Angiotensin converting enzyme inhibition decreases cell turnover in the neonatal rat heart

The renin angiotensin system plays an important role in growth and development. Exposure of the neonate to an ACE inhibitor increases mortality and results in growth retardation and abnormal development. We have demonstrated that ACE inhibition in the developing kidney increases apoptosis and decreases cell proliferation, which may account for renal growth impairment. To evaluate the role of endogenous angiotensin in cardiac development, the relationship between ACE inhibition, cell proliferation, apoptosis, several modulators of apoptosis (bcl-2, bcl-xl, and clusterin) was examined in the developing rat heart. Thirty-five newborn rat pups were treated with enalapril (30 mg/kg/d) or a vehicle (control group) for 7 d, and hearts were removed for rt-PCR and Western blotting of bcl-2, bcl-xl, and clusterin. An additional 10 rat pups were treated with hydralazine (10 mg/kg/d) or a vehicle, to serve as a hypotensive control. Cell proliferation was determined by PCNA immunostaining, and apoptosis was detected using the total TUNEL technique. Enalapril treatment resulted in a 24% mortality, reduced body weight, and decreased heart weight (p < 0.05). Enalapril decreased proliferating myocytes by 23%, and reduced proliferating cardiac interstitial cells by 8.1% (p < 0.05). Enalapril also decreased myocytes apoptosis by 60%, but the proportion of myocytes undergoing apoptosis was 10-fold less than that of proliferating cells. Cardiac bcl-2 mRNA, clusterin mRNA, bcl-2 protein, and bcl-xl protein content were not changed, but clusterin protein expression was decreased by enalapril treatment. Hydralazine did not alter cardiac cell proliferation or apoptosis. We conclude that ACE inhibition decreases cell turnover in the developing rat heart, which may contribute to cardiac growth impairment. The loss of myocytes may lead to greater myocyte hypertrophy and myocardial damage during later life.     

p38丝裂原活化蛋白激酶调节急性淋巴细胞性白血病CEM细胞株糖皮质激素受体功能的分子机制研究

目的探讨p38丝裂原活化蛋白激酶（p38MAPK）对地塞米松（DEX）诱导急性淋巴细胞性白血病CEM细胞株凋亡过程中糖皮质激素受体α（GRα）功能的影响。方法台盼蓝拒染法绘制增殖曲线;流式细胞仪解析细胞凋亡;Western blot检测糖皮质激素受体蛋白及磷酸化p38MAPK蛋白表达。结果（1）p38MAPK特异阻滞剂SB203580预处理后继续培养24-72h,细胞的增殖率分别由单用DEX时的62．3％、35．5％、11．6％升至82．8％、54．7％和48．1％（P〈0．01）;（2）5μmol／LDEX处理CEM细胞36h时,亚二倍体细胞为26．2％,显著高于对照组（P〈0．01）。SB203580预处理后,亚二倍体细胞由26．2％降至7．1％（P〈0．01）;（3）5μmol／L的DEX处理CEM细胞后,GRα总蛋白的表达水平从12h开始分别上调至117％（12h）、121％（24h）、122％（36h）、125％（48h）。未与DEX结合的GRα主要存在于细胞浆中,细胞核GRα与细胞浆GRα之比为0．27。从6-48h,GRα的胞核与胞浆之比分别为0．48、0．59、0．95、2．16、4．08;（4）5μmol／LDEX处理CEM细胞后,p-p38MAPK蛋白在15min时即隐约可见,1h开始表达增强并持续增强至6h,之后减弱至48h仍可见。SB203580预处理后,未检测到p-p38MAPK蛋白的表达;（5）SB203580预处理后,GRα的核浆比例由4．08降至0．43（P〈0．05）,GRα总蛋白无明显变化。结论DEX在诱导CEM细胞凋亡时上调GRα的蛋白表达水平并促进转位人核。p38MAPK信号转导通路的活化促进了GRα转位人核。     

谷氨酰胺对内毒素血症幼年大鼠肾脏ERK以及p38表达的影响

目的：通过观察谷氨酰胺(glutamine, Gln)对内毒素血症幼年大鼠肾脏信号转导通路ERK-2、p38及其mRNA表达的影响,探讨谷氨酰胺在大鼠内毒素血症中通过干预肾脏ERK-2以及p38信号转导途径来保护肾脏的机制。方法：选健康18日龄Wistar大鼠121只,按腹腔注射药物不同分为：0 h对照组(生理盐水,n=11), 内毒素组（LPS,n=55）,谷氨酰胺组(Gln, n=55); 后两组又分为2,4,6,24及72 h共5个时间点,每个时间点11只,在各指定时间点分别将大鼠断头分离肾脏,8只用逆转录 聚合酶链反应（RT-PCR）方法测定ERK-2以及p38的mRNA的表达,另3只取肾组织后用甲醛固定,制成石蜡切片,用免疫组化方法确定ERK-2以及p38蛋白质的表达。结果：LPS组各时点ERK-2 mRNA, p38MAPK mRNA, ERK-2, p38MAPK表达均较对照组增强,最高点在 6 h,差异有显著性,P<0.01;Gln组各时点ERK-2 mRNA, p38MAPK mRNA, ERK-2, p38MAPK表达趋势与LPS组一致,但明显低于LPS组,P<0.05或P<0.01。结论：ERK及p38在内毒素血症表达均明显增强。谷氨酰胺可使其表达下调,从而减轻肾脏损伤。［中国当代儿科杂志,2009,11（4）：301-305］     

急性淋巴细胞性白血病患儿血清高迁移率族蛋白1检测及其诱导白血病细胞分泌肿瘤坏死因子α的实验研究

目的探讨血清高迁移率族蛋白1（HMGB1）水平与儿童急性淋巴细胞性白血病（ALL）发病的关系;研究HMGB1对白血病细胞分泌肿瘤坏死因子α（TNF-α）的影响及其与丝裂原活化蛋白激酶（MAPK）信号通路的关系。方法采用Westernblot方法检测正常、ALL患儿初治期、完全缓解期各15例血清HMGB1水平;制备HMGB1重组蛋白刺激人K562白血病细胞损伤模型,采用ELISA和Westernblot方法检测HMGB1对TNF-Or．分泌及其p38、c-Jun氨基末端激酶（JNK）、细胞外信号调节激酶（ERK）MAPK信号通路活化的影响;采用ELISA方法检测MAPK抑制剂对HMGB1所致TNF-α．分泌的影响。结果初治ALL患者血清HMGB1[（43．78±4．62）μg／L]显著高于正常对照组[（0．60±0．48）μg／L,P〈0．01]和缓解组[（0．89±0．62）μg／L,P〈0．01],正常对照组与缓解组比较差异无统计学意义（P〉0．05）;HMGB1以时效和量效依赖性方式诱导K562细胞分泌TNF-α．并且促进了p38、JNK、ERK蛋白发生磷酸化;p38抑制剂（SB203580）、JNK抑制剂（SP600125）和MEK抑制剂（PD98059）抑制了HMGB1诱导K562细胞TNF-α．的分泌。结论血清HMGB1水平与儿童ALL发病、发展密切相关;HMGB1通过活化MAPK信号通路促进白血病细胞分泌TNF-α．参与肿瘤细胞免疫调节。     

p38丝裂素活化蛋白激酶在幼鼠高氧肺损伤模型中的表达及作用

目的：近年来的研究证实p38丝裂素活化蛋白激酶（p38MAPK）在多种因素诱导的肺损伤中发挥着重要的调控作用,但其在高氧肺损伤中的表达和作用尚不明了。该文通过建立幼鼠高氧肺损伤模型来研究p38MAPK在该模型中的表达及作用。方法：90％氧气暴露建立幼年Wistar大鼠高氧肺损伤模型,用免疫组化和蛋白质免疫印迹法观察p38MAPK在肺组织中的分布和表达,用TUNEL法观察肺组织的细胞凋亡指数,并同时观察p38MAPK抑制剂SB203580对肺组织细胞凋亡的影响。结果：高氧暴露后肺组织磷酸化p38MAPK蛋白表达明显增强,主要分布于肺泡上皮细胞、气道上皮细胞、血管内皮细胞、浸润炎症细胞。同时肺凋亡指数明显增加。使用SB203580抑制了p38MAPK活性的上升,同时降低了肺凋亡指数。结论：高氧诱导的急性肺损伤模型中,磷酸化的p38MAPK表达增加,并表达于肺内多种细胞,可能起促凋亡作用。［中国当代儿科杂志,2009,11（5）：389-392］     

人巨细胞病毒感染对人神经胶质瘤U251细胞p38MAPK表达和细胞凋亡及细胞周期的影响

目的研究人巨细胞病毒(HCMV)感染对人神经胶质瘤U251细胞p38MAPK表达水平、细胞凋亡及细胞周期的影响。方法以免疫印迹法(WesternBlotting)分析HCMV感染人神经胶质瘤U251细胞后不同时间总p38MAPK、磷酸化p38MAPK蛋白表达水平;用流式细胞仪检测HCMV感染后人神经胶质瘤U251细胞凋亡百分率变化和细胞周期的分布。结果HCMV感染后不同时间U251细胞总p38蛋白的表达水平不变,但磷酸化p38于HCMV感染后6至10h明显增高,6h、10h平均灰度值分别为186.33±7.51和188.00±7.02,与对照组比较差异有统计学意义,16h后下降至基础水平。HCMV感染能诱导U251细胞凋亡,于感染后第5天凋亡率最高,达(10.18±1.24)%,与对照组比较差异有统计学意义。HCMV感染使G1期细胞明显减少,感染后2、5、7d分别减少至(56.50±2.57)%、(62.33±2.64)%和(67.45±4.44)%,与对照组比较差异均有统计学意义,细胞周期阻滞于S期和G2期。结论HCMV感染U251细胞能够激活p38MAPK,并诱导U251细胞凋亡、细胞周期阻滞。     

IL-6 is constitutively expressed during lung morphogenesis and enhances fetal lung explant branching

Previous studies have shown that chorioamnionitis, with increased IL-6, promotes fetal lung maturation and decreases the incidence of respiratory distress syndrome in premature neonates. However, the expression pattern and the effects of IL-6 on fetal lung growth mechanisms remain unknown. IL-6 expression was assessed by in situ hybridization and by real-time PCR between 14.5 and 21.5 d postconception. Normal and nitrofen-induced hypoplastic lung explants were cultured with increasing IL-6 doses or IL-6 neutralizing antibodies. Branching, cellular proliferation (Ki-67) and MAPK phosphorylation in fetal lung explants were analyzed. Pulmonary primitive epithelium expressed IL-6 constitutively throughout all gestational ages, displaying highest levels during earliest stages. In normal and hypoplastic lung explants, IL-6 neutralizing antibodies significantly reduced, whereas IL-6 supplementation induced a biphasic effect (lower doses increased, while the highest dose did not accomplish additional effect) on branching and cellular proliferation. IL-6 enhanced p38-MAPK phosphorylation without changing MEK1/2 and JNK pathways. The present study suggests a physiological role for IL-6 on pulmonary branching mechanisms most likely involving p38-MAPK intracellular signalling pathway.     

Morphologic correlates of renal growth arrest in neonatal partial ureteral obstruction

To investigate the morphologic correlates of decreased renal blood flow and growth arrest resulting from chronic partial ureteral obstruction in the neonate, guinea pigs were subjected to unilateral ureteral constriction within the first 2 days of life and were studied at 3 and 8 wk of age. Severity of histologic changes in the obstructed and intact contralateral kidney was assessed by light microscopy. Morphometric glomerular measurements were made using a computerized tracing device. Since contralateral nephrectomy or administration of angiotensin converting enzyme inhibitor (enalapril) result in increased blood flow to the obstructed kidney, morphologic changes were also examined in separate groups of animals subjected to these maneuvers, and were compared to appropriate controls. Ipsilateral chronic partial ureteral obstruction resulted in decreased glomerular volume (p less than 0.0001) and increased glomerular crowding associated with tubular dilatation and progressive glomerular sclerosis, tubular atrophy, and interstitial fibrosis. The intact contralateral kidney underwent hypertrophy with increase in glomerular volume (p less than 0.0001) and decreased glomerular density. Contralateral nephrectomy prevented the decrease in glomerular volume in the obstructed kidney and resulted in decreased glomerular density and reduced tubular atrophy at 3 wk of age. Enalapril prevented the decrease in glomerular volume at 3 wk of age, but glomerular and tubular changes progressed and were unaffected by enalapril at 8 wk. Left kidney glomerular volume was directly related to renal blood flow (r = 0.71, p less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)     

狼疮性肾炎患儿肾组织细胞凋亡及凋亡相关蛋白PDCD5、Caspase-3的表达

目的探讨狼疮性肾炎(LN)肾脏细胞凋亡与细胞增殖的关系,加强LN发病机制的认识。方法用原位末端标记法(TUNEL)检测31例LN患儿及9例对照的肾组织细胞凋亡,用免疫组化及图像分析的方法检测PCNA(增殖细胞核抗原)及PDCD5、Caspase3的表达。结果(1)LN患儿肾组织中肾小球和肾小管的凋亡细胞、增殖细胞数、增殖细胞数/凋亡细胞数(P/A值)均明显高于对照组(LN组及对照组凋亡细胞、增殖细胞数、P/A值在肾小球分别为0.86±0.26比0.14±0.09,8.45±2.83比0.47±0.25,10.01±2.96比3.37±1.93,均P<0.01;在肾小管分别为1.16±0.42比0.16±0.07,13.73±3.54比1.32±0.15,12.81±3.91比8.94±1.79,均P<0.05)。(2)PDCD5在LN患儿肾小球的表达强度与对照组相比差异无统计学意义(0.09±0.05比0.08±0.02,P>0.05),在肾小管的表达强度明显低于对照组(0.13±0.05比0.21±0.07,P<0.01)。(3)Caspase3在LN患儿肾小球、肾小管的表达强度明显高于对照组(肾小球0.22±0.07比0.05±0.02,肾小管0.08±0.03比0.05±0.01,均P<0.01)。(4)直线相关分析显示:全部观察标本中肾小球凋亡细胞数与Caspase3的表达强度呈正相关(r=0.718,P<0.01),与PDCD5的表达强度无显著相关性(r=0.054,P>0.05);全部观察标本中肾小球、肾小管PDCD5的表达强度均与肾小球、肾小管Caspase3的表达强度无明显相关性(r=0.061,P>0.05,r=0.049,P>0.05)。多元逐步回归分析显示:在α=0.15水平上,全部观察标本中肾小管凋亡细胞数与PDCD5的表达强度呈负相关(回归系数为-3.686,t=-2.875,P=0.007),与Caspase3的表达强度呈正相关(回归系数为4.761,t=1.772,P=0.085)。结论(1)LN患儿肾组织细胞凋亡虽然比对照组增加,但相对细胞增殖来说表现为不足;(2)Caspase3参与了LN患儿肾组织肾小球、肾小管的细胞凋亡;(3)尚不能证实PDCD5参与了LN患儿肾小球细胞凋亡;但可能参与了肾小管的细胞凋亡。     

Pax2、增殖细胞核抗原、细胞凋亡在肾病综合征激素耐药中的作用

目的观察原发性肾病综合征（PNS）激素敏感与激素耐药组患儿Pax2、增殖细胞核抗原（PCNA）表达及细胞有效凋亡率，探讨Pax2在肾脏檐皮质激素（GC）耐药中的作用。方法采用免疫组织化学法观察PNS患儿40例肾活检组织Pax2、PCNA表达，采用荧光显微镜检测细胞凋亡。结果激素敏感组Pax2适量表达，与PCNA阳性表达呈正相关,激素耐药组，Pax2呈过度表达，与PCNA阳性表达无明显相关性。Pax2始终与细胞凋亡旱负相关。结论Pax2适量表达，通过促进细胞增殖及抑制凋亡，修复受损的小管-间质。Pax2过度表达，细胞增殖与凋亡进一步减少，导致肾小管病理改变加重，使其对激素不敏感共至耐药。     

p38MAPK对小儿急性肠梗阻肠坏死Occludin表达的影响及临床意义

目的 探讨p38MAPK信号通路在小儿急性肠梗阻肠坏死中对紧密连接蛋白Occludin表达的影响及临床意义.方法 选取2013年12月至2014年12月急诊入院的急性肠梗阻合并肠坏死患儿38例,术后病理切片均证实肠坏死为病变组;20例消化道畸形(梅克尔憩室、肠重复畸形无并发症)患儿为正常对照组.采用Western Blot检测切除肠管p38MAPK,p-p38MAPK含量;免疫组化法检测肠上皮间紧密连接蛋白Occludin表达(OD值);TUNEL法检测肠上皮细胞凋亡指数(AI);ELISA法检测术前、术后7d血清TNF-α含量;将各检测指标分别与Occludin蛋白OD值行相关性分析.结果 p38MAPK相对含量,病变组1.69±0.59,对照组1.89±0.63,P＞0.05;p-p38MAPK相对含量病变组1.15±0.14,对照组0.66±0.07;AI病变组52.10±1.7,对照组23.03±7.4;术前血清TNF-α含量病变组98.38±18.7,对照组50.02±11.1;Ocludin蛋白OD值病变组3.44±2.8,对照组8.66±4.6,P值均＜0.01.p-p38MAPK相对含量、AI、术前血清TNF-α含量分别与肠上皮细胞Occludin OD值相关性分析示:r=-0.616、r=-0.316、r=-0.562,三者间P＜0.01.结论 小儿急性肠梗阻肠坏死由于肠绞窄缺血缺氧p38MAPK信号通路被激活,启动细胞凋亡和TNF-α大量释放,使紧密连接Occludin蛋白表达下降,最终肠黏膜屏障破坏可能引发细菌移位.     

UCP2 基因siRNA 转染对脓毒症血清诱导心肌细胞炎症反应的影响

目的 研究解偶联蛋白2 基因（uncoupling protein 2, UCP2）siRNA 干扰大鼠H9C2 心肌细胞株后对脓毒症血清诱导炎症反应的影响,探讨UCP2 在脓毒症心肌病可能的机制。方法 制备正常大鼠血清和脓毒症大鼠血清。体外培养大鼠心肌细胞株（H9C2）,随机分为空白对照组、正常大鼠血清、10%脓毒症大鼠血清刺激12 h 组（脓毒症血清组）、UCP2-siRNA 干扰+10%脓毒症大鼠血清刺激12 h 组（UCP2-siRNA+ 脓毒症血清组）、阴性siRNA 转染+10%脓毒症大鼠血清刺激12 h 组（阴性siRNA+ 脓毒症血清组）。RT-PCR 检测各组TNF-α mRNA,IL-1βmRNA 表达;免疫印迹法检测磷酸化p38MAPK（p-p38MAPK）表达和核转录因子（NF-κB）的表达。结果 UCP2-siRNA+ 脓毒症血清组 p-p38、NF-κB 表达量均较脓毒症血清组明显增高（PP结论 UCP2 参与脓毒症血清诱导的H9C2 心肌细胞p38MAPK 活性和NF-κB 与下游炎症介质的表达调控。     

新生大鼠高氧肺损伤P38活化和MMP-2 mRNA表达的研究

目的：观察高氧肺损伤新生大鼠肺组织中P38-丝裂原活化蛋白激酶（P38）的活化与基质金属蛋白酶-2（MMP-2）mRNA表达水平的变化,并探讨P38的活化对MMP-2 mRNA表达的影响。方法：将36只新生Sprague-Dawley大鼠随机分为空气组、高氧组和SB203580干预组,每组12只。建立高氧肺损伤动物模型,处理组给予相应的干预。各组分别于第3天和第7天处死大鼠,常规苏木精-伊红染色,观察肺组织病理学变化;Western blot检测肺组织P38蛋白的活化水平,RT-PCR检测肺组织MMP-2 mRNA表达的变化。结果：高氧组大鼠肺组织P38的活化和MMP-2 mRNA表达水平较空气组和干预组明显增强,差异有统计学意义（P<0.01）。结论：P38在新生大鼠高氧肺损伤中活化增强,可能参与MMP-2 mRNA表达的调控。     

Signal transduction pathways involved in oxidative stress-induced intestinal epithelial cell apoptosis

Necrotizing enterocolitis (NEC) is a devastating inflammatory condition of the gut that occurs in premature infants. Ischemia-reperfusion gut injury with production of reactive oxygen species (ROS) is thought to contribute to NEC; the exact cellular mechanisms involved are largely unknown. The purpose of this study was to determine the intracellular signaling transduction pathways involved in oxidative stress-induced intestinal epithelial cell apoptosis. H2O2 treatment resulted in rat intestinal epithelial cell apoptosis in a dose- and time-dependent manner; the caspase inhibitor, zVAD-fmk, blocked this response. Western blotting was performed to determine phosphorylation of kinases and ELISA was used to assess DNA fragmentation, as a measure of apoptosis. A rapid increase in phosphorylation of extracellular signal-related kinase (ERK)1/2, c-Jun N-terminal kinase (JNK)1/2, and Akt was noted. Inhibition of ERK and JNK decreased H2O2-induced apoptosis. Additionally, inhibition of protein kinase C (PKC) and phosphatidylinositol 3-kinase (PI3-K) attenuated and enhanced H2O2-mediated apoptosis and mitochondrial membrane potential decrease, respectively. Furthermore, activation of PKC reduced the Akt phosphorylation, whereas inhibition of PKC attenuated H2O2-mediated activation of caspase-3 and enhanced the H2O2-induced Akt phosphorylation. This study shows that activation of multiple signaling transduction pathways occurs during oxidative stress-induced intestinal epithelial cell injury. In contrast to ERK, JNK, and PKC, PI3-K/Akt may play an important role as a protective cellular signaling pathway during this process.     

不同病理类型儿童狼疮性肾炎肾小球细胞凋亡的变化

目的检测不同病理类型狼疮性肾炎(lupusnephritis,LN)患儿肾组织细胞凋亡与增殖,分析LN患儿肾组织细胞凋亡与增殖的关系。方法用原位末端标记法(TUNEL)检测21例Ⅳ型、6例Ⅴ型LN患儿肾组织及9例对照肾组织中的细胞凋亡,用免疫组织化学法检测其细胞增殖(PCNA,增殖细胞核抗原),同时用免疫组织化学及图像分析法检测细胞凋亡相关基因蛋白PDCD5、Caspase-3的表达。结果1.Ⅳ型LN患儿肾小球凋亡细胞数(0.93±0.28)、增殖细胞数(9.97±1.90)及P/A值(11.15±2.49)均明显高于Ⅴ型LN患儿相应指标(0.66±0.11),(4.82±1.46)和(7.25±1.71),P<0.05,P<0.01和P<0.01。2.Ⅳ型和Ⅴ型LN患儿肾组织PDCD5、Caspase-3表达:PDCD5在Ⅳ型LN患儿肾小球的表达强度(0.08±0.02)与V型LN患儿肾小球的表达强度(0.07±0.03)无差异,P均>0.05;Caspase-3在Ⅳ型LN患儿肾小球的表达强度(0.25±0.05)明显高于Ⅴ型LN患儿肾小球的表达强度(0.16±0.03),P<0.01。结论Ⅳ型LN患儿相对于细胞增殖的程度而上调的细胞凋亡远不如Ⅴ型LN患儿;Caspase-3参与LN患儿肾小球细胞凋亡,而PDCD5未起主要作用或其促凋亡作用被抑制。     

邻苯二甲酸正丁酯对HL-60细胞凋亡影响及其作用机制

目的探讨邻苯二甲酸正丁酯(DBP)对急性早幼粒细胞性白血病细胞系HL-60细胞增殖和凋亡的影响及其作用机制。方法用细胞形态学观察、DNA断裂百分率及DNA片段凝胶电泳法观察DBP对HL-60细胞增殖与凋亡的影响,用Fu-ra-2AM方法检测对HL-60细胞内游离钙离子浓度的影响,用免疫组织化学方法检测对细胞c-myc和bcl-2蛋白表达的影响。结果DBP可剂量和时间依赖性地抑制HL-60细胞增殖,并诱导其凋亡;引起HL-60细胞内钙重新分布,促进细胞外钙离子内流,升高细胞内游离钙([Ca2 ]i);下调白血病细胞c-myc和bcl-2蛋白表达。结论DBP可能通过提升HL-60白血病细胞内钙离子水平启动凋亡,且下调c-myc和bcl-2蛋白表达促进细胞凋亡,从而发挥其净化HL-60细胞的作用。     

β抑制蛋白2及微管相关蛋白轻链3在大鼠肾脏缺血再灌注损伤中的改变及意义

目的 探讨β抑制蛋白2(β-arrestin2)及微管相关蛋白轻链3(microtubule-associated protein light chain 3,LC3)在急性肾脏缺血再灌注损伤中的表达及与肾脏损害程度的相关性.方法 选用生后3~4周的雄性SD大鼠,随机分为正常组、假手术组、急性缺血再灌注损伤组.通过右侧肾脏切除,无创动脉夹夹闭左侧肾动脉45 min之后松开动脉夹,恢复肾脏血流,建立肾脏急性缺血再灌注损伤模型.并在恢复肾脏血流后12、24、36、48、72、96 h取肾脏及血液样本.采用免疫组织化学方法及West-ern blot方法检测各组肾组织中β-arrestin2及LC3蛋白的表达水平,检测各组的肾功能,并对各组肾脏病理学进行评分.结果 与正常组及假手术组相比,缺血再灌注损伤组血肌酐及肾脏病理学评分均有显著升高,其中肾脏损伤程度以缺血再灌注损伤后24 h最为明显;β-arrestin2及LC3蛋白在正常组及假手术组肾脏中的表达较少,在缺血再灌注损伤后的肾脏中表达显著升高,其中以缺血再灌注损伤后12 h时表达上调最为显著;β-arrestin2及LC3的表达改变先于肾脏病理改变,并且与肾脏损害程度呈正相关(r=0.821,P<0.05;r=0.913,P<0.05).结论 在肾脏急性缺血再灌注损伤时,β-arrestin2可能作为一个上游调控蛋白,通过对自噬的调节参与急性肾损伤的病理过程.     

先天性心脏病胎儿心脏及胎盘组织中MAPK及Akt信号通路分子的变化

目的：检测先天性心脏病(congenital heart disease,CHD)胎儿心脏及胎盘组织中MAPK及Akt信号通路分子表达变化,探讨其在CHD发病过程中的作用。方法：选取10例因患严重CHD/或同时伴其他缺陷而引产的胎儿,采用Western blot技术检测心脏左心室及胎盘组织中p38、p38α、p-p38、MEF2、ERK、p-ERK Akt、p-AktSer473、p-AktThr308蛋白表达量,采用半定量RT-PCR方法检测左心室p38α mRNA水平。7例孕龄相似、无CHD畸形儿为对照组。结果：①Western blot结果示：与对照组相比,CHD组中分别有4例心脏组织中p38及其亚型p38α、6例p-p38、2例MEF2、8例p-ERK、4例Akt及8例p-AktSer473和p-AktThr308蛋白水平降低,2例p-p38及1例p-AktThr308升高;CHD组中3例胎盘组织中p-p38上升,2例p-ERK下降。②CHD组MAPK及Akt信号通路蛋白表达量在胎儿心脏与胎盘组织中的变化不一致。③与对照组相比,4例p38α蛋白减少的心脏样本p38α异构体2 mRNA水平均降低,只有1例p38α异构体1、3、4 mRNA也减少。结论：CHD胎儿MAPK及Akt信号通路分子蛋白表达变化具有组织特异性,心脏组织中MAPK及Akt信号通路的异常可能与人类CHD的发生有关。［中国当代儿科杂志,2010,12（5）：327-332］     

MS275对发育期惊厥大鼠p38 MAPK信号通路的调控机制研究

目的 探讨组蛋白去乙酰化酶抑制剂MS275在发育期大鼠惊厥发病中对p38 MAPK信号通路的调控机制。方法 将32只雄性大鼠随机分为对照组、戊四唑（PTZ）组、低剂量MS275组（3?mg/kg）及高剂量MS275组（6?mg/kg）（n=8）。通过腹腔注射PTZ制作发育期大鼠惊厥模型,对照组仅注射生理盐水;MS275在PTZ诱导惊厥前2?h腹腔注射给药。造模成功后24?h,每组取6只大鼠,Western blot法检测海马组织p38、MK2、CREB和IL-6蛋白表达;qRT-PCR法检测p38、MK2、CREB和IL-6 mRNA表达;苏木精-伊红（HE）染色观察脑组织病理变化;Western blot法检测小胶质细胞活化标志物CD11b的表达。结果 PTZ组大鼠海马组织中p38、MK2、CREB和IL-6 mRNA及其蛋白的表达均较对照组显著增高（P < 0.05）;而MS275能抑制上述指标mRNA及其蛋白在发育期惊厥大鼠中的表达水平（P < 0.05）;6?mg/kg MS275对CREB mRNA及其蛋白、IL-6 mRNA的抑制作用比3?mg/kg MS275更显著（P < 0.05）。HE染色结果可见PTZ组海马组织有明显的神经元凋亡及细胞水肿,伴有较多的炎性细胞浸润;而MS275干预组能减轻发育期惊厥大鼠神经元凋亡及细胞水肿,同时减少炎性细胞浸润。PTZ组小胶质细胞活化明显增多,而MS275能抑制发育期惊厥大鼠小胶质细胞活化（P < 0.05）;且6?mg/kg MS275的抑制作用比3?mg/kg MS275更显著（P < 0.05）。结论 组蛋白去乙酰化酶抑制剂MS275能抑制发育期惊厥大鼠p38 MAPK信号通路、海马神经元的凋亡和小胶质细胞活化,从而减少炎症反应及惊厥性脑损伤;且MS275的作用可能存在剂量依赖性。