急性鸭乙型肝炎病毒感染病毒清除机理研究

目的 :进一步阐明嗜肝病毒自然感染过程中病毒清除机理。方法 :7只 2～ 3月龄成年重庆麻鸭静脉接种 10～ 2 0mlDHBV阳性血清 (5× 10 8～ 1× 10 9genome) ,接种后每周采血检测外周血中DHBVDNA、DHBsAg和特异抗体的产生 ;感染后第10、35天分离外周血单个核细胞用于抗原特异细胞增殖实验 ;第 5、30、6 0天取肝组织标本进行DHBVDNASouthern杂交、DHB sAg免疫组化及肝组织病理检测。结果 :DHBV感染成年鸭在 1～ 2w潜伏期后出现急性、一过性感染 ,感染高峰期肝内存在多拷贝的DHBV所有复制中间体形式 ,包括cccDNA。进一步分析显示病毒血症的消失是在快速抗原特异细胞增殖反应和高滴度特异抗体产生之后 ;与此同时 ,整个急性感染期间 ,肝细胞并无明显的损害。结论 :非细胞直接损伤机制在嗜肝病毒清除过程中发挥了重要作用。     

Polymorphisms and the differential antiviral activity of the chicken Mx gene

The nucleotide sequence of chicken Mx cDNA was reported earlier using the White Leghorn breed in Germany, but it showed no enhanced resistance to viruses. In this study, the nucleotide sequences of chicken Mx cDNA were determined in many breeds. A total of 25 nucleotide substitutions, of which 14 were deduced to cause amino acid exchanges, were detected, suggesting that the chicken Mx gene is very polymorphic. Transfected cell clones expressing chicken Mx mRNA were established after the Mx cDNA was constructed with an expression vector and introduced into mouse 3T3 cells, and the Mx genes from some breeds were demonstrated to confer positive antiviral responses to influenza virus and vesicular stomatitis virus. On the basis of the comparison among the antiviral activities associated with many Mx variations, a specific amino acid substitution at position 631 (Ser to Asn) was considered to determine the antivirally positive or negative Mx gene. Thus, a single amino acid substitution influences the antiviral activity of Mx in domesticated chickens.     

Heteropentameric cholera toxin B subunit chimeric molecules genetically fused to a vaccine antigen induce systemic and mucosal immune responses: a potential new strategy to target recombinant vaccine antigens to mucosal immune systems

Noninvasive mucosal vaccines are attractive alternatives to parenteral vaccines. Although the conjugation of vaccine antigens with the B subunit of cholera toxin (CTB) is one of the most promising strategies for vaccine delivery to mucosal immune systems, the molecule cannot tolerate large-protein fusion, as it severely impairs pentamerization and loses affinity for GM1-ganglioside. Here we report a new strategy, in which steric hindrance between CTB-antigen fusion subunits is significantly reduced through the integration of unfused CTB "molecular buffers" into the pentamer unit, making them more efficiently self-assemble into biologically active pentamers. In addition, the chimeric protein took a compact configuration, becoming small enough to be secreted, and one-step affinity-purified proteins, when administered through a mucosal route, induced specific immune responses in mice. Since our results are not dependent on the use of a particular expression system or vaccine antigen, this strategy could be broadly applicable to bacterial enterotoxin-based vaccine design.     

Oral disodium cromoglycate and ketotifen for a patient with eosinophilic gastroenteritis, food allergy and protein-losing enteropathy

We present a case report of a 10 years old boy with protein-losing enteropathy and eosinophilic gastroenteritis who had positive histamine release tests, increased allergen-specific IgE antibodies to some food items, and low levels of total serum protein and albumin. Upper gastrointestinal endoscopy revealed a number of polyps and diffuse gastritis. Biopsy specimens of the stomach and duodenum showed widespread eosinophilia and neutrophilia. Although a restricted diet was recommended, a diet which excluded foods with positive results to both histamine release test and allergen-specific IgE antibodies was poorly tolerated, and the patient rejected systemic administration of corticosteroids. Thus, we initiated an oral disodium cromoglycate (DSCG) and ketotifen therapy. After oral DSCG and ketotifen administration, the patient's condition improved gradually. Therefore, oral DSCG and ketotifen therapy might be considered as treatment option in patients with eosinophilic gastroenteritis and protein-losing enteropathy caused by food allergy.     

Retrograde dopaminergic neuron degeneration following intrastriatal proteasome inhibition

Recent studies have suggested that defects in the ubiquitin-proteasome system (UPS) contribute to the etiopathogenetic mechanisms underlying dopaminergic neuronal degeneration in Parkinson's disease. The present study aims to study the effects of proteasome inhibition in the nerve terminals of nigrostriatal dopaminergic neurons in the substantia nigra pars compacta (SNpc). Following a unilaterally intrastriatal injection of lactacystin, a selective proteasome inhibitor, dopaminergic neurons in the ipsilateral SNpc progressively degenerated with alpha-synuclein-immunopositive intracytoplasmic inclusions. When lactacystin was administered at a high concentration, the striatum was simultaneously involved, and alpha-synuclein-immunopositive extracytoplasmic granules appeared extensively within the SN pars reticulata (SNpr). In addition, during the retrograde neuron degeneration in SN, the level of heme oxygenase-1 immunopositivity, an oxidative stress marker, was markedly increased in SNpc neurons. These results reveal that intrastriatal proteasome inhibition sufficiently induces retrograde dopaminergic neuronal degeneration with abundant accumulation of alpha-synuclein in the SN.     

Immunolocalization of 3β-hydroxysteroid dehydrogenase in human adrenal cortex and in its disorders

Immunohistochemical localization of 3β-hydroxysteroid dehydrogenase/Δ^5→4-isomerase (3β-HSD), which converts Δ⁵-3β-hydroxysteroids to Δ⁴-3-ketosteroids, was performed in the human adrenal gland and in its disorders by employing a specific antibody raised against the enzyme purified from human placenta. Immunoreactivity of 3β-HSD was present in all three cortical zones of the adrenal glands obtained at autopsy, while in surgically removed adrenal glands, immunoreactivity was dominant in the zona fasciculata (ZF), with faint immunoreactivity in the zona glomerulosa (ZG) and the zona reticularis (ZR). Intracortical localization of 3β-HSD in the adrenal glands obtained at autopsy may represent an adrenal adaptation to antemortem stress, with shifting of adrenal pregnenolone- a substrate of 3β-HSD - as well as steroid 17α-hydroxylase from adrenal androgen synthesis to glucocorticoid synthesis. In adrenocortical hyperplasia, marked immunoreactivity was observed in the ZG and outer ZF in adrenal glands with idiopathic hyperaldosteronism and in the ZF and ZR, especially in cortical micronodules, in the adrenal glands associated with Cushing’s disease. In aldosteronoma and Cushing’s adenoma, immunoreactivity of the enzyme was much more intense in large clear tumor cells than in small compact tumor cells. Immunolocalization of 3β-HSD can yield important information toward an understanding of adrenal steroid metabolism in both physiological and pathological processes.     

Effects of chloride and potassium channel blockers on apoptotic cell shrinkage and apoptosis in cortical neurons

K⁺ and Cl^– homeostasis have been implicated in cell volume regulation and apoptosis. We addressed the hypothesis that K⁺ and Cl^– efflux may contribute to apoptotic cell shrinkage and apoptotic death in cultured cortical neurons. CLC-2 and CLC-3 chloride channels were detected in cultured cortical neurons. The Cl^– channel blockers 4,4-diisothiocyanatostilbene-2,2-disulfonic acid (DIDS), 4-acetamido-4-isothiocyanatostilbene-2,2-disulfonic acid (SITS) and 5-nitro-2-(3-phenylpropylamino)-benzoic acid (NPPB) inhibited the outwardly rectifying Cl^– current, prevented apoptotic cell shrinkage, and mildly attenuated cell death induced by staurosporine, C₂-ceramide, or serum deprivation. Cl^– channel blockers, however, at concentrations that prevented cell shrinkage had no significant effects on caspase activation and/or DNA fragmentation. Cell death in the presence of a Cl^– channel blocker was still sensitive to blockade by the caspase inhibitor z-Val-Ala-Asp(OMe)-fluoromethyl ketone (z-VAD-fmk). Electron microscopy revealed that, although DIDS prevented apoptotic cell shrinkage, certain apoptotic ultrastructural alterations still took place in injured neurons. On the other hand, the K⁺ channel blocker tetraethylammonium (TEA), clofilium, or the caspase inhibitor z-VAD-fmk prevented cell shrinkage as well as caspase activation and/or DNA damage, and showed stronger neuroprotection against apoptotic alterations and cell death. The results indicate that neurons may undergo apoptotic process without cell shrinkage and imply distinct roles for Cl^– and K⁺ homeostasis in regulating different apoptotic events.     

Temporal profile of potassium channel dysfunction in cerebrovascular smooth muscle after experimental subarachnoid haemorrhage

The pathogenesis of cerebral vasospasm after subarachnoid haemorrhage (SAH) involves sustained contraction of arterial smooth muscle cells that is maximal 6–8 days after SAH. We reported that function of voltage-gated K⁺ (K_V) channels was significantly decreased during vasospasm 7 days after SAH in dogs. Since arterial constriction is regulated by membrane potential that in turn is determined predominately by K⁺ conductance, the compromised K⁺ channel dysfunction may cause vasospasm. Additional support for this hypothesis would be demonstration that K⁺ channel dysfunction is temporally coincident with vasospasm. To test this hypothesis, SAH was created using the double haemorrhage model in dogs and smooth muscle cells from the basilar artery, which develops vasospasm, were isolated 4 days (early vasospasm), 7 days (during vasospasm) and 21 days (after vasospasm) after SAH and studied using patch-clamp electrophysiology. We investigated the two main K⁺ channels (K_V and large-conductance voltage/Ca²⁺-activated (K_Ca) channels). Electrophysiologic function of K_Ca channels was preserved at all times after SAH. In contrast, function of K_V channels was significantly decreased at all times after SAH. The decrease in cell size and degree of K_V channel dysfunction was maximal 7 days after SAH. The results suggest that K_V channel dysfunction either only partially contributes to vasospasm after SAH or that compensatory mechanisms develop that lead to resolution of vasospasm before K_V channels recover their function.     

心肌炎患者高频心电图QRS波群的功率谱分析

本文选取６９名心肌炎患者，９４名正常人做为对照。依其年龄不同，共分６组：幼儿心肌炎组，正常幼儿组，学龄心肌炎组，正常学龄组，成人心肌炎组，正常成人组。分别对这６组的Ⅱ导、ａＶＦ导、Ｖ２导、Ｖ５导ＱＲＳ波群０－１０００Ｈｚ、８０－３００Ｈｚ、１００－１０００Ｈｚ３个频段的功率谱（用绝对能量表示）进行了分析。结果提示，不同年龄的心肌炎患者，其心电信号高频频段的能量变化规律不一致。     

系列高频成分节段在正常人中的分布

将１３４例正常人按年龄分为幼儿组、学龄组、青年组和中老年组，分别观察分析了系列高频成分节段的分布特点。其分布无年龄、性别差异（Ｐ＞０．０５），系列高频成分节段对急性病毒性心肌炎的诊断特异性为８２．４１％。各年龄组的特异性分别是幼儿组９０％、学龄组８３．３％、青年组７２．４１％。中老年组８２．０５％。