Nitrosothiol Formation and Protection against Fenton Chemistry by Nitric Oxide-induced Dinitrosyliron Complex Formation from Anoxia-initiated Cellular Chelatable Iron Increase

Dinitrosyliron complexes (DNIC) have been found in a variety of pathological settings associated with ^•NO. However, the iron source of cellular DNIC is unknown. Previous studies on this question using prolonged ^•NO exposure could be misleading due to the movement of intracellular iron among different sources. We here report that brief ^•NO exposure results in only barely detectable DNIC, but levels increase dramatically after 1–2 h of anoxia. This increase is similar quantitatively and temporally with increases in the chelatable iron, and brief ^•NO treatment prevents detection of this anoxia-induced increased chelatable iron by deferoxamine. DNIC formation is so rapid that it is limited by the availability of ^•NO and chelatable iron. We utilize this ability to selectively manipulate cellular chelatable iron levels and provide evidence for two cellular functions of endogenous DNIC formation, protection against anoxia-induced reactive oxygen chemistry from the Fenton reaction and formation by transnitrosation of protein nitrosothiols (RSNO). The levels of RSNO under these high chelatable iron levels are comparable with DNIC levels and suggest that under these conditions, both DNIC and RSNO are the most abundant cellular adducts of ^•NO.     

NdhO,a Subunit of NADPH Dehydrogenase,Destabilizes Medium Size Complex of the Enzyme in Synechocystis sp. Strain PCC 6803

Two mutants that grew faster than the wild-type (WT) strain under high light conditions were isolated from Synechocystis sp. strain PCC 6803 transformed with a transposon-bearing library. Both mutants had a tag in ssl1690 encoding NdhO. Deletion of ndhO increased the activity of NADPH dehydrogenase (NDH-1)-dependent cyclic electron transport around photosystem I (NDH-CET), while overexpression decreased the activity. Although deletion and overexpression of ndhO did not have significant effects on the amount of other subunits such as NdhH, NdhI, NdhK, and NdhM in the cells, the amount of these subunits in the medium size NDH-1 (NDH-1M) complex was higher in the ndhO-deletion mutant and much lower in the overexpression strain than in the WT. NdhO strongly interacts with NdhI and NdhK but not with other subunits. NdhI interacts with NdhK and the interaction was blocked by NdhO. The blocking may destabilize the NDH-1M complex and repress the NDH-CET activity. When cells were transferred from growth light to high light, the amounts of NdhI and NdhK increased without significant change in the amount of NdhO, thus decreasing the relative amount of NdhO. This might have decreased the blocking, thereby stabilizing the NDH-1M complex and increasing the NDH-CET activity under high light conditions.     

Cardiac Restricted Overexpression of Membrane Type-1 Matrix Metalloproteinase Causes Adverse Myocardial Remodeling following Myocardial Infarction

The membrane type-1 matrix metalloproteinase (MT1-MMP) is a unique member of the MMP family, but induction patterns and consequences of MT1-MMP overexpression (MT1-MMPexp), in a left ventricular (LV) remodeling process such as myocardial infarction (MI), have not been explored. MT1-MMP promoter activity (murine luciferase reporter) increased 20-fold at 3 days and 50-fold at 14 days post-MI. MI was then induced in mice with cardiac restricted MT1-MMPexp (n = 58) and wild type (WT, n = 60). Post-MI survival was reduced (67% versus 46%, p < 0.05), and LV ejection fraction was lower in the post-MI MT1-MMPexp mice compared with WT (41 ± 2 versus 32 ± 2%,p < 0.05). In the post-MI MT1-MMPexp mice, LV myocardial MMP activity, as assessed by radiotracer uptake, and MT1-MMP-specific proteolytic activity using a specific fluorogenic assay were both increased by 2-fold. LV collagen content was increased by nearly 2-fold in the post-MI MT1-MMPexp compared with WT. Using a validated fluorogenic construct, it was discovered that MT1-MMP proteolytically processed the pro-fibrotic molecule, latency-associated transforming growth factor-1 binding protein (LTBP-1), and MT1-MMP-specific LTBP-1 proteolytic activity was increased by 4-fold in the post-MI MT1-MMPexp group. Early and persistent MT1-MMP promoter activity occurred post-MI, and increased myocardial MT1-MMP levels resulted in poor survival, worsening of LV function, and significant fibrosis. A molecular mechanism for the adverse LV matrix remodeling with MT1-MMP induction is increased processing of pro-fibrotic signaling molecules. Thus, a proteolytically diverse portfolio exists for MT1-MMP within the myocardium and likely plays a mechanistic role in adverse LV remodeling.     

Microtubule-associated distribution of specific granules and secretion of atrial natriuretic factor in primary cultures of rat cardiomyocytes

A close spatial relationship between specific granules containing atrial natriuretic factor (ANF) and microtubules was demonstrated in primary cultures of neonatal rat cardiac myocytes. For the detection of specific granules and microtubules, the myocytes were double immunolabelled with antibodies against -ANF and -tubulin and examined by conventional fluorescence or laser scanning confocal microscopy. In addition, the ultrastructural distribution of specific granules was demonstrated by electron microscopy. In the atrial myocytes, ANF was stored in numerous specific granules that were mainly localized in the perinuclear sarcoplasm. In the ventricular myocytes, however, a minority of the cells (10%) exhibited limited ANF immunoreactivity after 4 days in culture. Microtubules were present throughout the sarcoplasm of the myocytes. They were most densely packed in the perinuclear regions. Depolymerization of the microtubules with nocodazole was followed by dispersal of ANF immunostaining both in the atrial myocytes and in the ventricular myocytes exhibiting ANF immunoreactivity. When the microtubules were allowed to recover, the perinuclear distribution of specific granules, as seen in non-treated myocytes, reappeared. Measurements of secreted immunoreactive ANF by radioimmunoassay revealed that the secretion of ANF from atrial myocytes into the medium was significantly reduced following nocodazole treatment, whereas a similar decrease in secretion from ventricular myocytes was not observed. These findings indicate that ANF-containing specific granules are closely associated with microtubules within the myocytes. It is suggested that secretion of ANF from the atrial myocytes, in contrast to the ventricular myocytes, is microtubule-dependent.     

草鱼染色体的包装（间期—中期）

以草鱼ＺＣ７９０１细胞株为材料,观察鱼类细胞从间期染色质到中期染色体的包装过程。主要通过（１）分裂期与间期细胞融合,诱导染色体早熟凝集;（２）染色体“伸长”处理;（３）培养细胞的低渗处理;（４）染色质辅展等方法,制作染色体标本,进行扫描和透射电镜观察。观察表明,鱼类染色质的基本结构与哺乳类细胞相同,也是直径约１０ｎｍ的核丝。染色体的色装有两种形式：一种是多级螺旋化形成直径约３００ｎｍ的染色单体,     

Activity of amylin at CGRP1-preferring receptors coupled to positive contractile response in rat ventricular cardiomyocytes

Calcitonin gene-related peptide (CGRP) exerts a positive contractile response directly in rat ventricular cardiomyocytes. This response is mediated by receptors of the CGRP₁-subtype. Amylin is 46% homologous with CGRP and binds to receptors selective for CGRP in a range of tissues. The ability of amylin to influence ventricular contractility has been assessed using cardiomyocytes isolated from the ventricles of adult rats. Cardiomyocytes were subjected to biphasic electrical stimulation at 0.5 Hz. CGRP produced a concentration-dependent positive contractile response which became maximal 4 min after initial stimulation. CGRP increased the contractile amplitude maximally at 1 nM and to a value which was 23.3% greater than in the absence of peptide (EC₅₀ VALUE = 21 pM). Amylin increased the contractile amplitude maximally at 20 nM and to a value which was 17.3% greater than in the absence of peptide (EC₅₀ VALUE = 216 pM). In the presence of amylin (20 nM), the concentration-dependence of the contractile response to CGRP was shifted to the left, so that the response became maximal when CGRP was present at 50 pM. In the presence of CGRP_8–37 (100 nM), a selective antagonist at CGRP₁-preferring receptors, the concentration-dependence of the contractile response to CGRP was shifted to the right (dose RATIO = 54). Similarly, in the presence of CGRP_8–37 (100 nM), the contractile response to amylin was inhibited significantly (P ≤ 0.01). Amylin_8–37 (100 nM) did not inhibit the concentration-dependence of the contractile responses to CGRP and amylin significantly (dose RATIOS = 4.2 and 2.4, respectively). In conclusion, these data indicate that amylin exerts a contractile response directly in rat ventricular cardiomyocytes via CGRP₁-preferring receptors. This effect could assume greater significance in non-insulin-dependent diabetes mellitus and in hypertensive states, in which the concentration of amylin is elevated in plasma.     

Complex interaction between different proteinaceous components within the cell-wall structure ofCandida albicans

We have previously described a monoclonal antibody, MAb DC3:H10, which recognized an epitope preferentially expressed on the surface ofCandida albicans germ tubes. In the present study we examined the MAb-reactive material further. Immunoblot analysis of the material purified partially by Sephadex G-200 and DEAE-Sephacel chromatography reacted with antibodies to theC. albicans C3d receptor (CR2). In an ELISA, MAb DC3:H10 captured antigen that was recognized by both anti-CR2 and anti-mp58 fibrinogen binding mannoprotein polyclonal antibodies. The MAb DC3:H10 failed to compete with either of these antisera in an ELISA. Indirect immunofluo-rescence (IIF) analysis showed differences in surface distribution for the MAb DC3:H10, the CR2, and the mp 58 epitopes. Dual labeling IIF experiments showed MAb DC3:H10 binding to be unaffected by binding of fibrinogen or anti-mp58 antibody. However, the binding patterns of MAb DC3:H10 were modified in the presence of anti-CR2 antibody, suggesting a complex interaction between these cell wall components.     

南京地区早产儿坏死性小肠结肠炎病原体特点和益生菌干预效果及预后影响因素分析

目的探究南京地区早产儿坏死性小肠结肠炎(NEC)的病原体特点,并观察益生菌辅助治疗对患儿血清炎症因子水平的影响,分析影响NEC早产儿预后的因素。方法将2014年5月至2018年12月于东南大学附属中大医院就诊的87例NEC早产儿选为研究对象,分析其病原体特点,观察不同治疗方法对患儿血清炎症因子的影响,并通过Logistic回归分析影响NEC患儿预后的危险因素。结果革兰阴性菌是NEC早产儿的主要致病菌,主要包括肺炎克雷伯菌(25株,27.78%)和大肠埃希菌(21株,23.33%)。肺炎克雷伯菌和大肠埃希菌对美罗培南、亚胺培南/西司他丁、哌拉西林/他唑巴坦、多粘菌素和阿米卡星敏感。益生菌干预后的NEC早产儿血清IL-1β、IL-10、TNF-α水平及TLR4的表达均显著低于常规治疗患儿(均P0.05)。发病前3 d内输血、新生儿窒息、益生菌干预、非营养性吮吸、腹膜炎、败血症、加奶速度、PDA均是影响NEC早产儿预后的相关因素(均P0.05),其中发病前3 d内输血、新生儿窒息、加奶速度、PDA是影响NEC早产儿预后的独立危险因素(均P0.05)。结论革兰阴性菌是NEC早产儿的主要致病菌,益生菌干预可有效降低NEC早产儿血清炎症因子水平。对于NEC早产儿的治疗可选用碳青霉烯类、氨基糖苷类等抗生素,同时需关注影响NEC早产儿预后的独立危险因素。     

Mechanisms of the acute ischemia-induced arrhythmogenesis--a simulation study

The underlying ionic mechanisms of ischemic-induced arrhythmia were studied by the computer simulation method. To approximate the real situation, ischemic cells were simulated by considering the three major component conditions of acute ischemia (elevated extracellular K(+) concentration, acidosis and anoxia) at the level of ionic currents and ionic concentrations, and a round ischemic zone was introduced into a homogeneous healthy sheet to avoid sharp angle of the ischemic tissue. The constructed models were solved using the operator splitting and adaptive time step methods, and the perturbation finite difference (PFD) scheme was first used to integrate the partial differential equations (PDEs) in the model. The numerical experiments showed that the action potential durations (APDs) of ischemic cells did not exhibited rate adaptation characteristic, resulting in flattening of the APD restitution curve. With reduction of sodium channel availability and long recovery of excitability, refractory period of the ischemic tissue was significantly prolonged, and could no longer be considered as same as APD. Slope of the conduction velocity (CV) restitution curve increased both in normal and ischemic region when pacing cycle length (PCL) was short, and refractory period dispersion increased with shortening of PCL as well. Therefore, dynamic changes of CV and dispersion of refractory period rather than APD were suggested to be the fundamental mechanisms of arrhythmia in regional ischemic myocardium.     

随机、平行对照评价陶瓷膜室间隔缺损封堵器安全性和有效性

目的:随机、平行对照的临床试验验证先健科技(深圳)有限公司生产的TM陶瓷膜VSD封堵器治疗室间隔缺损的安全性及有效性。方法:根据试验的要求入选术前确诊为室间隔缺损的病人42例,随机分为实验组(A组)和对照组(B组),每组各21例,对A组病人植入CeraTM陶瓷膜VSD封堵器,对B组病人植入普通镍钛合金VSD封堵器,在超声心动图下或X线下观察室间隔缺损大小、位置、形态、类型,选择合适的封堵伞,经输送系统置入封堵器闭合VSD,分别于术前、术后24小时、术后30天、90天及术后180天进行随访,主要随访内容包括临床症状和体征、超声心动图、心电图、胸片、24小时动态心电图检查等。比较这两组患者的即刻手术成功率以及随访过程中终点事件发生率的不同。结果:试验组和对照组的成功率均为100%,残余分流率分别为0和4.8%,试验组的并发症发生率与对照组未见明显差异(P0.005)。结论:试验组与对照组安全性与疗效相当,应用国产陶瓷封堵器是安全、有效的。但由于该项技术是一项新技术,临床试验样本量较少,其安全性和疗效特别是中远期疗效仍需进一步观察。