柯萨奇B3病毒经晚期孕鼠胎盘致母婴感染的小鼠动物模型研究

目的：探讨柯萨奇B3病毒（CVB3m）在间经胎盘垂直传播的可能性和条件，并建立小鼠动物模型。方法：对不同孕期的母鼠分别接种最佳感染量柯萨奇B3病毒嗜心肌株（CVB3m），剖腹取胎或产仔，对母鼠、胎鼠、仔鼠及胎盘分别进行病毒学、血清学及病理检查。结果：晚期孕鼠接种适量的CVB3m后次日形成病毒血症，此时血中无抗CVB3m中和抗体。病毒经胎盘传播形成母婴垂直感染。胎盘、胎鼠、母鼠心肌及仔鼠血清和心肌中均查到较高滴度的病毒。电镜下可见受染仔心肌细胞线粒体肿胀、嵴突缺失、肌纤维断裂、明暗带模糊等初期病变。结论：晚期孕鼠感染CVB3m后形成病毒血症，通过胎盘屏障将病毒传至胎儿，引起胎鼠及仔鼠心肌病变，可作为防治CVB3围产期感染的动物模型。     

抗柯胶囊总皂苷对柯萨奇病毒B型诱导细胞凋亡的抑制作用

目的 观察抗柯胶囊总皂苷抑制柯萨奇病毒B型(CVB)诱导细胞凋亡的作用,探讨其可能的机制.方法 通过接种病毒液的方式建立体外细胞感染模型,采用细胞病变效应法(CPE)测定抗柯胶囊总皂苷对CVB3、CVB4、CVB5株50%抑制浓度(IC50)及治疗指数(TI);CCK-8试剂盒检测细胞存活情况;TdT介导的dUTP缺口...     

我国克山病情处于稳定低发态势

哈尔滨医科大学王铜教授及其课题组通过监测查明,我国16个病区省、327个病区县、2954个病区乡的克山病病情已基本处于稳定低发态势。近年,全国克山病监测点慢型克山病的平均检出率在0.6%左右,潜在型克山病的平均检出率在3．5％左右。目前,克山病监测已由初期注重重点人群发病、病情转归等的疫情监测,转变为着重现患检出率和总体估计的病情监测。北方省区基本消除了急型、亚急型克山病,慢型、潜在型克山病发病大幅度下降。西南地区小儿亚急型克山病得到有效控制。通过膳食中补充硒碘盐的预防措施,     

三七总黄酮对病毒性心肌炎模型治疗作用的药效学研究

目的评价三七总黄酮对柯萨奇B3病毒感染体外、体内模型的治疗作用。方法采用柯萨奇B3型病毒感染原代培养Wistar乳鼠心肌细胞方法,建立体外病毒感染模型;Balb/c小鼠腹腔注射柯萨奇B3型病毒感染建立病毒性心肌炎体内实验动物模型。结果三七总黄酮能明显抑制体外培养心肌细胞病变;治疗组小鼠生存率明显增加,心肌酶释放活性明显降低;小鼠干扰素水平上升,病毒滴度降低;炎性浸润作用明显减轻。结论三七总黄酮对柯萨奇B3病毒所致小鼠病毒性心肌炎具有一定治疗作用。     

中药抗柯萨奇B3病毒的实验研究进展

柯萨奇B3病毒（CVB3）属小RNA病毒科（Picomaviridae）肠道病毒属（enterviruses），是引起病毒性心肌炎的主要原因，它可在心肌长期潜伏，持续性病毒感染不但会直接损伤心肌细胞，还可引起免疫反应性病理损伤，形成扩张性心肌病，对人类的健康造成极大的威胁。目前尚无有效的抗CVB3药物，以往的研究结果显示中药在抗CVB3方面具有抑制病毒复制，调节免疫功能，保护心肌细胞等作用，临床应用前景广阔。     

小柴胡汤对体外感染柯萨奇病毒心肌细胞琥珀酸脱氢酶的影响

为探讨小柴胡汤对病毒性心肌炎的治疗作用,以体外培养的新生大鼠心肌细胞感染柯萨奇病毒B3(CVB3)作为病毒性心肌炎模型,用酶组织化学方法及图像分析观测心肌细胞感染CVB348h后琥珀酸脱氢酶(SDH)的活性,并观察小柴胡汤治疗后的影响。结果表明,CVB3感染心肌细胞48h,SDH活性明显下降;经小柴胡汤治疗后,SDH活性接近正常水平。结论:小柴胡汤具有抗柯萨奇病毒感染和保护心肌细胞作用。     

柯萨奇病毒B组5型研究进展

 柯萨奇病毒B组5型（coxsackievirus B5,CVB5）近年在中国引起多起手足口病和无菌性脑膜炎暴发。分子流行病学研究显示出现新型CVB5传播,从而给防控肠道病毒感染所致疾病带来了新的挑战。此文就CVB5的流行病学、进化特征、动物模型以及实验室诊断等方面的研究进行综述。     

2006年河北省张北县克山病监测点监测结果分析

河北省克山病分布于河北省北部11个县,历史上曾多次爆发流行。通过综合防治,近20多年无急型、亚急型克山病发生,克山病病情相对稳定。为掌握当前克山病病情动态,指导全省克山病防治工作,2006年选择在自然环境、主要农作物和生活习惯等具有普遍代表性的克山病历史病区张北县战海乡水泉洼村作为克山病病情监测点开展病情监测,监测结果分析报告如下。     

小儿克山病心肌酶活性对比观察

我室于1985年至1986年间,分别对克山病患儿和健康儿童进行心肌酶学活性对比观察,以寻找病区健康儿童心肌酶学指标,判别不同临床型克山在心肌损伤不同时期的生化特点,为小儿克山病的早期诊断提供有价值的临床资料。一、材料和方法 1.对象及分组本文选择我所住院治疗的小儿克山病患者为实验组,平均年龄8.7岁;居住郊区的职工户健康子女为对照组,平均年龄9.1岁。两组经临床体检以及X线、心电图、心肌图、超声心动图等项实验室检查,确诊亚急型克山病13例,慢型克山病     

蛋白酶体抑制剂MG-132对柯萨奇B3病毒性心肌炎小鼠的作用研究

目的：研究泛素蛋白酶体抑制剂MG-132对柯萨奇B3（CVB3）病毒性心肌炎小鼠的作用,探讨泛素蛋白酶体系统在病毒性心肌炎发病学中的作用机制。方法：随机将80只雄性BALB／C小鼠分为4组（,2—20）：正常对照组、心肌炎组、心肌炎＋处理组、正常＋处理组。腹腔接种CVB3诱发急性心肌炎,次日腹腔注射MG-132,0．75mg／kg;连续给药7d,对照组腹腔注射PBS。第8天小鼠取材,观察心脏病理变化,测定心肌CVB3病毒复制及血清肌钙蛋白、脑钠肽水平。结果：MG-132显著减轻心肌炎小鼠心脏病理损伤,显著降低心脏重量／身体重量比值,MG-132干预后第8天小鼠血清肌钙蛋白、脑钠肽水平显著降低,同时荧光定量PCR显示CVB3mR—NA复制水平显著降低。结论：MG-132通过抑制CVB3病毒复制,显著减轻心肌炎小鼠心脏病理损伤,起到保护心肌作用。     

西罗莫司对CVB3感染大鼠心肌成纤维细胞增殖及mTOR表达的影响

目的:观察西罗莫司(又名雷帕霉素,Rap)对柯萨奇病毒B3(CVB3)感染的心肌成纤维细胞增殖及哺乳类雷帕霉素靶蛋白(mTOR)mRNA和蛋白表达的影响。方法:SD大鼠心肌成纤维细胞分对照组、CVB3组、Rap组及CVB3+Rap组,用重复感染率(MOI)为0.5噬斑形成单位(PFU)/cell的CVB3诱导心肌成纤维细胞,采用四氮唑蓝(MTT)法检测细胞增殖,RT-PCR及Western Blot分别检测其mTOR mRNA表达及mTOR蛋白表达。结果:MTT法测得对照组、CVB3组、Rap组及CVB3+Rap组的D值分别为0.55±s 0.09、0.66±0.09,0.42±0.09、0.36±0.12,CVB3组高于对照组,Rap组和CVB3+Rap组均低于对照组及CVB3组(P<0.05)。用RT-PCR检测上述4组的mTOR mRNA表达,mTOR/β-actin光密度比值分别为0.25±0.09、0.87±0.15、0.19±0.04、0.22±0.06,CVB3组高于对照组(P<0.05),Rap组和CVB3+Rap组低于对照组及CVB3组(P<0.05)。Western Blot显示mTOR蛋白表达结果与RT- PCR结果相符。结论:Rap可抑制CVB3病毒诱导的大鼠心肌成纤维细胞增殖及mTOR表达,Rap抑制细胞增殖可能是通过mTOR信号途径起作用的。     

The human fatty acid synthase: a new therapeutic target for coxsackievirus B3-induced diseases?

Coxsackievirus is linked to a large variety of severe human and animal diseases such as myocarditis. The interplay between host factors and virus components is crucial for the fate of the infected cells. However, host proteins which may play a role in coxsackievirus-induced diseases are ill-defined. Two-dimensional gel electrophoresis of protein extracts obtained from coxsackievirus B3 (CVB3)-infected and uninfected HeLa or HepG2 cells combined with spot analysis revealed several proteins which are exclusively up-regulated in infected cells. One of these proteins was identified as the fatty acid synthase (FAS). By using cerulenin and C75, two known inhibitors of FAS we were able to significantly block CVB3 replication. FAS appears to be directly involved in CVB3-caused pathology and is therefore suitable as a therapeutic target in CVB3-induced diseases.     

New pleconaril and [(biphenyloxy)propyl]isoxazole derivatives with substitutions in the central ring exhibit antiviral activity against pleconaril-resistant coxsackievirus B3

Amino acid 1092 (AA1092) in capsid protein 1 of coxsackievirus B3 (CVB3) is located in close vicinity to the central phenoxy group of capsid binders (i.e. pleconaril). Whereas isoleucine is associated with drug susceptibility, leucine and methionine confer resistance to pleconaril. In the present study, novel analogues with different substitutions in the central phenoxy group were synthesized to study their influence on anti-CVB3 activity with the aim to overcome pleconaril resistance. Two [(biphenyloxy)propyl]isoxazoles and pleconaril were synthesized without methyl groups in the central phenoxy ring using Suzuki coupling reaction and tested for antiviral activity against the pleconaril-resistant CVB3 Nancy. Furthermore, pleconaril with 3-methyl, 3-methoxy, 3-bromine, 2,3-dimethyl in the central ring as well as the external rings in meta position were synthesized for structure-activity relationship analysis with CVB3 variants containing leucine, methionine or isoleucine at position 1092, other coxsackieviruses B (CVB) as well as several rhinoviruses. The results demonstrate a high impact of substituents in the central ring of capsid inhibitors for anti-enteroviral activity. Pleconaril resistance of CVB3 based on Leu1092 or Met1092 was overcome by unsubstituted analogues or by monosubstitution with 3-methyl as well as 3-bromine in the central phenyl. The 3-bromine derivative inhibited a broad spectrum of CVB and rhinoviruses.     

Viral infection and PBDE exposure interact on CYP gene expression and enzyme activities in the mouse liver

In the present study coxsackievirus B3 (CVB3) adapted to Balb/c mice was used to examine whether infection affects xenobiotic-metabolising CYP1A1 and CYP2B gene expression (measured by RT-PCR) and the corresponding enzyme activities of ethoxyresorufin-O-deethylase (EROD) and pentoxyresorufin-O-depentylase (PROD), as observed on day 3 of infection. To study the simultaneous effects of xenobiotic exposure, mice were administered the polybrominated diphenyl ether (PBDE) compounds BDE-99 (single congener) and Bromkal 70-5 DE (commercial mixture). Serum thyroxine levels were also measured. High numbers of CVB3 were found in the livers of infected mice but no significant effects of PBDE on virus replication were observed. In infected mice gene expression and CYP activities were decreased in comparison with non-infected mice, especially for CYP2B. PBDE exposure in the non-infected mice was characterised by an increase in both CYP2B and PROD levels/activities, whereas CYP1A levels increased and EROD activity decreased. In general, PBDE exposure in the infected mice did not increase EROD and PROD activities to the same extent as in the non-infected exposed mice. Infected mice exposed to BDE-99 showed significantly higher CYP2B and PROD levels than both the infected non-exposed and Bromkal-exposed groups. T(4) levels were greatly decreased by infection and a tendency of reduced T(4) levels after PBDE exposure could be observed in non-infected mice. In conclusion, infection reduced the detoxifying capacity of the liver and the serum T(4) levels. PBDE exposure can modify these effects. Notably, in the infected mice differences between BDE-99 and Bromkal were observed on CYP2B gene expression and PROD activity.     

The antiviral effect of jiadifenoic acids C against coxsackievirus B3

Coxsackievirus B type 3 (CVB3) is one of the major causative pathogens associated with viral meningitis and myocarditis, which are widespread in the human population and especially prevalent in neonates and children. These infections can result in dilated cardiomyopathy (DCM) and other severe clinical complications. There are no vaccines or drugs approved for the prevention or therapy of CVB3-induced diseases. During screening for anti-CVB3 candidates in our previous studies, we found that jiadifenoic acids C exhibited strong antiviral activities against CVB3 as well as other strains of Coxsackie B viruses (CVBs). The present studies were carried out to evaluate the antiviral activities of jiadifenoic acids C. Results showed that jiadifenoic acids C could reduce CVB3 RNA and proteins synthesis in a dose-dependent manner. Jiadifenoic acids C also had a similar antiviral effect on the pleconaril-resistant variant of CVB3. We further examined the impact of jiadifenoic acids C on the synthesis of viral structural and non-structural proteins, finding that jiadifenoic acids C could reduce VP1 and 3D protein production. A time-course study with Vero cells showed that jiadifenoic acids C displayed significant antiviral activities at 0–6 h after CVB3 inoculation, indicating that jiadifenoic acids C functioned at an early step of CVB3 replication. However, jiadifenoic acids C had no prophylactic effect against CVB3. Taken together, we show that jiadifenoic acids C exhibit strong antiviral activities against all strains of CVB, including the pleconaril-resistant variant. Our study could provide a significant lead for anti-CVB3 drug development.KEY WORDS: CVB3, Jiadifenoic acids C, Antiviral activityAbbreviations: CAR, coxsackievirus and adenovirus receptor; CPE, cytopathic effect; CVB3, coxsackievirus B type 3; CVBs, coxsackie B viruses; DAF, decay accelerating factor; DCM, dilated cardiomyopathy; IC₅₀, 50% inhibitory concentration; IRES, internal ribosomal entry site; MOI, multiplicity of infection; NTR, non-translated region; RBV, ribavirin; RdRp, RNA-dependent RNA polymerase; SI, selectivity index; Vero, African green monkey kidney cells     

Selenium and viral virulence

A mouse model of coxsackievirus-induced myocarditis is being used to investigate nutritional determinants of viral virulence. This approach was suggested by research carried out in China which showed that mice fed diets composed of low selenium ingredients from a Keshan disease area suffered more extensive heart damage when infected with a coxsackie B4 virus than infected mice fed the same diet but supplemented with selenium by esophageal intubation. Selenium deficiency in our mice increased the virulence of an already virulent strain of coxsackievirus B3 (CVB3/20) and also allowed conversion of a non-virulent strain (CVB3/0) to virulence. Such conversion of CVB3/0 was accompanied by a change in the viral genome to more closely match that of the virulent virus, CVB3/20. As far as the authors are aware, this is the first report of host nutrition influencing the genetic make-up of an invading pathogen. Nutritionists may need to consider this mechanism of increased viral virulence in order to gain a better understanding of diet/infection relationships.     

牛磺酸对感染柯萨奇B3病毒大鼠心肌细胞L型电压依赖性钙通道的影响

AIM: To study the effects of taurine on L-type voltage-dependent Ca2+ channel (VDCC) in adult rat cardiomyocytes infected with coxsackievirus B3 (CVB3). METHODS: Whole-cell Ca2+ current of L-type VDCC was obtained by patch-clamp techniques. RESULTS: The density of L-type Ca2+ current was 4.1 +/- 0.8 pA/pF in normal cardiomyocytes, but increased to 4.9 +/- 1.4 pA/pF with CVB3 infection. At 16 mmol.L-1, taurine decreased the density to 3.5 +/- 0.5 pA/pF in normal cardiomyocytes, and to 3.8 +/- 0.8 pA/pF in CVB3-infected cardiomyocytes. In addition, CVB3 shifted the membrane potential depolarizing to peak current (Vp) from 8 +/- 8 mV to 5 +/- 3 mV which could also be reverted to 8 +/- 4 mV by taurine. CONCLUSION: Taurine inhibited the increase of Ca2+ inflow through L-type VDCC and normalized the decreased Vp induced by CVB3 infection. The effect of taurine on L-type VDCC was the mechanism of taurine attenuating the intracellular Ca2+ accumulation and abnormal electric activities induced by CVB3 infection.     

Polybrominated diphenyl ether exposure suppresses cytokines important in the defence to coxsackievirus B3 infection in mice

Environmental pollutants can adversely affect the immune system. The host defence during infection depends on cytokine signalling and proper function of immune cells. However, no studies have addressed how polybrominated diphenyl ethers (PBDEs) affect cytokine responses. We investigated the combined effects in Balb/c mice of human coxsackievirus B3 (CVB3) infection and exposure to PBDEs (BDE-99 or Bromkal mixture) on 21 serum cytokines. The mice were infected (i.p.) on day 0, orally treated with BDE-99 or Bromkal on day 1 (20 mg/kg bw) and put to death on day 3. CVB3 was quantitatively measured in the liver and pancreas by RT-PCR. The Luminex 200 multi-analyte system was used for cytokine analysis. High numbers of viral copies were found in the liver and pancreas. Infection increased TNF-α, IL-6, MCP-1, IL-12p40, KC and RANTES levels. Notably, PBDE-exposure resulted in a marked decrease, or even lack, of IL-13, MIP-1β, RANTES, IFN-γ and KC levels in non-infected mice. However, the effects of PBDE-exposure on cytokines did not affect viral replication during early CVB3 infection. In conclusion, PBDEs causes a selective block in immune signalling pathways but the consequences of this need to be further studied in different host resistance models of infection.     

柯萨奇B3病毒对心脏发育因子GATA-4、NKx2.5表达的影响及叶酸对其的保护作用

【摘要】目的探讨叶酸对孕期感染柯萨奇B3病毒(CVB3)的新生鼠心脏发育因子GATA-4、NKx2.5的基因与蛋白表达的影响。方法SD雌鼠分为对照组、叶酸组、CVB3组、CVB3+叶酸组。叶酸组及CVB3+叶酸组孕前给予叶酸灌胃2周;CVB3组及CVB3+叶酸组孕第7天腹腔注射CVB3,连续5d,自然分娩后取新生鼠心脏组织,病理切片HE染色观察新生鼠心脏组织形态学变化;逆转录聚合酶链反应（RT-PCR）及Western blot测定新生鼠心脏组织中转录因子GATA-4、NKx2.5基因及蛋白表达水平。结果CVB3组新生鼠心肌损伤明显,心肌细胞排列紊乱,心肌纤维断裂,CVB3+叶酸组状态改善。CVB3组新生鼠心脏组织GATA-4、NKx2.5基因及蛋白表达低于对照组（P＜0.05）,CVB3+叶酸组GATA-4、NKx2.5基因及蛋白表达高于CVB3组（P＜0.05）。结论孕鼠早期感染CVB3抑制新生鼠心脏发育因子正常表达,补充叶酸对新生鼠心脏发育具有明显保护作用。