细胞周期蛋白D1在支气管哮喘小鼠肺组织中的表达及其与气道重塑的关系

目的 研究细胞周期蛋白D1(Cyclin D1)在支气管哮喘(简称哮喘)小鼠肺组织中的表达,探讨Cyclin D1在哮喘及气道重塑中的作用.方法 将40只SPF级BALB/c小鼠按随机数字表法分为正常对照组(A组)、哮喘雾化2周组(B组)、哮喘雾化4周组(C组)、哮喘雾化8周组(D组)4组,每组10只.用10%鸡卵白蛋白(OVA)致敏和1%OVA激发小鼠建立哮喘模型.分析支气管肺泡灌洗液(BALF)中嗜酸粒细胞(EOS)计数及分类;用动物肺功能仪检测各组小鼠肺功能状况;用苏木精-伊红(HE)染色观察气道炎症及细胞浸润情况;用图像分析软件观察气道壁及平滑肌层变化情况;用逆转录-聚合酶链反应(RT-PCR)及实时定量(Real-time)PCR测定肺组织中Cyclin D1 mRNA水平表达变化;用Western blot法观察肺组织中Cyclin D1的蛋白表达变化.结果 BALF分析结果提示,B、C、D组EOS计数分别为(42.6±0.9)×104/L、(54.7±1.4)×104/L、(44.8±2.4)×104/L,与A组[(3.4±0.5)×104/L]比较差异有统计学意义(q值分别为79.75、91.42、84.82,P均＜0.01);对小鼠呼气阻力检测发现,乙酰胆碱浓度为45 μg/kg时B、C、D组分别为(5.27±0.16)cm·L-1·min-1、(6.68士0.20)cm·L-1·min-1、(7.14±0.41)cm·L-1·min-1,与A组[(4.11±0.15)cm·L-1·min-1]比较差异有统计学意义(q值分别为5.58、6.39、7.11,P均＜0.05);支气管平滑肌面积/管腔内周长(Wam/Pi)B组为2.8±0.6,C组为4.8±0.6,D组为6.4±0.7,与A组(2.4±0.4)比较差异有统计学意义(q值分别为6.40、8.28、9.27,P＜0.05);管壁面积/管腔内周长(Wat/Pi)B组为6.4±0.8,C组为8.3±1.2,D组为9.3±1.0,与A组(5.6±1.0)比较差异有统计学意义(q值分别为2.80、4.83、6.37,P均＜0.05);Western blot检测发现Cyclin D1在B、C、D组表达量分别为0.587±0.015、0.808±0.029、0.826±0.022,与A组(0.404±0.016)比较差异有统计学意义(q值分别为5.87、8.08、8.26,P均＜0.01);相关性分析结果提示呼气阻力和Cyclin D1水平表达呈正相关(r=0.83,P＜0.05).结论 Cyclin D1在哮喘小鼠肺组织中表达增加,其表达与气道反应性呈正相关,Cyclin D1可能通过细胞外信号调节激酶(ERK)信号通路参与气道重塑过程.     

Peg10基因在黏着斑激酶介导的肝癌细胞耐药中的作用

目的初步探讨Peg10基因与黏着斑激酶(FAK)介导的肝癌细胞耐药(CAM-DR)的关系及分子机制。方法采用MTT法检测5-氟尿嘧啶(5-Fu)、阿霉素(ADR)对LO2细胞、ADR耐药的人肝癌细胞BEL-7404/ADR(7404/ADR)和Peg10基因沉默的siRNA-BEL-7404/ADR(siRNA-7404/ADR)细胞增殖的影响;建立7404/ADR和siRNA-7404/ADR裸鼠荷瘤模型,48只裸鼠随机分为6组,每组8只,A、C、D组注射7404/ADR细胞,B、E、F组注射siRNA-7404/ADR细胞;A、B组尾静脉注射生理盐水,C、E组给予5-Fu,D、F组给予尾静脉注射ADR,测量瘤块体积,RT-PCR检测Peg10基因的表达,Western Blot法检测PEG10、p-FAK、p-JNK、p-ERK和p38MAPK蛋白的表达。两组间比较采用t检验,多组间比较采用单因素方差分析。结果 ADR和5-Fu对siRNA-7404/ADR 24 h的IC50值均较7404/ADR的降低,差异均具有统计学意义(t值分别为7.641,7.560,P值均<0.01)。B、C、D组肿瘤体积较A组小,差异均有统计学意义(P值均<0.05),E、F组肿瘤体积较B组小,差异均具有统计学意义(P值均<0.01);E组与C组比较,F组与D组的肿瘤体积差异亦均有统计学意义(P值均<0.05)。B、E、F组Peg10的mRNA极少表达,C、D组Peg10的mRNA表达较A组有所降低。B组PEG10蛋白极少表达,与A组比较,差异具有统计学意义(P<0.01),其pFAK、p-JNK、p-ERK和p38MAPK蛋白表达与A组比较,差异均有统计学意义(P值均<0.05)。C、D组PEG10、p-FAK、pJNK、p-ERK和p38MAPK蛋白的表达较A组均降低,差异均有统计学意义(P值均<0.05)。E、F组p-FAK、p-JNK、p-ERK和p38MAPK蛋白表达与B组比较,差异均有统计学意义(P值均<0.01),而C组与E组、D组与F组相比,PEG10、p-FAK、p-JNK、p-ERK和p38MAPK蛋白的表达差异亦均有统计学意义(P值均<0.01)。结论 Peg10基因失活后可增加ADR耐药的BEL-7404细胞株对5-Fu和ADR的敏感性,其作用机制可能与其下调p-FAK、p-JNK、p-ERK和p38MAPK蛋白的表达有关。     

线粒体膜电位对缺氧人肺动脉平滑肌细胞内氧自由基的变化及细胞增殖的作用

目的研究线粒体膜上ATP敏感钾通道(MitoKATP)的开放剂二氮嗪和线粒体膜电位在缺氧引起的人肺动脉平滑肌细胞(HPASMC)内氧自由基的变化及细胞增殖/凋亡失衡中的作用。方法培养HPASMC并将所培养的细胞分为6组正常对照组(A组);MitoKATP阻断剂5-羟基癸酸盐(5-HD)组(B组);MitoKATP开放剂二氮嗪组(C组);慢性缺氧组(D组);慢性缺氧+二氮嗪组(E组);慢性缺氧+5-HD组(F组),每组样本数均为6。利用激光共焦显微镜成像检测线粒体膜电位,荧光染色检测细胞内氧自由基含量,免疫组化法检测增殖细胞核抗原(PCNA)、c-fos及c-jun的蛋白表达和四甲基偶氮唑盐(MTT)法检测细胞增殖情况。结果C、D、E组细胞线粒体膜电位(以R123的荧光强度表示)分别为105±4、95±13、126±8,较A组(75±7)明显去极化(q值分别为5.474、3.659、9.213,P均<0.05);C、D、E组细胞内氧自由基含量分别为3045±126、3116±34、3236±31,与A组(2772±49)比较差异有统计学意义(q值分别为6.882、7.448、16.289,P均<0.05);C、D、E组细胞增殖活性[以MTT法检测出的A值表示]分别为0.305±0.022、0.328±0.078、0.440±0.023,与A组(0.237±0.013)比较差异有统计学意义(q值分别为2.993、4.017、8.919,P均<0.05),且E组线粒体膜电位、细胞内氧自由基含量、细胞增殖活性与D组比较差异有统计学意义(q值分别为5.554、8.841、4.902,P均<0.05)。F组线粒体膜电位、细胞内氧自由基含量、细胞增殖活性分别为71±4、2863±132、0.264±0.045,与D组(95±13、3116±34、0.328±0.078)比较差异有统计学意义(q值分别为4.367、5.907、2.832,P均<0.05)。结论二氮嗪或缺氧能够通过开放HPASMC线粒体膜上ATP敏感的钾通道,引起线粒体膜电位去极化,增加细胞内氧自由基的含量,最终导致HPASMC的增殖/凋亡的失衡,从而促进了缺氧性肺动脉重塑过程。     

实验性支气管哮喘豚鼠肺和内脏感觉传入系统蛋白激酶C的表达及神经生长因子的调节作用

目的探讨支气管哮喘(简称哮喘)豚鼠肺及内脏感觉传入系统(C7～T5脊神经节及对应的脊髓后角)蛋白激酶C(PKC)的表达及神经生长因子(NGF)的调节作用。方法40只豚鼠按随机数字表法分为4组生理盐水对照组(A组)8只、单纯致敏对照组(B组)8只、哮喘组(C组)12只和NGF抗体组(D组)12只。用免疫组织化学方法检测各组豚鼠C7～T5脊神经节和对应的脊髓后角PKC的免疫反应变化;采用Westernblot方法分别检测各组豚鼠肺组织、C7～T5脊神经节和对应的脊髓后角NGF与PKC的表达;采用LuzexF实时图像分析系统和凝胶成像分析系统分别对以上结果进行图像分析。结果(1)免疫组织化学A、B组豚鼠C7～T5脊神经节和对应的脊髓后角PKC的平均吸光度(A)值分别为0.102±0.009、0.113±0.009、0.106±0.005、0.116±0.007,二者比较差异无统计学意义(P>0.05)。C组豚鼠C7～T5脊神经节和对应的脊髓后角PKC的A值分别为0.215±0.014、0.176±0.010,C组与A组比较差异有统计学意义(P<0.01)。D组豚鼠C7～T5脊神经节和对应的脊髓后角PKC的A值分别为0.140±0.008、0.130±0.011,D组与C组比较差异有统计学意义(P<0.01)。(2)Westernblot蛋白印迹C组肺组织、C7～T5脊神经节神经元及相应脊髓节段PKC蛋白表达量(其A相对值分别为1.51±0.01、1.40±0.03、2.22±0.02)与A组(其A相对值分别为0.51±0.02、0.43±0.01、0.92±0.02)和B组比较均明显增加,而D组PKC蛋白表达量(其A相对值分别为0.80±0.03、0.83±0.01、1.12±0.02)明显低于C组。结论PKC可能参与哮喘的发病过程,而NGF可上调哮喘豚鼠PKC的表达。     

树Qu实验性肝癌发生过程p53基因的变化

目的探讨由人乙型肝炎病毒(HBV)和黄曲霉毒素B1(AFB1)诱发的树鼩肝细胞癌变过程,p53基因的表达及变化.方法将树鼩分为四组A组HBV+AFB1,B组只感染HBV;C组只摄入AFB1;D组作空白对照.定期肝活检,用免疫组织化学、分子生物学等技术对实验树鼩肝及肿瘤组织进行检测.结果 (1)接受HBV及AFB1双因素的A组,肝细胞癌(HCC)发生率(66.7%)明显高于只接受HBV的B组或AFB1的C组(30%),而且HCC的平均发生时间也明显早于C组,(120.0±16.6)周与(153.3±5.8)周,t=3.336,P＜0.01.(2)在第75周前各组动物肝均未检出突变的p53蛋白.(3)105周时,A组p53蛋白表达率为78.6%,B组为60%,C组为71.4%,D组为10%(x2≥5.03,P＜0.05).在A、C组检出p53基因异常带.(4)树鼩肝癌p53基因突变点分别位于2 7 5、7 8及1 3密码子;其野生型p 5 3基因的核苷酸及氨基酸序列与人的p 5 3基因的核苷酸及氨基酸序列的同源性分别为91.7%、93.4%.结论再次证实HBV和AFB1有协同致肝癌作用;突变的p53蛋白出现于肝细胞发生癌变之前,p 5 3基因的突变促进了肝癌的发生和演进.HBV可能协同AFB1致p 5 3基因突变.     

全反式维甲酸对人肺动脉平滑肌细胞表型和迁移的影响及可能机制

目的探讨全反式维甲酸(atRA)对培养的人肺动脉平滑肌细胞(PASMC)表型和迁移的影响及可能机制。方法培养的人PASMC株随机分成8组:对照组(A组)、溶媒组(B组)、10%胎牛血清干预组(C组)、atRA干预组分别加入浓度为0.001μmol/L(D组)、0.01μmol/L(E组)、0.1μmol/L(F组)、1μmol/L(G组)、10μmol/L(H)组的atRA,干预24 h后用逆转录-聚合酶链反应(RT-PCR)法检测各组细胞JWA基因mRNA表达,用改良Boyden小室法检测各组迁移细胞数,用RT-PCR法及免疫细胞化学法检测各组细胞内平滑肌细胞α肌动蛋白(SM-α-actin)mRNA及蛋白表达。结果人PASMC内JWA mRNA表达量A、B、C组分别为0.125±0.014、0.164±0.018、0.164±0.006,3组间比较差异无统计学意义(P均>0.05);而D组(0.326±0.018)、E组(0.440±0.033)、F组(0.460±0.040)、G组(0.589±0.024)、H组(0.821±0.050)分别与A、B、C组比较差异均有统计学意义(P均<0.01);迁移细胞数C组为(30.4±7.4)个/高倍视野(HP),与A组[(7.2±1.9)个/HP]、B组(6.8±2.3)个/HP]比较差异有统计学意义(P均<0.01);而D组[(21.8±2.9)个/HP]、E组[(17.2±2.3)个/HP]、F组[(14.4±3.5)个/HP]、G组[(12.6±3.4)个/HP]、H组[(8.8±2.4)个/HP]与C组比较差异有统计学意义(P均<0.01);但G、H组与A、B组比较差异无统计学意义(P均>0.05);细胞内SM-α-actin蛋白表达量C组为0.219±0.018,与A组(0.319±0.011)、B组(0.325±0.005)比较差异均有统计学意义(P均<0.01);E组(0.328±0.016)、F组(0.386±0.025)、G组(0.442±0.017)、H组(0.501±0.018)与C组比较差异均有统计学意义(P均<0.01);而F、G、H组与A、B组比较差异均有统计学意义(P均<0.01);细胞内SM--αactin mRNA表达量C组为0.144±0.009,与A组(0.299±0.023)、B组(0.296±0.041)比较差异有统计学意义(P均<0.01);而D组(0.487±0.014)、E组(0.501±0.020)、F组(0.611±0.018)、G组(0.774±0.013)、H组(0.851±0.026)与A、B、C组比较差异均有统计学意义(P均<0.01)。结论atRA能诱导人PASMCs内JWA基因表达增加,使细胞表现为收缩表型,细胞迁移受到抑制。     

持续吸入变应原引起支气管哮喘小鼠免疫耐受

目的探讨吸入变应原引起支气管哮喘(简称哮喘)过敏性气道炎症免疫耐受形成的机制。方法BALB/c小鼠60只,按随机数字表法分为实验组(50只)和空白对照组(10只),实验组小鼠先给予腹腔注射卵清白蛋白(OVA)1mg,每周1次,共3周。雾化吸入OVA每天1h(含OVA80μg),连续10d。依据吸入OVA时间分为A、B、C、D、E5组,每组10只。A组雾化吸入10d后处死。B、D组继续每天1次,每次1h,每周5次,分别吸入OVA4周及8周,然后每天1h,连续10d吸入OVA后处死。C组停止吸入OVA4周后再次吸入OVA,每天1h,连续10d后处死。E组每天1次,每次1h,每周5次,吸入OVA4周,停止雾化吸入OVA4周,然后每天1h,连续10d吸入OVA后处死。测定各组小鼠支气管肺泡灌洗液(BALF)中细胞总数,嗜酸粒细胞、淋巴细胞、CD4+、CD8+、CD4+IL-10+分类及BALF中白细胞介素4(IL-4)、γ干扰素(IFN-γ)、IL-10的含量。测定血清中IL-4、IFN-γ、IL-10、OVA、IgE、IgG1、IgG2a水平,并对各组小鼠肺组织病理学进行分析。结果空白对照组BALF中嗜酸粒细胞、B淋巴细胞、CD4+IL-10+细胞分别为0.010±0.000、2.1±1.9、4.9±1.5,A组分别为0.480±0.110、5.1±2.6、5.1±2.3,B组分别为0.120±0.020、8.9±3.6、10.4±3.6,C组分别为0.560±0.050、4.7±1.7、6.3±3.1,D组分别为0.070±0.030、10.1±2.9、12.7±4.5,E组分别为0.680±0.030、5.6±3.2、6.1±3.4,各组间比较差异有统计学意义(F值分别为36.46、31.89、167.89,P均<0.01)。B、D组BALF中CD4+IL-10+细胞数与A组比较差异有统计学意义(q=5.8、6.4,P均<0.05);空白对照组BALF中IL-4、IL-10水平分别为(21±3)pg/ml、(44±12)pg/ml,A组分别为(128±23)pg/ml、(68±18)pg/ml,B组分别为(54±12)pg/ml、(127±27)pg/ml,C组分别为(133±21)pg/ml、(78±17)pg/ml,D组分别为(8±18)pg/ml、(135±34)pg/ml,E组分别为(143±26)pg/ml、(76±15)pg/ml,组间比较差异有统计学意义(F分别为37.20、143.78,P均<0.01)。B、D两组BALF中IL-10水平与A组比较差异有统计学意义(q分别为7.8、9.6,P均<0.05)。结论持续吸入变应原可使小鼠气道炎症减轻,产生免疫耐受,调节T淋巴细胞产生的IL-10参与了耐受形成。     

异氟醚吸入对新生大鼠认知功能的影响及p38MAPK抑制剂的干预作用

目的探讨p38细胞内丝裂原活化蛋白激酶通路（MAPK）通路在异氟醚诱发新生大鼠认知功能障碍中的作用。方法将60只7日龄SD大鼠随机分为5组各12只。其中A组正常饲养;B、C组均置于自制麻醉气体吸入箱中并分别吸入1.2%、1.8%异氟醚,D、E组处理分别同B、C组,但在吸入异氟醚前30 min腹腔注射p38MAPK抑制剂SB203580。6周后各组均进行行为学实验观察逃逸潜伏期（T1）、空间探索时间（T2）,并取海马组织采用Western blot法测定p38蛋白含量。结果 B、C组T1均显著长于A组,而D、E组分别显著短于B、C组（P均〈0.05）;B、C组T2均显著短于A组,而D、E组分别显著长于B、C组;B、C组海马区p38蛋白含量显著高于A组,而D、E组分别显著低于B、C组（P均〈0.05）。结论异氟醚诱发新生大鼠认知功能障碍与p38MAPK通路活化有密切关系,抑制该通路活化可减轻认知功能损害。     

甘露醇与尼莫通合用治疗大鼠脑缺血再灌注损伤机制的研究

目的 探讨甘露醇与尼莫通联合应用治疗大鼠局灶性脑缺血再灌注损伤的机制。 方法将60只SD大鼠随机分成5组,每组12只。A组:假手术组;B组:局灶性脑缺血再灌注组;C组:甘露醇组;D组:尼莫通组;E组:甘露醇与尼莫通合用组。采用硝酸还原酶法及TUNEL法分别检测各组大鼠脑组织中一氧化氮的含量及神经细胞凋亡数目。 结果 (1)B组大鼠脑组织一氧化氮含量及神经细胞凋亡数目分别为(0.54±0.11)μmol／g蛋白及(46.6±11.0)个／高倍视野;A组大鼠一氧化氮含量(0.19±0.03)μmol／g蛋白及神经细胞凋亡数目为(1.1±0.6)个／高倍视野(P<0.01);(2)C、D、E组大鼠脑组织一氧化氮含量分别为(0.32±0.07)μmol／g蛋白、(0.29±0.04)μmol／g蛋白及(0.24±0.02)μmol／g蛋白,神经细胞凋亡数目分别为(30.8±9.2)个／高倍视野、(25.7±9.6)个／高倍视野及(15.5±4.7)个／高倍视野,均低于B组(P<0.05)。(3)E组分别与C、D组大鼠比较,疗效最显著(P<0.05)。结论 甘露醇、尼莫通及两药合用均可通过下调缺血再灌注后大鼠脑组织一氧化氮含量,从而减少神经细胞凋亡,发挥其脑保护作用,两药合用效果更佳。     

缺氧诱导因子1α与其脯氨酰羟化酶相互调控对大鼠缺氧性肺动脉高压的作用

目的探讨慢性缺氧性肺动脉高压(HPH)形成过程中缺氧诱导因子1α亚基(HIF-1α)及脯氨酰羟化酶(PHD1、PHD2、PHD3)在肺内的动态表达及相互调控作用。方法 40只成年雄性 SD 大鼠随机分为常氧对照组(C 组)和缺氧3 d、7 d、14 d、21 d 组(H_3、H_7、H_(14)、H_(21)组),每组8只,常压缺氧复制慢性 HPH 大鼠模型。测各组大鼠平均肺动脉压(mPAP)、右心室肥大指数(RVHI)、血管形态学指标;原位杂交、逆转录-聚合酶链反应(RT-PCR)检测肺内 HIF-1α及 PHD1、PHD2、PHD3的mRNA 表达水平,免疫组化、Western blot 检测其蛋白表达水平。结果(1)H_7组大鼠 mPAP 为(21.7±2.4)mm Hg(1mm Hg=0.133 kPa),管壁面积与血管面积比值为(43.9±5.3)%,肺小血管中膜厚度分别为(10.0±0.7)μm,与 C 组[mPAP 为(16.6±1.6)mm Hg,管壁面积与血管面积比值为(36.3±4.8)%,肺小血管中膜厚度为(8.5±1.3)μm]比较差异均有统计学意义(q 分别5.591、4.082、2.929,P 均<0.05),H_(14)组稳定于高水平;H_(14)组 RVHI 为(27.6±1.4)%,与 C 组[(23.6±2.9)%]比较差异有统计学意义(q=5.817,P<0.05);(2)HIF-1α蛋白在 C 组肺小动脉表达不明显(0.080±0.009),H_3组表达均开始升高(0.196±0.018,与 C 组比较 q=18.864,P<0.05),H_7组达高峰(0.203±0.022),H_(14)和 H_(21)组表达稍下降(0.174±0.020、0.156±0.016)。HIF-1α mRNA 在 C 组肺小动脉(0.139±0.017)和肺组织表达阳性,H_(14)组表达略升高(0.176±0.019,与 C 组比较 q=5.401,P<0.05);(3)C 组 PHD1、PHD2 mRNA(0.260±0.031、0.196±0.023)和蛋白(0.244±0.030、0.205±0.025)呈阳性表达,PHD3 mRNA、蛋白表达相对不明显(0.110±0.013、0.153±0.019)。PHD1 mRNA 缺氧后无显著变化,H_3组 PHD2、PHD3 mRNA 表达升高(0.246±0.023、0.262±0.025,与 C 组比较 q 值分别为5.268、15.831,P 均<0.05),PHD3 mRNA 升高更明显。PHD1蛋白缺氧14 d下降(0.210±0.023,与 C 组比较 q=3.885,P<0.05),缺氧21 d 保持较低水平,H_(14)组 PHD3蛋白显著升高(0.259±0.024,与 C 组比较 q=12.975,P<0.05),缺氧7 d 保持高水平,H_(14)、H_(21)组下降(0.206±0.025、0.189±0.019,与 H_7组比较 q 分别6.441、8.526,P<0.05)。直线相关分析结果表明,HIF-1α蛋白与 PHD2、PHD3 mRNA 呈显著正相关(r 分别为0.580、0.690,P 均为0.000),缺氧组大鼠 PHD2蛋白与 HIF-1α蛋白表达呈负相关(r=-0.704,P<0.05)。结论在缺氧性肺动脉高压形成过程中 HIF-1α表达调节主要发生在转录后水平,HIF-1的表达增加可能激活 PHD2、PHD3的转录,在一定程度上对 HIF-1α亚基的表达具有负反馈调节作用。PHD 可能还存在其他转录后调节机制。     

Rosiglitazone prevents murine hepatic fibrosis induced by Schistosoma japonicum

AIM： To evaluate the effect of rosiglitazone in a murine model of liver fibrosis induced by Schistosoma japonicum infection. METHODS： A total of 50 mice were randomly and averagely divided into groups A, B, C, D and E. The mice in group A served as normal controls, while those in the other four groups were infected with Schistosoma japonicum to induce the model of liver fibrosis. Besides, the mice in groups C, D and E were treated with praziquantel, rosiglitazone and praziquantel plus rosiglitazone, respectively. NF-κB binding activity and expression of PPARγ-mRNA were determined by Western blot assay and real-time quantitative PCR. Radioimmunonassay technique was used to detect the serum content changes of TNF-α and IL-6. Histological specimens were stained with HE. Expression of TGF-β1, a-smooth muscle actin and type Ⅰand type Ⅲ collagen was detected by immunohistochemistry and multimedia color pathographic analysis system.
RESULTS： Inflammation and fibrosis in the rosiglitazone plus praziquantel treatment group （group E） were lightest among the mice infected with Schistosoma （P 〈 0.05）. To further explore the mechanism of rosiglitazone action, we found that rosiglitazone can significantly increase the expression of PPARγ [E： -18.212 ± （-3.909） vs B： -27.315 ± （-6.348） and C： -25.647 ± （-5.694）, P 〈 0.05],reduce the NF-κB binding activity （E： 88.89 ± 19.34 vs B： 141.11 ± 15.37, C： 112.89 ± 20.17 and D： 108.89 ± 20.47, P 〈 0.05）, and lower the serum level of TNF-α （E： 1.613 ± 0.420 ng/mL vs B： 2.892 ± 0.587 ng/mL, C： 2.346 ± 0.371 ng/mL and D： 2.160 ± 0.395 ng/mL, P 〈 0.05） and IL-6 （E： 0.106 ± 0.021 ng/mL vs B： 0.140 ± 0.031 ng/mL and C： 0.137 ± 0.027 ng/mL, P 〈 0.05） in mice with liver fibrosis. Rosiglitazone can also substantially reduce the hepatic expression of TGF-β1, α-SMA type Ⅰand type Ⅲ collagen in mice with liver fibrosis. CONCLUSION： The activation of PPARγ by its ligand can retard liver fi     

Effects of emodin and double blood supplies on liver regeneration of reduced size graft liver in rat model

AIM: To study the influences of emodin and reconstruction of double blood supplies on liver regeneration of reduced size graft liver in rat model. METHODS: A total of 45 SD-SD rat reduced size liver transplantation models were randomly divided into three groups (A-C). The conventional reduced size liver transplantation was performed on rats in group A, while the hepatic artery blood supply was restored in groups B and C. The emodin (1.5 mg/kg/d) was given by intraperitoneal route in group C only. The recipients were killed on the seventh day after the operation. The proliferative cell nuclear antigen (PCNA), TBil and ALT of serum were detected, and the pathological changes of liver cell were observed. RESULTS: The numbers of the rats that survived in A, B, and C group on the seventh day after operation were 14, 13, 13, respectively. The levels of TBil (31.5±5.2 μmol/L, 23.2±3.1 μmol/L vs 38.6±6.8 μmol/L), and ALT (5 351±1 050 nKat, 1300±900 nKat vs 5779±1202 nKat) in serum in groups B and C were lower than those in group A (P<0.05), while the expression of PCNA in groups B or C was higher than that in group A (22.0±3.5%, 28.2±4.2% vs 18.6±3.2%, P<0.05). The deeper staining nuclei, double nuclei, multi-nuclei and much glycogen were observed in liver cells of groups B and C, especially in group C, while fewer were found in liver cells of group A. CONCLUSION: The reconstruction of arterial blood supply is very important for rat liver regeneration after reduced size liver transplantation. Emodin has the effect of promoting liver regeneration and improving liver function in rats after reduced size transplantation. The possible mechanism is improving proliferation of liver cell and protecting liver cells from injury.     

Effect of 1,25-dihydroxyvitamin D3 on preventing allograft from acute rejection following rat orthotopic liver transplantation

AIM: To study the mechanism and the preventive role of 1,25-dihydroxyvitamin D3 in acute rejection following orthotopic liver transplantation.METHODS: Rats were randomly divided as donors or recipients for orthotopic liver allotransplantation model. Four groups were designed in the study, Group Ⅰ: syngenic control(Wistar to Wistar); Group Ⅱ: acute rejection (SD to Wistar);Group Ⅲ: acute rejection treated with cyclosporine A, and Group Ⅳ: acute rejection treated with 1,25-(OH)2D3. Liver function, rejection activity index and mRNA of IFN-γ, IL-10 intragraft in recipients were measured on day 1, 5, 7, 15,30 posttransplant for assessing graft function, severity of acute rejection and immune state of recipients.RESULTS: Survival time of recipients in Group Ⅳ was significantly prolonged (4/6 recipients survived for over 100 days. vs Group Ⅱ, P&lt;0.001; vs Group Ⅲ, P&gt;0.05). After treatment with 1,25-(OH)2 D3, mean value of all the assay tested on each experimental time was compared, liver function in group IV was significantly improved (AST 127&#177;41U/L-360&#177;104U/L, BIL 13&#177;5mmol/l-38&#177;11mmol/l; vs Group Ⅱ, P&lt;0.05; vs Group Ⅲ, P&gt;0.05. Rejection activity index was significantly decreased (0-3.3&#177;1.6; vs Group Ⅱ, P&lt;0.05;vs Group Ⅲ, P&gt;0.05). Level of hepatic IFN-γ, mRNA in group IV was decreased, while level of hepatic IL-10 mRNA was increased (vs Group Ⅱ, P&lt;0.05; vs Group Ⅲ, P&gt;0.05).CONCLUSION: Our results indicated that 1,25-(OH)2D3 induced the secretion of oltokine toward to Th2 type, which would alleviate acute rejection, protect liver function and prolong survival of recipient after orthotopic liver transplantation.     

p38信号转导途径对离体肝脏缺血再灌注损伤的影响

目的 研究离体肝脏缺血再灌注期间p38信号转导途径的激活对其损伤程度的影响。 方法 通过自行建立的兔离体肝脏缺血再灌注模型,将一定剂量的特异性p38丝裂原激活蛋白激酶(MAPK)抑制剂SB202190加入到保存液中,根据原位灌注液及保存液中加入SB202190的剂量再将离体肝分为A、B、C、D4组。分别于冷保存前、冷保存末及再灌注5、10、15、30、60、120 min获取离体肝组织及受体兔静脉血液标本。分别应用免疫印迹杂交和免疫沉淀法测定离体肝组织磷酸化p38M A P K的活性。用全自动生化分析仪测定离体兔肝功酶学含量;并测定再灌注120 min内的胆汁分泌总量。 结果 在正常肝组织中p38 MAPK即有一定的基础活性;经冷保存后有一定的升高,再灌注10 min时达到峰值,然后逐渐下降,至再灌注120 min时降低至正常水平,而在冷保存液中加入SB202190后,再灌注期间p38 MAPK的活性受到显著性抑制,其中B、C、D组p38 MAPK活性峰值仅分别为A组的53.9％,12.8％,9.6％;再灌注期间各组受体肝酶学水平均表现为A>B>C>D,而胆汁分泌量表现为A相似文献   

Trans-arterial gene therapy for hepatocellular carcinoma in a rabbit model

AIM： To study the effect of adenovirus （Ad）-p53 gene therapy on hepatocellular carcinoma （HCC） in a rabbit model.
METHODS： VX2 tumor was grown in the liver of 24rabbits. Animals were divided into four groups： group A receiving trans-arterial gene therapy （Ad-p53） only,group B receiving combined Ad-p53 therapy and transarterial embolization （lipiodol）, group C receiving transarterial chemoembolization （lipiodol ＋ mitomycin C）,control group （D） receiving sodium chloride. Tumor volume （V1） was measured by using MRI （d 13）.Interventional procedure was applied （d 14）.Tumor volume （V2） was assessed by MRT （d 21） and the mean ratio （V2/V1） was calculated. After the second MRI,specimens of the liver were abstained and examined immunohistochemically using mutant-type p53 antibody.The positive expression was scored.
RESULTS： Compared with control group （（^-x） = 3.14± 0.64）, therapeutic groups all showed a significant decrease in the tumor growth ratio （P 〈 0.05）. A slight difference was found between group A （（^-x） =2.35 ±0.59） and group B （（^-x） = 1.75 ± 0.28） （P = 0.048）. Nostatistically significant difference was observed between group B and group C （（^-x） = 2.00 ± 0.44）. The positive expression rate of mutant-type p53 was the lowest in group B and significantly different between group A and group C （P 〈 0.05）.Compared to the control subjects, groups A and C both showed a decrease in the expression of mutant-type p53, but there was no significant difference between them.
CONCLUSION： Trans-arterial Ad-p53 gene therapy can reduce tumor growth of HCC in rabbit model.     

Early steroid withdrawal after liver transplantation for hepatocellular carcinoma

AIM: To evaluate the impact of early steroid withdrawal on the incidence of rejection, tumor recurrence and complications after liver transplantation for advanced- stage hepatocellular carcinoma. METHODS: Fifty-four patients underwent liver transplantation for advanced-stage hepatocellular carcinoma from April 2003 to June 2005. These cases were divided into a steroid-withdrawal group (group A, n = 28) and a steroid-maintenance group (group B, n = 26). In group A, steroid was withdrawn 3 mo after transplantation. In group B, steroid was continuously used postoperatively. The incidence of rejection, 6-mo and 1-year recurrence rate of carcinoma, 1-year survival rate, mean serum tacrolimus trough level, and liver and kidney function were compared between the two groups. RESULTS: In the two groups, no statistical difference was observed in the incidence of rejection (14.3 vs 11.5%, P > 0.05), mean serum tacrolimus trough levels (6.9 ± 1.4 vs 7.1 ± 1.1 μg/L, P > 0.05), liver and kidney function after 6 mo [alanine aminotransferase (ALT): 533 ± 183 vs 617 ± 217 nka/L, P > 0.05; creatinine: 66 ± 18 vs 71 ± 19 μmol/L, P > 0.05], 6-mo recurrence rate of carcinoma (25.0 vs 42.3%, P > 0.05), and 1-year survival rate (64.2 vs 46.1%, P > 0.05). The 1-year tumor recurrence rate (39.2 vs 69.2%, P < 0.05), serum cholesterol level (3.9 ± 1.8 vs 5.9 ± 2.6 mmol/L, P < 0.01) and fasting blood sugar (5.1 ± 2.1 vs 8.9 ± 3.6 mmol/L, P < 0.01) were signifi cantly different. These were lower in the steroid-withdrawal group than in the steroid- maintenance group. CONCLUSION: Early steroid withdrawal was safe after liver transplantation in patients with advanced-stage hepatocellular carcinoma. When steroids were withdrawn 3 mo post-operation, the incidence of rejection didnot increase, and there was no demand to maintain tacrolimus at a high level. In contrast, the tumor recurrence rate and the potential of adverse effects decreased signifi cantly. This may have led to an increase in long-term survival rate.     

外源性一氧化碳对小肠缺血再灌注大鼠不同组织p38 MAPKs蛋白表达的影响

目的:探讨外源性一氧化碳(carbonmonoxide,CO)对小肠缺血再灌注(intestinal ischemia-reperfusion,IIR)所致多器官损伤防治作用的机制.方法:♂Wistar大鼠64只,随机分为8组,给予不同实验方法处理:A组:假手术对照组,不阻断肠系膜上动脉(superior mesenteric artery,SMA),其余手术过程同其他组;B组:小肠缺血再灌注组,经T型管吸入空气;C组:缺血前10 min CO吸入组,按吸入CO浓度(100μL/L,250μL/L)分为两个亚组(C1组和C2组);D组:再灌注开始时CO吸入组,按吸入CO浓度(100μL/L,250μL/L)分为两个亚组(D1组和D2组);E组:再灌注后60 min CO吸入组,按吸入CO浓度(100μL/L,250μL/L)分为两个亚组(E1组和E2组).实验结束时取不同组织以免疫蛋白印迹杂交法检测p38 MAPKs的密度表达.结果:与对照组A组比较,单纯IIR的B组的小肠、肺、肝组织中p38 MAPKs蛋白表达升高,但不显著(0.468±0.213 vs 0.474±0.151;0.439±0.111vs 0.482±0.103;0.622±0.112vs 0.654±0.016,all P>0.05);与单纯IIR的B组比较,外源性应用CO的C1、C2、D1、D2、E1、E2组的小肠、肺、肝组织中p38 MAPKs蛋白表达均明显增高(1.540±0.346,1.626±0.277,1.365±0.233,1.483±0.265,1.353±0.234,1.372±0.2731 vs 0.474±0.151;1.654±0.211,1.701±0.101,1.398±0.245,1.444±0.272,1.288±0.218,1.366±0.244 vs 0.482±0.103;1.695±0.234,1.723±0.213,1.423±0.221,1.586±0.254,1.322±0.261,1.411±0.296 vs 0.654±0.016,均P<0.05).结论:调节细胞内p38 MAPKs表达是外源性CO防治IIR所致多器官损伤作用的分子生物学基础之一.     

Age-associated decline in cardiac allograft rejection

The influence of age on cardiac allograft rejection was studied in 57 consecutive recipients. Twenty-one subjects were 54 years of age or older (mean, 57.7 +/- 0.6 years [+/- SEM]; range, 54 to 63 years) and 36 subjects were 52 years of age or younger (mean, 39.9 +/- 1.8 years; range, 16 to 52 years; p less than 0.001). The older recipients had fewer rejection episodes during the first four months following cardiac transplantation (0.24 +/- 0.05 episodes per month versus 0.72 +/- 0.09 episodes per month; p less than 0.001) and during the total duration of follow-up (0.20 +/- 0.03 episodes per month versus 0.40 +/- 0.07 episodes per month; p = 0.045), and experienced their first rejection episode later (50.4 +/- 4.0 days versus 27.7 +/- 8.5 days; p = 0.008). Younger age was found to add significantly as a predictor of rejection in a multivariate analysis that controlled for sex, immunosuppressive agents, cause of heart failure, and pretransplantation lymphocyte cross-match status (r = 0.64, p less than 0.05). Decreased rejection frequency occurred without a concomitant increase in the serious infection rate (67 percent in both groups). The 12-month actuarial survival was 100 percent in the older group and 94 percent in the younger group (p = NS). Decreased rejection in the older recipients is likely a manifestation of an age-associated decline in immune function and might represent an advantage in transplantation for carefully selected older patients.