沉默信息调节因子1对胃癌侵袭的影响及其机理研究

目的 研究沉默信息调节因子1 （SIRT1）蛋白在胃癌组织中的表达及其对胃癌侵袭的影响和可能机理。方法 采用免疫组化SP法检测46例胃癌组织中SIRT1和血管内皮生长因子A （VEGF-A）蛋白的表达;采用Kaplan-Meier法分析SIRT1蛋白和VEGF-A蛋白表达对胃癌患者预后的影响;采用Western blot法检测GES-1人胃黏膜细胞株及SGC7901人胃癌细胞株中SIRT1蛋白和VEGF-A蛋白的表达;采用siRNA小干扰技术特异性干扰SGC7901细胞株中SIRT1基因的表达后,检测SIRT1蛋白和VEGF-A蛋白的表达;通过细胞侵袭实验检测不同处理后SGC7901细胞侵袭能力的变化。结果 与正常胃黏膜组织比较,胃癌组织中SIRT1蛋白和VEGF-A蛋白均呈高表达（P＜0.050）。SIRT1蛋白阳性表达患者的生存情况较差（P=0.001）,SIRT1及VEGF-A蛋白均呈阳性表达者的生存情况较差（P=0.006）,但VEGF-A蛋白表达与胃癌患者的预后无关（P=0.091）。SGC7901细胞中SIRT1蛋白（P=0.010） 和VEGF-A蛋白（P=0.020） 的表达较GES-1细胞上调。构建SIRT1-siRNA特异性抑制SIRT1基因的表达后（siRNA阳性组）,SIRT1蛋白和VGEF-A蛋白的表达均下调（P=0.010）,且SGC7901细胞的侵袭能力下降（P=0.000）。结论 SIRT1基因可能通过促进VEGF-A蛋白的表达而促进胃癌的侵袭,其可能是抑制胃癌侵袭的一个治疗靶点。     

Cystic fibrosis transmembrane conductance regulator is correlated closely with sperm progressive motility and normal morphology in healthy and fertile men with normal sperm parameters

Cystic fibrosis transmembrane conductance regulator (CFTR) has been demonstrated to be expressed in mature spermatozoa and correlated with sperm quality. Sperm CFTR expression in fertile men is higher than that in infertile men suffering from teratospermia, asthenoteratospermia, asthenospermia and oligospermia, but it is unknown whether CFTR is correlated with sperm parameters when sperm parameters are normal. In this study, 282 healthy and fertile men with normal semen parameters were classified into three age groups, group (I): age group of 20–29 years (98 cases, 27.1 ± 6.2), group (II): age group of 30–39 years (142 cases, 33.7 ± 2.6) and group (III): age group of more than or equal to 40 years (42 cases, 44.1 ± 4.6). Sperm concentration, total count and progressive motility were analysed by computer‐assisted sperm analysis. Sperm morphology was analysed by modified Papanicolaou staining. Sperm CFTR expression was conducted by indirect immunofluorescence staining. There was a significant positive correlation (P < 0.001) between CFTR expression and sperm progressive motility (r = 0.221) and normal morphology (r = 0.202), but there were no correlations between sperm CFTR expression and semen volume, sperm concentration, sperm total count as well as male age (P > 0.05). Our findings show that CFTR expression is associated with sperm progressive motility and normal morphology in healthy and fertile men with normal sperm parameters, but not associated with the number of spermatozoa and male age.     

Prolonged administration of secretin to normal rats increases biliary proliferation and secretin-induced ductal secretory activity

Background and aim

Cholangiocyte proliferation is coordinately regulated by a number of gastrointestinal hormones/peptides, some of which display stimulatory effects and some have inhibitory actions on cholangiocyte proliferation. Enhanced biliary proliferation [for example after bile duct ligation (BDL) and partial hepatectomy] is associated with increased expression of secretin receptor (SR), cystic fibrosis transmembrane conductance regulator (CFTR) and Cl^–/HCO₃^– anion exchanger 2 and secretin-stimulated ductal secretion, whereas loss/damage of bile ducts [for example after acute carbon tetrachloride (CCl₄) administration] is associated with reduced secretin-stimulated ductal secretory activity. There is growing information regarding the role of gastrointestinal hormones the regulation of biliary growth. For example, while gastrin, somatostatin and serotonin inhibit bile duct hyperplasia of cholestatic rats by downregulation of cAMP signaling, secretin has been shown to stimulate the proliferation of normal mice by activation of cyclic adenosine 3'',5''-monophosphate (cAMP)-dependent signaling. However, no information exists regarding the stimulatory effects of secretin on biliary proliferation of normal rats. Thus, we evaluated the in vivo and in vitro effect of secretin on biliary proliferation, the expression of markers key of ductal secretion and secretin-stimulated ductal secretion.

Methods

Normal male rats were treated with saline or secretin (2.5 nmoles/kg BW/day by osmotic minipumps for one week). We evaluated: (I) intrahepatic bile duct mass (IBDM) in liver sections and PCNA expression in purified cholangiocytes; (II) SR and CFTR mRNA expression and secretin-stimulated cAMP levels in purified cholangiocytes; and (III) secretin-stimulated bile and bicarbonate secretion in bile fistula rats. In vitro, normal rat intrahepatic cholangiocyte lines (NRIC) were treated with BSA (basal) or secretin (100 nM) for 24 to 72 hours in the absence/presence of a PKA or a MEK inhibitor before evaluating proliferation by MTS assays.

Results

Prolonged administration of secretin to normal rats increased IBDM and PCNA expression in purified cholangiocytes compared to saline-treated normal rats. Also, secretin increased the expression of proteins (SR and CFTR) that are key in the regulating ductal secretion and enhanced secretin-stimulated cAMP levels and bile and bicarbonate secretion. In vitro, secretin increased the proliferation of NRIC, increase that was prevented by PKA and MAPK inhibitors.

Conclusions

We have demonstrated that secretin stimulates both in vivo and in vitro biliary proliferation and secretin-stimulated ductal secretory activity in normal rats. We suggest that the stimulatory effect of secretin on biliary proliferation and secretion may be important for preventing biliary dysfunction during ductopenic disorders.     

CFTR Protein Repair Therapy in Cystic Fibrosis

Cystic fibrosis is a single gene, autosomal recessive disorder, in which more than 1900 mutations grouped into 6 classes have been described. It is an example a disease that could be well placed to benefit from personalized medicine. There are currently 2 very different approaches that aim to correct the basic defect: gene therapy, aimed at correcting the genetic alteration, and therapy aimed at correcting the defect in the CFTR protein. The latter is beginning to show promising results, with several molecules under development. Ataluren (PTC124) is a molecule designed to make the ribosomes become less sensitive to the premature stop codons responsible for class i mutations. Lumacaftor (VX-809) is a CFTR corrector directed at class ii mutations, among which Phe508del is the most frequent, with encouraging results. Ivacaftor (VX-770) is a potentiator, the only one marketed to date, which has shown good efficacy for the class iii mutation Gly551Asp in children over the age of 6 and adults. These drugs, or a combination of them, are currently undergoing various clinical trials for other less common genetic mutations. In the last 5 years, CFTR has been designated as a therapeutic target. Ivacaftor is the first drug to treat the basic defect in cystic fibrosis, but only provides a response in a small number of patients. New drugs capable of restoring the CFTR protein damaged by the most common mutations are required.     

尼克酰胺磷酸核糖转移酶对心肌缺血/再灌注的影响研究进展

背景 尼克酰胺磷酸核糖转移酶(nicotinamide phosphoribosyltransferase,NAMPT)是合成烟酰胺腺嘌呤二核苷酸(nicotinamide adenine dinucleotide,NAD)的限速酶.NAMPT可存在于细胞内和细胞外,它们的作用不尽相同.NAMPT在心血管系统中发挥着重要作用,但在心肌缺血/再灌注(ischemia/reperfusion,I/R)中的作用尚存争议. 目的 进一步了解NAMPT的心血管作用及对心肌I/R的影响. 内容 就目前研究对NAMPT的来源及潜在的心血管作用、细胞内NAMPT(intracellular NAMPT,iNAMPT)及细胞外NAMPT(extracellular NAMPT,eNAMPT)、以及对心肌I/R的影响作一综述. 趋向 促进对NAMPT的作用及机制的了解,NAMPT有望成为治疗心肌I/R损伤的新靶点.     

沉默信息调节蛋白1在缺血性脑损伤中的神经保护作用

近年来,沉默信息调节蛋白1（silent information regulator protein 1,SIRT1）作为依赖NAD+的组蛋白去乙酰化酶被广泛地研究。SIRT1存在于机体各组织细胞中,通过去乙酰化修饰,调节细胞的多种生理过程,在能量守恒、细胞氧化、衰老及凋亡等方面发挥重要作用。目前认为,SIRT1在缺血性脑损伤中起到神经保护作用,可能通过调控细胞能量代谢,抗炎性反应、抗细胞凋亡等方面发挥作用。本文主要综述了SIRT1在缺血性脑损伤中的神经保护作用。     

Beta2-glycoprotein I, anti-beta2-glycoprotein I, and fibrinolysis

β₂-glycoprotein-I (β₂GPI) is a phospholipid-binding plasma protein that consists of five homologous domains. Domain V is distinguished from others by bearing a positively charged lysine cluster and hydrophobic extra C-terminal loop. β₂GPI has been known as a natural anticoagulant regulator. β₂GPI exerts anticoagulant activity by inhibition of phospholipid-dependent coagulation reactions such as prothrombinase, tenase, and factor XII activation. It also binds factor XI and inhibits its activation. On the other hand, β₂GPI inhibits anticoagulant activity of activated protein C. According to the data from knockout mice, β₂GPI may contribute to thrombin generation in vivo. Phospholipid-bound β₂GPI is one of the major target antigens for antiphospholipid antibodies present in patients with antiphospholipid syndrome (APS). Binding of pathogenic anti-β₂GPI antibodies increases the affinity of β₂GPI to the cell surface and disrupts the coagulation/fibrinolysis balance on the cell surface. These pathogenic antibodies activate endothelial cells via signal transduction events in the presence of β₂GPI. Impaired fibrinolysis has been reported in patients with APS. Using a newly developed chromogenic assay, we demonstrated lower activity of intrinsic fibrinolysis in euglobulin fractions from APS patients. Addition of monoclonal anti-β₂GPI antibodies with β₂GPI also decreased fibrinolytic activity in this assay system. β₂GPI is proteolytically cleaved by plasmin in domain V (nicked β₂GPI) and becomes unable to bind to phospholipids, reducing antigenicity against antiphospholipid antibodies. This cleavage occurs in patients with increased fibrinolysis turnover. Nicked β₂GPI binds to plasminogen and suppresses plasmin generation in the presence of fibrin, plasminogen, and tissue plasminogen activator (tPA). Thus, nicked β₂GPI plays a role in the extrinsic fibrinolysis via a negative feedback pathway loop.     

β2-glycoprotein I, anti-β2-glycoprotein I, and fibrinolysis

β₂-glycoprotein-I (β₂GPI) is a phospholipid-binding plasma protein that consists of five homologous domains. Domain V is distinguished from others by bearing a positively charged lysine cluster and hydrophobic extra C-terminal loop. β₂GPI has been known as a natural anticoagulant regulator. β₂GPI exerts anticoagulant activity by inhibition of phospholipid-dependent coagulation reactions such as prothrombinase, tenase, and factor XII activation. It also binds factor XI and inhibits its activation. On the other hand, β₂GPI inhibits anticoagulant activity of activated protein C. According to the data from knockout mice, β₂GPI may contribute to thrombin generation in vivo. Phospholipid-bound β₂GPI is one of the major target antigens for antiphospholipid antibodies present in patients with antiphospholipid syndrome (APS). Binding of pathogenic anti-β₂GPI antibodies increases the affinity of β₂GPI to the cell surface and disrupts the coagulation/fibrinolysis balance on the cell surface. These pathogenic antibodies activate endothelial cells via signal transduction events in the presence of β₂GPI. Impaired fibrinolysis has been reported in patients with APS. Using a newly developed chromogenic assay, we demonstrated lower activity of intrinsic fibrinolysis in euglobulin fractions from APS patients. Addition of monoclonal anti-β₂GPI antibodies with β₂GPI also decreased fibrinolytic activity in this assay system. β₂GPI is proteolytically cleaved by plasmin in domain V (nicked β₂GPI) and becomes unable to bind to phospholipids, reducing antigenicity against antiphospholipid antibodies. This cleavage occurs in patients with increased fibrinolysis turnover. Nicked β₂GPI binds to plasminogen and suppresses plasmin generation in the presence of fibrin, plasminogen, and tissue plasminogen activator (tPA). Thus, nicked β₂GPI plays a role in the extrinsic fibrinolysis via a negative feedback pathway loop.     

Loss of vancomycin bactericidal activity against accessory gene regulator (agr) dysfunctional Staphylococcus aureus under conditions of high bacterial density

The impact of accessory gene regulator (agr) dysfunction and high bacterial density on vancomycin killing and resistance was evaluated among 10 clinical methicillin-resistant Staphylococcus aureus bloodstream isolates using time kill experiments. Under conditions of high inocula and agr dysfunction, vancomycin activity was markedly attenuated, amplifying resistant mutants by 72 h.