞的凋亡率,以Western blot法检测p38丝裂原活化蛋白激酶(MAPK)、细胞外信号调节激酶(ERK)及c-Jun N末端激酶(JNK)等蛋白的表达。结果随着α-毒素作用时间的延长,磷酸化-p38 MAPK和JNK1/2等蛋白的表达逐渐增高,而磷酸化-ERK1/2蛋白的表达不变。用SB203580和SP600125阻断p38 MAPK和JNK1/2后,单核细胞凋亡率明显降低,分别为(17.7±1.8)%和(15.3±1.6)%,与感染60 min时(35.7±3.6)%比较,差异有统计学意义(P<0.05)。结论 MAPK信号通路的成员p38 MAPK和JNK1/2在金黄色葡萄球菌的致病过程中起重要作用。     

p38 MAPK抑制剂通过抑制JNK活性抑制培养的小脑颗粒神经元凋亡

【目的】研究特异性p38分裂原激活的蛋白激酶（MAPK）抑制剂SB203580对低钾诱导的小脑颗粒神经元凋亡的作用。【方法】把体外培养的小脑颗粒神经元从含去极化浓度钾离子（KCl25mmol*L-1）的培养基中转移至低钾培养基（KCl5mmol*L-1）中诱导神经元凋亡。凝胶电泳分析DNA片段,SAPK/JNK分析盒测定c-JunN-末端蛋白激酶(JNK)活性。【结果】低钾诱导小脑颗粒神经元的具有典型形态学和生化特征的凋亡。特异性的p38MAPK抑制剂SB203580通过抑制细胞凋亡,促进低钾环境中培养的小脑颗粒神经元存活。这种保护作用具有浓度依赖性。培养于低钾环境中的颗粒神经元,c-Jun表达和磷酸化水平升高了,且激活了JNK活性。当小脑颗粒神经元生长在含SB20358025μmol*L-1的低钾培养基中,c-Jun表达、磷酸化水平和JNK活性都明显降低。【结论】SB203580抑制JNK活性,降低c-Jun的磷酸化而对低钾培养的小脑颗粒神经元具有保护作用。     

PI3K/Akt信号途径抑制烧伤后大鼠缺血缺氧心肌细胞凋亡

目的 探讨PDK/Akt信号途径在烧伤后大鼠缺血缺氧心肌细胞凋亡中的作用及机制.方法 首先建立模拟烧伤的动物模型,通过Western blot观察P13K/Akt信号途径的活化规律,TUNEL实验观察烧伤大鼠心肌凋亡.然后建立离体培养的缺血缺氧心肌细胞模型,并分为对照组、单纯缺血缺氧组、LY294002处理组或IGF-1处理组,应用ELISA观察缺血缺氧心肌细胞凋亡.并通过DEVD荧光检测caspase-3酶活性,RT-PCR检测p53、Bax的转录活性.结果 烧伤后大鼠心肌组织内P13K和pAkt表达逐渐增加,伤后3 h达高峰;烧伤3 h后,大鼠心肌细胞凋亡率显著增加;应用LY294002特异性抑制P13K/Akt通路后,大鼠心肌细胞凋亡率较对照组增加更为显著(P<0.05).缺血缺氧导致体外培养心肌细胞凋亡增加,caspase-3活性逐渐增强,p53、Bax mRNA表达量逐渐升高;与单纯缺血缺氧组相比,应用LY294002阻断P13K/Akt途径活化后,心肌细胞凋亡数量增加更显著(P<0.05),caspase-3活性增高更为明显(P<0.05),053、Bax mRNA表达量均显著升高(P<0.05);用IGF-1预先激活P13K/Akt途径后,与单纯缺血缺氧组比较,心肌细胞caspase-3活性明显降低(P<0.05).结论 P13K/Akt信号途径在缺血缺氧心肌细胞中具有抗凋亡作用,该作用与P13K/Akt调控促凋亡基因p53、Bax的表达,抑制caspase-3凋亡反应有关,并为心肌缺血缺氧损害的临床防治提供新思路及实验依据.     

褪黑素通过p38/MAPK信号通路调控oxLDL诱导的人脐静脉内皮细胞MLCK的表达

目的 探讨褪黑素(MLT)通过p38/MAPK信号通路调控oxLDL诱导的人脐静脉内皮细胞肌球蛋白轻链激酶(MLCK)的表达.方法 体外培养人脐静脉内皮细胞(HU-VEC),分空白对照组、DMSO对照组、DMSO oxLDL组、DM-SO oxLDL SB203580组、DMSO oxLDL MLT组、DMSO oxLDL MLT SB203580组.RT-PCR、Western blot、γ-32>P-ATP掺入法分别检测各组HUVEC MLCK转录、表达和活性及MLT对p38磷酸化的影响.结果 RT-PCR显示oxLDL组MLCK转录增强,MLT组与SB203580组均抑制MLCK的转录;Western blot结果 表明oxLDL组MLCK表达、p38磷酸化增强,MLT组与SB203580组均抑制MLCK的表达及p38磷酸化;γ-32P-ATP掺入法检测MLCK活性提示oxLDL组MLCK活性增强,MLT与SB203580降低MLCK活性.结论 MLT可能通过p38/MAPK通路调控MLCK的转录、表达和活性.     

阿霉素可通过MAPK信号转导通路促进成骨肉瘤细胞凋亡

目的: 探讨阿霉素在适宜的加药浓度、作用时间下诱导成骨肉瘤细胞凋亡的分子机制.方法: 首先通过FCM,MTT等方法探讨引起成骨肉瘤细胞凋亡的最佳药物作用浓度及时间;在此条件下应用不同信号转导通路的抑制剂观察导致细胞凋亡的可能通路;最后应用Western-Blotting来检测在成骨肉瘤细胞凋亡过程中相关信号分子的表达水平及磷酸化水平的变化.结果: 0.4 mg/L阿霉素作用成骨肉瘤细胞12 h即可引起细胞凋亡,并呈时间依赖性;用p38的抑制剂SB203580和MEK4/7的抑制剂U0126可明显抑制细胞的凋亡;Western Blotting检测发现阿霉素作用于细胞后,MAPK家族成员JNK、ERK和p38等各激酶的表达水平未发生明显变化,但JNK,p38的磷酸化水平提高了ERK的磷酸化水平有所下降;用Western Blotting检测到阿霉素作用后p53蛋白的表达水平有提高.结论: 阿霉素可导致成骨肉瘤细胞凋亡,而且此凋亡过程与MAPK通路有关.     

Aβ25～35通过激活p38MAPK信号通路促进大鼠心肌细胞凋亡

目的 观察β淀粉样蛋白25～35 (Aβ25～35)对培养大鼠心肌细胞的毒性作用,并阐明可能的机制.方法 体外培养大鼠心肌细胞,给予不同浓度Aβ25～35刺激,采用流式细胞术检查细胞凋亡率,采用Western blot方法测定凋亡相关蛋白Bax和Bcl-2的表达水平.观察Aβ25～35对p38丝裂原活化蛋白激酶(MAPK)磷酸化的影响,并进一步评估p38MAPK特异性抑制剂对Aβ25～35诱导心肌细胞凋亡的影响.结果 Aβ25～35可以诱导体外培养大鼠心肌细胞凋亡,升高促凋亡蛋白Bax的表达,下调抗凋亡蛋白Bcl-2的表达,且呈现浓度依赖的关系.同时发现给予Aβ25～35后p38MAPK磷酸化水平增加,而给予p38MAPK特异性抑制剂SB203580减少心肌细胞的凋亡.结论 Aβ25～35可以导致体外培养的大鼠心肌细胞发生凋亡,而p38MAPK信号通路可能参与了该过程.本研究探索阿尔兹海默病(AD)患者心肌损伤的可能发病机制,为将来有效防治AD相关性心肌损伤提供新思路.     

APPLICATION OF LORNOXICAM TO PATIENT-CONTROLLED ANALGESIA IN PATIENTS UNDERGOING ABDOMINAL SURGERIES

FOR anesthesiologis s ,treatingpostoperativepainhas alwaysbeen a problem.Althoughopioidshave been provedtobe effective,theirsideeffectscouldnotbeignored.With thedevelopmentofscienceand pharmacology,many drugs with aspectsof satisfactoryanalgesicefficacyand couldbe welltoleratedby patientshave been developed.And lornoxicamisone of them, which isa non-steroidalanti-inflammatorydrug (NSAID ), with analgesic, anti-infl-ammatory,andantipyreticproperties.Itseliminationhalf-time(3 to 5 hours) isle…     

Dr.Zhang Ren''''s Experience in the Acupuncture Treatment of Different Diseases with the Same Therapeutic Principle

Dr.Zhang Ren,the chief physician,is the chairman of Shanghai Acupuncture and Moxibustion Association.Having been engaged in medicine for about 40 years,he is experienced in treating various intractable diseases.In his long years of clinical practice,he advocates taking the TCM differentiation as the basis to seek for the acupuncture method for treatment of modern intractable diseases.The author of this essay had the fortune to follow Dr.Zhang in study.The following is a summary of Dr.Zhang's experience in the acupuncture treatment for different intractable diseases with the same therapeutic principle.     

Luo Lingjie''''s Experience in Treating Chronic Nephropathy

In treating chronic nephropathy,Luo Lingjie,a chief physician,pays attention to regulating the balance between yin and yang,treating infection if present,and removing pathogenic factors.He prescribes gentle drugs and uses carefully strongly warming-tonifying ones,emphasizes the importance of persuading the patient to persist in treatment with medication and nurse one's health for recuperation,and is good at combined use of TCM and western medicine therapy and brings the merits of various therapies into full play,with obvious theraoeutic effects.     

Acupuncture Combined with Spinal Tui Na for Treatment of Primary Dysmenorrhea in 30 Cases

Objective: To observe the therapeutic effects in acupunture treatment of primary dysmenorrhea combined with spinal Tui Na, and study its mechanism. Methods: Thirty cases of the treatment group were treated by acupuncture combined with spinal Tui Na, and thirty cases in the control group were treated by routine acupuncture. Results: The total effective rate was 93.3% in the treatment group, and 73.3% in the control group, with a significant difference between the two groups (P<0.05). Conclusions: Acupuncture combined with spinal Tui Na has good prospects for treatment of primary dysmenorrhea.     

猪肺磷脂注射液联合经鼻持续气道正压通气对呼吸衰竭早产儿的临床疗效及肌酸激酶同工酶活性的影响

目的 探讨猪肺磷脂注射液联合经鼻持续气道正压通气(NCPAP)对呼吸衰竭早产儿的临床疗效及肌酸激酶同工酶活性(CK-MB)的影响.方法 选取呼吸衰竭早产儿80例,分为观察组和对照组各40例.对照组采用NCPAP给氧治疗,观察组给予NCPAP给氧联合猪肺磷脂气管内给药.观察两组患儿治疗前及治疗12h、24 h后PaO2、PaCO2、血氧饱和度(SaO2)、pH的变化情况,检测治疗前及治疗5d后血清CK-MB水平;评估两组患儿的临床治疗效果.结果 两组患儿PaO2、PaCO2、SaO2、pH比较,差异均有统计学意义(P＜0.05),其中观察组治疗后的PaO2、SaO2、pH均高于对照组,PaCO2则低于对照组.两组的PaO2、SaO2、pH均随观察时间延长而升高(P＜0.05),PaCO2均随观察时间的延长而降低(P＜0.05).观察组治疗有效率为87.5％,显著高于对照组的70.0％ (P ＜0.05).治疗5d后两组患儿血清CK-MB水平均较前降低(P＜0.05),且观察组明显低于对照组(P＜0.05).结论 猪肺磷脂注射液气管内给药联合NCPAP可以显著降低呼吸衰竭早产儿CK-MB的含量,提高治疗有效率,起到很好的呼吸循环支持作用.     

Survey and practice of reporting quality of randomized controlled clinical trials on traditional Chinese medicine

Evidence obtained from randomized controlled trials （RCTs） has been generally accepted as the gold standard in the evaluation of clinical effectiveness. Readers need to understand the trial design, implementation, results, analysis and interpretation, so as to fully Jnderstand the results of RCTs. Thus, the investigators of RCTs have to report these items in a complete, accurate and clear manner. Since 1998, we have conducted several evaluations on the reporting quality of RCTs published in Chinese journals on traditional Chinese medicine （TCM） and results have shown that there is an urgent need for higher quality RCTs on TCM.     

TCM Treatment of Ankylosing Spondylitis by Tonifying the Kidney and Strengthening the Governor Vessel

Ankylosing spondylitis is a chronic and progressive disorder with inflammation mainly involving the central axis joints. It mainly affects the cervical spine and the lumbosacral area, with the pathogenesis closely related to the kidney and the Governor Vessel (GV). TCM holds that the syndrome is deficiency in origin and excess in superficiality, which is due to insufficiency of the kidney, deficiency of GV, and blocking of the channels with the invasion of exogenous evil, leading to poor circulation of qi and blood and malnutrition of the bones, muscles and joints. The TCM method of tonifying the kidney and strengthening GV to regulate circulation of qi and blood and check the arthralgia pain should be adopted, with the Kidney-Tonifying and GV Strengthening Decoction (益肾强督汤) prescribed.     

IMPROVING P-gp EXPRESSION IN HUMAN MONONUCLEAR CELLS IN VITRO TRANSFECTED BY MULTIDRUG RESISTANCE-1 mRNA

CHEMOTHERAPY playsa greatrolein the treat- ment of malignanttumors,especiallyingynecolo- gicalones.But inanticancerchemotherapy,leuko-cytopeniaisfrequentlytheprimarydose-limitingsideeffect factor.Moreover,cancersarefrequentlychemoresistantbe-causeof overexpressionof P-glycoprotein(P-gp), which isencodedby multidrugresistancegene (MDR1 ) and detectableinup to50% ofhuman cancersand renderscellsresistancetoanticancerdrugs.The safetyand potentialtherapeuticbenefitof mdr1 gene transferredto h…