Interleukin-4 Restores Insulin Sensitivity in Lipid-Induced Insulin-Resistant Adipocytes

Obesity and latent inflammation in adipose tissue significantly contribute to the development of insulin resistance (IR) and type 2 diabetes. Here we studied whether the antiinflammatory interleukin-4 (IL-4) can restore insulin sensitivity in cultured 3T3-L1 adipocytes. The activity of key components of the insulin signaling cascade was assessed by immunoblotting using phospho-specific antibodies to insulin receptor substrate IRS1 (Tyr612), Akt (Thr308 and Ser473), and AS160 (Ser318) protein that regulates translocation of the GLUT4 glucose transporter to the plasma membrane. IR was induced in mature adipocytes with albumin-conjugated palmitate. IR significantly reduced phosphorylation levels of all the above-mentioned proteins. Addition of IL-4 to the culturing medium during IR induction led to a dose-dependent stimulation of the insulin-promoted phosphorylation of IRS1, Akt, and AS160. At the optimal concentration of 50 ng/ml, IL-4 fully restored activation of the insulin cascade in IR cells, but it did not affect insulin signaling activation in the control cells. IL- 4 neither upregulated expression of key adipogenesis markers GLUT4 and PPARγ nor caused lipid accumulation in the adipocytes. These results demonstrate that IL-4 can restore insulin sensitivity in adipocytes via mechanisms not associated with induced adipogenesis or de novo formation of lipid depots.     

Arrhythmia and sudden death associated with elevated cardiac chloride channel activity

The identification and analysis of several cationic ion channels and their associated genes have greatly improved our understanding of the molecular and cellular mechanisms of cardiac arrhythmia. Our objective in this study was to examine the involvement of anionic ion channels in cardiac arrhythmia. We used a transgenic mouse model to overexpress the human cystic fibrosis transmembrane conductance regulator (CFTR) gene, which encodes a cAMP-regulated chloride channel. We used RNase protection and in situ hybridization assays to determine the level of CFTR expression, and radiotelemetry and in vivo electrophysiological study in combination with pharmacological intervention to analyse the cardiac function. Cardiac CFTR overexpression leads to stress-related sudden death in this model. In vivo intracardiac electrophysiological studies performed in anaesthetized mice showed no significant differences in baseline conduction parameters including atrial-His bundle (AH) or His bundle-ventricular (HV) conduction intervals, atrioventricular (AV) Wenckebach or 2:1 AV block cycle length and AV nodal functional refractory period. However, following isoproterenol administration, there was marked slowing of conduction parameters, including high-grade AV block in transgenic mice, with non-sustained ventricular tachycardia easily inducible using programmed stimulation or burst pacing. Our sudden death mouse model can be a valuable tool for investigation of the role of chloride channels in arrhythmogenesis and, potentially, for future evaluation of novel anti-arrhythmic therapeutic strategies and pharmacological agents.     

Crystallization and preliminary X-ray crystallographic analysis of spruce budworm antifreeze protein.

Antifreeze proteins have the ability to bind to ice with high affinity and inhibit further crystal growth. The insect antifreeze protein from spruce budworm exhibits very high thermal hysteresis activity and is implicated in the protection of overwintering larvae from freezing. This protein has been crystallized in 20-25% polyethylene glycol (Mr 6000), 0.4 M NaCl, 0.1 M Tris-HCl, pH 8.5, by vapor diffusion using the hanging drop method. The resulting crystals are very thin (typically <0.01 mm in the shortest dimension), and only after repeated seeding could crystals be grown large enough for data collection using synchrotron radiation. The crystals belong to the monoclinic space group C2, with cell dimensions a = 82.28 A, b = 62.29 A, c = 63.63 A, and beta = 113.7 degrees. Molecules in the asymmetric unit are related by a twofold axis of symmetry with two molecules present. Native data to a resolution of 2.6 A have been collected with 90.3% completeness and a Rsym of 6.9%.     

Efficient mucosal vaccination mediated by the neonatal Fc receptor

Almost all infectious diseases are initiated at mucosal surfaces, yet intramuscular or subcutaneous vaccination usually provides only minimal protection at sites of infection owing to suboptimal activation of the mucosal immune system. The neonatal Fc receptor (FcRn) mediates the transport of IgG across polarized epithelial cells lining mucosal surfaces. We mimicked this process by fusing a model antigen, herpes simplex virus type-2 (HSV-2) glycoprotein gD, to an IgG Fc fragment. Intranasal immunization, together with the adjuvant CpG, completely protected wild-type, but not FcRn knockout, mice after intravaginal challenge with virulent HSV-2 186. This immunization strategy induced efficient mucosal and systemic antibody, B- and T-cell immune responses, with stable protection for at least 6 months after vaccination in most of the immunized animals. The FcRn-IgG transcellular transport pathway may provide a general delivery route for subunit vaccines against many mucosal pathogens.     

大型绿藻浒苔转化表达系统选择标记的筛选

主要研究了条浒苔对抗生素氯霉素和除草剂Basta的敏感性,以确定适合的阳性选择标记基因。应用不同浓度氯霉素(0、25、50、75、100、125μg/ml)和Basta(0、5、12.5、25、37.5、50μg/ml)对不同发育时期条浒苔细胞存活率影响进行了测定。实验结果表明:不同发育时期条浒苔对氯霉素和Basta的敏感性不同。其中最大浓度125μg/ml浓度的氯霉素在15d内对条浒苔孢子和小苗两个不同发育时期的细胞均难以达到全部杀死效果,相对存活率仍分别为1%和20%;而Basta对条浒苔孢子和小苗均具有很强的杀生作用,其中5μg/ml浓度的Basta在3d内可将条浒苔孢子全部杀死,12.5μg/ml浓度下约一周时间可以将浒苔小苗全部致死。本实验结果提示bar基因有可能成为浒苔基因工程较理想的选择标记基因。     

SpltMNPV日本分离株gp41的克隆表达及gp41和ph的进化分析

从斜纹夜蛾核型多角体病毒(Spodopteralitura multicapsid nucleopolyhedrovirus,SpltMNPV)日本分离株(C3)基因组中克隆了gp41基因.该基因编码区含993bp核苷酸,编码分子量为36.9kDa的多肽.将该基因克隆至原核表达载体pET28a,经IPTG诱导后在大肠杆菌BL21(DE3)中获得了表达.应用CLUSTAL程序分析表明,SpltMNPV日本株(C3)gp41的核苷酸序列和氨基酸序列与SpltMNPV中国G2株相似性最高,均达99.9%.用MEGA分别构建了基于gp41和ph的聚类分析图和分子进化树,发现它们具有相似的拓扑结构.将这两个基因序列结合在一起构建进化树,该树的结构与基于gp41的进化树相似.突变率分析显示gp41的突变率高于ph,这意味着在杆状病毒进化过程中,gp41和ph面临不同的选择压力.     

Precise Reconstruction of Veins and Bile Ducts in Rat Liver Transplantation

Rat orthotopic liver transplantation (ROLT) remains a technically demanding procedure, especially regarding the reconstruction of the suprahepatic vena cava (SHVC). In this study, a new microsuture technique was developed for anastomosis of the SHVC, and a special single-groove cuff and blade-cut stent were introduced. With these modified techniques, we aimed to make a precise anastomosis of the SHVC and to provide optimal cuffs and stents for the reconstruction of the veins and bile ducts. According to different microsuture techniques for the SHVC and different types of cuffs and stents, three ROLT groups were created to compare the operation times and prognoses. Sham operations were performed as controls in the fourth group. The time expenditures with each step were compared among the transplantation groups. Biochemical parameters were tested at the end of a 1-month observation period. The short- and long-term survival rates of the transplantation groups were recorded and compared. Our new microsuture technique was faster than the conventional continuous suture technique for SHVC anastomosis (P < 0.05). The use of a single-groove cuff for reconstruction of the portal vein and the infrahepatic vena cava shortened the anastomotic time (P < 0.05). The use of blade-cut stents resulted in fewer biliary complications and better survival over the short and long terms (P < 0.05). Our new microsuture technique and the single-groove cuffs proved to be a precise method for venous reconstruction which shortened the anhepatic time and the anastomotic time significantly. The blade-cut stents apparently reduced the incidence of biliary complications. In summary, with this precise microsuture technique and delicate cuffs and stents, excellent long-term survival can be achieved easily and stably for ROLT.     

颈内动脉支架置入术对颈动脉狭窄患者认知功能影响的研究

目的：研究颈动脉狭窄对认知功能的影响,探讨颈动脉支架置入术（CAS）患者术后不同时期认知功能的变化及其可能的机制。方法：选取2010年1月至2012年12月我院神经内科收住的急性脑梗死（前循环）患者75例,按颈内动脉狭窄程度,分成无狭窄组,轻度狭窄组（狭窄程度〈30％）,中度狭窄组（30-69％）和治疗组（70．99％）,前三组给予药物治疗,治疗组同时给予颈内动脉支架置入术,应用蒙特利尔认知评估量表（MoCA）、简易精神状态检查表（MMSE）、搭火柴测验（stickTest）,对患者进行认知功能评估,比较术前、术后不同时期认知功能的变化,同时利用SPECT／CT对治疗组20例患者术后局部脑血流改善状况进行评价。结果：治疗组和对照组在术前比较MoCA总分、MMSE评分、StickTest评分,发现治疗组评分低于对照组,且随着颈动脉狭窄程度的加重,认知功能受损越明显;治疗组在术后1周与术前比较,评分反而降低,差异有统计学意义;术后3个月、术后6个月与术前比较,评分均有所提高,差异有统计学意义;通过SPECT／CT对治疗组术前感兴趣区血流量与同侧小脑平均脑血流量比较,术后再与同侧小脑比较,结果显示术后脑灌注明显改善。结论：颈动脉狭窄与血管性认知功能障碍有关,而且认知功能障碍的程度与颈动脉狭窄程度呈正相关,颈内动脉支架置入术可最终改善认知功能,尤其表现在视空间／执行能力,延迟回忆能力,注意力等方面。     

亚抑菌浓度头孢西丁对耐药质粒接合转移的影响

目的探讨亚抑菌浓度头孢西丁对耐药质粒接合转移的影响,研究病原菌耐药性播散的产生机制。方法采用聚合酶链反应（PCR）分析临床分离的63株产ESBLs肺炎克雷伯菌株SHV型β-内酰胺酶编码基因。质粒接合转移试验采用肉汤接合法。研究不同亚抑菌浓度头孢西丁（0、1、0.5、0.25、0.125μg/ml）和不同作用时间（2、4、6、8、10、12 h）下临床产ESBLs肺炎克雷伯菌株供体菌与受体菌E.coliC600接合转移频率的变化。结果63株临床分离的产ESBLs肺炎克雷伯菌株有41株扩增出SHV型基因,阳性率为65.08%。随着亚抑菌浓度头孢西丁作用时间的增加,接合转移频率有随之增加的趋势。在相同作用时间下,头孢西丁浓度0.125μg/ml作用下的接合转移频率高于其他亚抑菌浓度的作用。结论应合理使用抗菌药物,减少抗菌药物的选择性压力,防止耐药细菌传播。