去甲肾上腺素对人肝细胞系表达瘦素及瘦素受体的影响

目的探讨交感神经递质去甲肾上腺素(NE)对体外培养人肝细胞系L02表达瘦素及瘦素受体的影响。方法用不同浓度的NE作用于体外培养的人肝细胞系L02,分别于24、48及72 h收集细胞,Western印迹法检测其表达瘦素及瘦素受体蛋白的影响;RT-PCR检测NE对肝细胞系L02瘦素及瘦素受体mRNA的影响。结果①Western印迹结果显示,1、10、100μmol/L NE作用于L02细胞24 h后,瘦素蛋白表达明显高于对照组(1.02±0.08,2.24±0.09,2.35±0.12 vs 0.62±0.09,P<0.05);瘦素受体蛋白表达亦明显高于对照组(1.35±0.13,2.37±0.12,2.39±0.15 vs 0.85±0.13,P<0.05)。②RT-PCR检测L02瘦素以及瘦素受体mRNA的表达情况,1、10、100μmol/L NE作用于L02 24 h后,瘦素mRNA表达明显高于对照组(1.54±0.08,2.37±0.09,2.72±0.12 vs 1.00±0.07,P<0.05);瘦素受体mRNA表达亦明显高于对照组(1.61±0.08,2.27±0.10,3.39±0.11 vs1.00±0.06,P<0.01)。结论交感神经递质NE对体外培养的肝细胞系瘦素以及瘦素受体的表达均有促进作用,从而参与了肝纤维化进程。     

糖原合成酶基因多态性与高血压的关系

目的研究糖原合成酶基因多态性与原发性高血压易感性的关系。方法采用内切酶Ｘｂａ１对８２例原发性高血压患者和７２例正常血压对照者的糖原合成酶基因片段ＰＣＲ产物进行酶解。结果原发性高血压患者的Ａ２等位基因频率显著高于正常血压者（Ｐ＝０．０２５）。结论糖原合成酶基因Ｘｂａ１多态性可能作为原发性高血压发病的一种标志。     

瘦素和瘦素受体mRNA表达与哮喘和肥胖的关系

目的探讨瘦素(LEP)和瘦素受体(LEPR)mRNA表达与哮喘和肥胖的关系。方法采用实时荧光定量PCR(RT-qPCR)方法测定各组PBMC LEP和LEPR mRNA表达水平,分析LEP mRNA与血浆瘦素浓度、EOS%、IgE和FEV1占预计值(%)的关系。结果哮喘、肥胖、哮喘合并肥胖组的PBMC LEP mRNA表达量与健康对照组相比明显升高(P均0.01),中、重度哮喘与轻度哮喘相比明显升高(P均0.05);肥胖组LEPR mRNA表达量与其余各组相比明显升高(P均0.01);哮喘组PBMC LEP mRNA表达量与血浆LEP浓度呈正相关(r=0.211,P=0.012),与FEV1占预计值(%)成负相关(r=-0.314,P=0.043),与EOS%、IgE无相关关系(r=0.002、0.001,P=0.58、0.96)。结论 LEP mRNA表达升高可能参与哮喘和肥胖的发生,并且导致较为严重的哮喘表型,其机制可能与血浆瘦素浓度增高有关,其相关的哮喘是非嗜酸细胞性、非过敏性的;LEPR mRNA高表达参与肥胖发生,但与哮喘无关。     

瘦素对肝细胞葡萄糖氧化及葡萄糖激酶基因表达的影响

目的观察瘦素（Ｌｅｐｔｉｎ）对肝细胞葡萄糖氧化的影响及其剂量效应,并观察其对葡萄糖激酶基因表达的影响。方法正常大鼠肝细胞,以不加Ｌｅｐｔｉｎ与加不同剂量的Ｌｅｐｔｉｎ培养１小时后,以液体闪烁计数法检测肝细胞氧化〔Ｕ１４Ｃ〕葡萄糖的量。以ＲＴＰＣＲ法检测Ｌｅｐｔｉｎ１００μｇ／Ｌ处理组与对照组肝细胞葡萄糖激酶（ＧＫ）ｍＲＮＡ的表达。结果当Ｌｅｐｔｉｎ浓度为１０μｇ／Ｌ时,肝细胞葡萄糖氧化与对照组无明显差异,当Ｌｅｐｔｉｎ浓度上升为５０μｇ／Ｌ后,葡萄糖氧化较对照组显著下降（Ｐ＜０．０５）,并随剂量增大抑制作用加强。Ｌｅｐｔｉｎ１００μｇ／Ｌ处理组ＧＫｍＲＮＡ表达水平（以βａｃｔｉｎ校正）亦显著低于对照组（０．４３±０．０７比０．８７±０．０９,Ｐ＜０．０１）。结论低剂量瘦素对肝细胞葡萄糖氧化无明显影响,高浓度瘦素则具剂量依赖性抑制作用,并至少部分通过抑制ＧＫ基因表达而达到     

糖原合成酶激酶-3β对巨噬细胞系RAW264.7活化的影响

目的探讨糖原合成酶激酶-3β（GSK-3β）对巨噬细胞系RAW264.7活化的影响。方法将生长状态良好的RAW264.7细胞分为3组：实验对照组、脂多糖（LPS）组和GSK-3β特异抑制剂SB216763干预组,在12 h、24 h进行指标检测。采用Western印迹法检测细胞GSK-3β、p-GSK-3β^ser9蛋白的表达,ELISA试剂盒法检测细胞上清IL-10、TNF-α的变化,RT-PCR检测细胞中5-LO mRNA变化,免疫荧光法检测ED1表达,电镜下观察RAW264.7细胞形态变化。结果12 h和24 h LPS组与对照组相比,p-GSK-3β^ser9表达减少,GSK-3β表达不变,活性升高;TNF-α、IL-10及5-LO mRNA表达增多（P〈0.05）;ED1表达绿色荧光明显增多;透射电镜观察LPS组巨噬细胞形态不规则,吞噬坏死物质细胞变多。12 h和24 h SB216763干预组与LPS组进行比较,p-GSK-3β^ser9表达明显增多,GSK-3β表达不变,活性降低;TNF-α及5-LO mRNA表达减少（P〈0.05）,IL-10表达明显增多（P〈0.05）;ED1绿色荧光表达减少;透射电镜观察细胞形态较规则。结论 GSK-3β对于RAW264.7细胞活化发挥重要的作用。     

瘦素对培养大鼠原代肝细胞葡萄糖吸收及肝细胞膜胰岛素受体磷酸化的影响

目的：观察瘦素对培养大鼠原代肝细胞葡萄糖吸收及肝细胞膜胰岛素受体磷酸化的影响,探讨瘦素参与胰岛素抵抗的分子生物学机制。方法：体外培养大鼠肝细胞,根据培养基中加入瘦素的浓度将实验分为空白对照组以及50、100、200、500／μg／L瘦素组,共5组,每组分别培养2h和24h,用全自动生化分析仪检测培养基中剩余的葡萄糖浓度,用ELISA方法测定肝细胞膜磷酸化胰岛素受体β亚基（pY 1158）含量。结果：①培养基剩余葡萄糖检测,瘦素抑制肝细胞对葡萄糖的吸收,且抑制效应随瘦素浓度增加和时间延长而增强（P〈0.01）;②肝细胞膜磷酸化胰岛素受体β亚基（pY1158）水平测定,50／μg／L瘦素组与对照组比较差异无统计学意义（P〉0．05）,100、200、500／2μg／L瘦素组与对照组比较差异有极显著性（P〈0．01）,并随瘦素剂量上升含量进一步下降。结论：瘦素与胰岛素抵抗相关,抑制细胞膜胰岛素受体β亚基（pY1158）磷酸化可能是其参与胰岛素抵抗的机制之一。     

原发性肝细胞癌患者组织中瘦素及其受体表达与临床病理特征的关系

目的检测瘦素（leptin）和瘦素受体（OB—R）在原发性肝细胞癌（HCC）患者癌及癌旁组织中mRNA和蛋自水平表达情况。方法应用RT-PCR和Westernblot方法检测leptin其受体在33例HCC患者癌及癌旁组织中mRNA和蛋白表达,并分析其与临床病理特征之间的关系。结果leptin和OB—RmRNA及蛋白在癌旁组织表达水平高于癌组织（P〈0．05）。leptin和OB—R的表达与各临床病理特征均无显著相关（P〉0．05）。结论leptin及OB-R在HCC患者的癌及癌旁组织中均表达。     

假酸浆水提液对2型糖尿病模型大鼠肝糖原合成关键酶表达的影响

目的研究假酸浆(NPG)水提液对2型糖尿病(T2DM)模型大鼠肝糖原合成关键酶表达的影响。方法用50只Wistar大鼠作为受试对象制作糖尿病模型。一次性腹腔注射链脲佐菌素(STZ)制备2型糖尿病大鼠模型。以NPG水提液进行灌胃,测定血糖及肝脏糖原合成酶、葡萄糖-6-磷酸酶的表达水平。结果 NPG能有效地降低血糖。NPG水提液低剂量组(2.5 ml/kg)能增加大鼠肝脏组织糖原合成酶mRNA的表达(P<0.05),降低大鼠肝脏葡萄糖-6-磷酸酶mRNA的表达(P<0.01)。结论 NPG是一种有效的降血糖药,可以加强糖原合成,进而降低血糖。     

Epidermal growth factor stimulates glycogen synthase activity in cultured cells.

Addition of epidermal growth factor (EGF) to quiescent cultured cells was found to stimulate the activity of glycogen synthase (UDPglucose:glycogen 4-alpha-D-glucosyltransferase, EC 2.4.1.11), an enzyme subjected to regulation by covalent modification. In Swiss mouse 3T3 cells, the activation by EGF paralleled the effect seen with insulin; the time course and dose-response curves of the two polypeptide factors were similar. Stimulation of enzyme activity ratio [(activity in the absence of glucose 6-phosphate)/(activity in the presence of glucose 6-phosphate)] was maximal after 20-30 min of incubation. Both factors caused a maximal stimulation of 2.5-fold in synthase activity ratio at approximately equal to 10 nM, and the half-maximal effect was observed at 0.1-1 nM. Insulin and EGF exhibited partial additivity in effecting this enzyme activation. In contrast, human A431 cells showed no response to insulin. Although quantitatively different, the EGF effect in the latter cells was time dependent, reaching a maximum at 90 min, and dose dependent, with a maximal stimulation of 4-fold in synthase activity ratio at 10 nM. Half-maximal effect was observed at 0.3 nM EGF. Direct quantitation of allosteric effectors (glucose 6-phosphate, adenine nucleotides, and Pi) present in the enzyme assay mixtures indicated that the observed activation was not simply a consequence of changes in metabolite concentrations. These results suggest that EGF may be important in regulating glycogen synthesis through phosphorylation/dephosphorylation mechanisms.     

注射用血栓通对肝星状细胞基因表达的影响

目的:探讨注射用血栓通对肝星状细胞 LX02基因表达的影响及其机制.方法:将103 mg/L的注射用血栓通与人肝星状细胞 LX02 共同孵育 48 h,提取 mRNA,并逆转录成cDNA.与芯片杂交,根据杂交信号强弱筛选相关基因.结果:41 条肝星状细胞基因差异表达,其中表达上调的基因21条,下调基因20条,这些基因主要与能量代谢、细胞黏附、DNA结合、磷酸化、甲基化等相关,还有部分基因功能不详.结论:经注射用血栓通作用后,肝星状细胞基因表达发生变化,其可能通过调控这些基因的表达实现了对肝星状细胞功能的抑制作用.     

Fat feeding impairs glycogen synthase activity in mice without effects on its gene expression

To examine whether the effects of high-fat feeding on glycogen synthase (GS) activity and mRNA levels differ between diabetes-prone (C57BL/6J) and diabetes-resistant mice (NMRI), we measured GS activity and mRNA levels in muscle from C57BL/6J and NMRI mice fed a high-fat or normal chow diet for 3, 6, and 15 months. As compared with chow feeding, fat feeding increased plasma insulin levels in C57BL/6J mice at 15 months (464 +/- 29 v 267 +/- 47 pmol/L, P =.005), which was associated with elevated plasma glucose levels at 15 months (5.3 +/- 0.3 v 3.8 +/- 0.2 mmol/L, P =.001). Fat feeding increased plasma insulin levels also in NMRI mice at 15 months (705 +/- 145 v 275 +/- 64 pmol/L, P =.01) without, however, a rise of plasma glucose levels. In parallel with increased insulin levels, decreased muscle GS fractional velocity (FV) was observed at 6 (49.0% +/- 2.6% v 69.1% +/- 7.3%, P =.04) and 15 (45.8% +/- 1.8% v 53.4% +/- 1.6 %, P <.01) months but not at 3 months in the fat-fed C57BL/6J mice. Similarly, there was a significant decrease in GS fractional activity at 3 (57.9% +/- 4.3% v 70.4% +/- 2.6 %, P <.03) and 15 (47.3% +/- 2.4% v 56.4% +/- 2.1%, P =.02) but not at 6 months in the fat-fed NMRI mice. The decrease in GS activity was not associated with changes in mRNA levels at any time points. We conclude that (1) fat feeding results in similar elevation of plasma insulin levels and impairs GS activity in C57BL/6J and NMRI mice, and (2) the changes in GS activity do not involve effects on gene expression.     

JAZF1基因过表达对鼠肝细胞糖脂代谢相关基因表达的影响

目的探讨JAZF1过表达对鼠肝细胞IAR-20及Hepa1-6糖脂代谢相关基因的影响。方法克隆鼠JAZF1编码区并构建pIRES2-EGFP-JAZF1表达载体,利用脂质体法将目的基因转染到鼠肝细胞IAR-20及Hepa1-6中,采用实时荧光定量PCR测定各处理组JAZF1 mRNA表达情况,以及糖脂代谢相关基因SREBP1、AOC、FAS、HSL、PPARa、ATGL、G1uT-1、G1uT-4和细胞因子FGF-21mRNA的变化情况。结果JAZFl基因在鼠肝细胞中过表达导致脂代谢基因SREBP1,ACC,FAS表达降低（P〈0．01）,PPARa和HSL表达增加（P〈0．01）,对AXG-L无影响（P〉0．05）,使糖转运相关基因GluT-1表达增加（P〈0．01）,GluT-4无显著变化（P〉0．05）,对FGF-21无影响（P〉0．05）。结论JAZF1过表达可以抑制肝细胞的脂肪生成,促进脂肪分解,并可能增加基础葡萄糖转运。     

Effects of Fuzhenghuayu decoction on collagen synthesis of cultured hepatic stellate cells, hepatocytes and fibroblasts in rats

ＩＮＴＲＯＤＵＣＴＩＯＮＬｉｖｅｒｆｉｂｒｏｓｉｓｉｓｔｈｅｃｏｍｍｏｎｐａｔｈｏｌｏｇｉｃａｌｆｅａｔｕｒｅｏｆｃｈｒｏｎｉｃｌｉｖｅｒｄｉｓｅａｓｅｓ,ａｎｄｉｓｃｌｏｓｅｌｙａｓｏｃｉａｔｅｄｗｉｔｈｃｈａｎｇｅｓｏｆｌｉｖｅｒｃｅｌｆｕｎｃｔ...     

α干扰素对肝星状细胞凋亡及其基因表达的影响

临床研究发现,α干扰素(IFN-α)有特异的抗肝纤维化作用而不依赖于其抗病毒作用,但其抗纤维化机制尚不清楚。目前IFN-α对活化的肝星状细胞(HSC)凋亡的影响未见报道。以体外培养的鼠HSC为研究对象,观察IFN-α对活化状态的HSC凋亡及其相关基因表达的影响     

Effects of long-term restricted feeding on plasma leptin, hepatic leptin expression and leptin receptor expression in juvenile Atlantic salmon (Salmo salar L.)

Leptin is a pleiotropic hormone and plays a key role in body weight regulation, energy homeostasis and lipid store utilization in mammals. In this study, we investigated the effect of feed-restriction on leptin genes (lepa1 and lepa2), leptin receptor (lepr) gene expression and plasma leptin levels in juvenile Atlantic salmon parr. Feed restriction was performed from late April to mid-June, in order to gain insight into the role of the leptin system in energy balance regulation and adiposity in juvenile salmon. A significant increase in lepa1 expression as well as higher levels of plasma leptin was found in feed-restricted fish in June compared to fully fed controls, while lepa2 gene expression decreased in both groups during the treatment period. Lepa2 was, however significantly higher in the feed-restricted group in June. Leptin receptor expression was up regulated during the period of enhanced growth and lipid deposition in the fully fed control, indicating a seasonal effect on the receptor expression in the brain. Both lepa1 and lepa2 genes very mainly expressed in the liver in juvenile salmon, while lepr was expressed in the brain but showed also considerable expression in various peripheral tissues. The study provides evidence that the leptin system is sensitive to the metabolic status of the fish as both season and restricted feeding affect lepa1 and lepa2 gene expression in the liver and brain leptin receptor expression, however, for lepa1 expression and leptin plasma level in an opposite way as that observed in the mammalian system.