Comparative study of histopathology changes between the PS1/APP double transgenic mouse model and Aβ1-40 -injected rat model of Alzheimer disease

Objective To identify the genetype of the PS1/APP double transgenie mouse model, then to analyse the histopathological changes in the brain and compare the differences between the transgenie mice models and Aβ1-40-injeeted rats models of Alzheimer disease. Methods The modified congo red staining, Nissl＇s staining and immunohistology staining was used to observe the Aβ deposits, activation of astrocyte respectively. Results ①The PS1/APP transgenic mouse extensively displayed Aβ deposits in the cortex and hippocampal structures, and GFAP positive cells were aggregated in mass and surrounded the congo red-positive plaque. ②The Aβ1-40-intrahippocmnpal-injeeted rat model showed the Aβ plaque deposits in the dentate gyrus of the hippocampus, with the astrocyte surrounded. The neurons loss was significant in the injection point and pin hole of injection with Nissl＇s staining methods. GFAP-positive cells increased significantly compared with the uninjected lateral of the hippocampus. Conclusion Although Aβ1-40-injected rat models could simulate some characteristic pathological features of human Alzheimer diseases, Aβ deposits and neurons loss in partial hippocampal, it would not simulate the progressive degenenration in the brain of AD. The double transgenie PS1/APP mice could simulate the specific pathogenesis and progressive changes of AD, mainly is Aβ deposits and the spongiocyte response , while no neurons loss were observed in this model.     

Pilot-scale production and characterization of paramyosin, a vaccine candidate for schistosomiasis japonica

Despite effective chemotherapy, schistosomiasis remains a major public health problem in the developing world, with at least 200 million active infections resulting in significant morbidity. Rapid reinfection after treatment, accompanied by extensive residual morbidity, mandates alternative control strategies, including vaccine development. Paramyosin, a myofibrillar protein found only in invertebrates, has been widely studied as a vaccine candidate for both Schistosoma mansoni and Schistosoma japonicum. Recently, we demonstrated that Th2-biased immune responses to paramyosin are associated with resistance to reinfection with S. japonicum in humans; however, challenges in the pilot-scale production of schistosome paramyosin have hampered further studies of this promising vaccine candidate. Here we report a method for the pilot-scale expression and purification of recombinant S. japonicum paramyosin (rSj97). rSj97 was extracted from Escherichia coli inclusion bodies and purified with sequential anion-exchange, hydroxyapatite, and size exclusion chromatography. The purified rSj97 was >95% pure as judged by sodium dodecyl sulfate-polyacrylamide gel electrophoretic analysis and was free of significant endotoxin contamination. We demonstrate that, like native paramyosin, rSj97 adopts an alpha-helical coiled-coil tertiary structure and binds immunoglobulin and collagen. Naïve mice infected with S. japonicum produce anti-rSj97 immunoglobulin G (IgG) antibodies as early as 4 weeks postinfection, while sera collected from S. japonicum-infected individuals contain anti-rSj97 IgE antibodies. Our method for pilot-scale production of recombinant full-length paramyosin will facilitate preclinical evaluation of paramyosin as a vaccine for schistosomiasis.Schistosomiasis remains a major public health concern in the developing world, with 200 million individuals infected and 600 million at risk of infection (6). The disease is caused by parasitic helminths of the genus Schistosoma and is prevalent in sub-Saharan Africa, Asia, Latin America, and the Middle East. Severe morbidity and mortality are associated with end-organ liver or urinary tract pathology, while recent work has highlighted subtle morbidities associated with the chronic nature of the disease, such as anemia and malnutrition (11). Because of rapid reinfection, the prevalence of infection and morbidity has not been adequately reduced despite effective chemotherapy with praziquantel. For this reason, “vaccine-linked chemotherapy” has been advocated as an alternative control strategy (1). Even with a nonsterilizing, suboptimal vaccine, mathematical models predict significant long-term reductions in infection prevalence and intensity when this approach is targeted either to humans (3) or to livestock for the zoonotic species Schistosoma japonicum (25). These models justify the search for an antischistosomal vaccine to effect durable reductions in the prevalence and intensity of infection and to limit subtle morbidities that persist despite continued treatment.Paramyosin, a 97-kDa muscular protein found exclusively in invertebrates, is a recognized priority vaccine candidate for both Schistosoma mansoni and S. japonicum (2). Paramyosin adopts a coiled-coil dimer structure composed of two parallel alpha-helices wrapped in a left-handed, supercoiled twist (10). Paramyosin was identified as the major immunogen in S. mansoni freeze-thawed larvae, a crude parasite preparation that confers significant protection in murine challenge studies (14). Independently, S. japonicum paramyosin (Sj97) was identified as the target of a monoclonal immunoglobulin E (IgE) antibody that conferred protection upon passive immunization of mice (12). In addition to its subtegumental location, paramyosin is also expressed on the surfaces of schistosomula (9), providing a target for immune attack via antibody-dependent cellular cytotoxicity, presumably mediated by eosinophils (12). Surface-expressed paramyosin has been implicated in immune evasion strategies due to its Ig (17), collagen (13), and complement protein (5) binding activities.Murine immunization studies using both biochemically purified and recombinant paramyosins have consistently demonstrated significant protection from challenge infection. Studies with S. mansoni demonstrate a 24 to 56% reduction in worm burdens, while protection against S. japonicum ranged from 32 to 86% (reviewed in reference 8). Maximal protection (62 to 86%) against S. japonicum was observed by using biochemically purified paramyosin followed by challenge with the Philippine strain of S. japonicum (20).Subsequently, several immunoepidemiologic studies conducted in areas where schistosomiasis is endemic have associated antigen-specific immune responses to paramyosin with resistance to infection. In a cohort study in Brazil, uninfected individuals had threefold-higher levels of IgG antibody to paramyosin than stool-positive individuals. Importantly, after antischistosomal treatment, individuals who remained stool negative showed elevated antiparamyosin antibodies over 32 months compared to those who continued to excrete eggs (4). Recently, we demonstrated in a longitudinal treatment-reinfection study in Leyte, Philippines, that Th2-biased cytokine responses to Sj97 predict a significantly longer time to reinfection and a 30 to 41% lower intensity of reinfection with S. japonicum following treatment with praziquantel (15).Further development of paramyosin as a vaccine candidate for schistosomiasis has been halted due to the inability to express and purify this protein at a pilot scale. Specifically, poor yields and insolubility in bacterial and yeast expression systems (19, 27) have resulted in paramyosin being “shelved” from the vaccine priority list (1). Here we report a robust process for the pilot-scale production of recombinant full-length S. japonicum paramyosin (rSj97) with assessment of the protein''s structure, binding properties, and antigenicity.     

An ingenious method for the determination of the relative and absolute configurations of compounds containing aryl-glycerol fragments by 1H NMR spectroscopy

A concise method was established to determine the relative and absolute configurations of aryl-glycerols that depend on the chemical shift differences (Δδ) of the diastereotopic methylene protons (H-3) by ¹H NMR spectroscopy. When using DMSO-d₆ as the preferred solvent, the threo configuration corresponded to a larger Δδ_H3a–H3b value (>0.15 ppm), whereas the erythro configuration (<0.07 ppm) corresponded to a smaller value. Furthermore, the absolute configurations were determined with the aid of a simple acylation reaction through camphanoyl chloride. In the threo enantiomers, the Δδ value of the 1R,2R configuration was <0.15 ppm, and that of the 1S,2S configuration was >0.20 ppm. In the erythro enantiomers, the Δδ value of 1R,2S was >0.09 ppm, and that of 1S,2R was <0.05 ppm. Remarkably, this empirical rule is invalid in CDCl₃. In addition, this method was also verified by a quantum ¹H NMR calculation.

A concise method was established to determine the relative and absolute configurations of aryl-glycerols that depend on the chemical shift differences (Δδ) of the diastereotopic methylene protons (H-3) by ¹H NMR spectroscopy.     

肺癌侵袭力的影像学表现与c-erbB-2及bcl-2基因蛋白表达的关系

目的和方法：利用免疫组化技术ＬＳＡＢ法和计算机图像分析系统的结合,测定经手术病理证实之３７例肺鳞癌及４８例肺腺癌组织中ｃ－ｅｒｂＢ－２及ｂｃｌ－２基因蛋白的表达水平,分析其表达或共表达与反映肺癌侵袭力的影像征象的关系。结果：ｃ－ｅｒｂＢ－２及ｂｃｌ－２基因蛋白在两种不同类型肺癌组织中的阳性表达及表达定位有明显的差异,并且,ｃ－ｅｒｂＢ－２基因蛋白表达阳性的肺癌,肿块毛刺征的出现率明显高于表达阴性者;ｂｃｌ－２基因蛋白表达阳性的肺癌,肿块毛刺征及三级支气管受累的出现率明显高于表达阴性者（Ｐ＜００５）;当两种基因蛋白在肺鳞癌中共表达时,肿块毛刺征及肺门或纵隔淋巴结转移的出现率明显高于它们单独表达者（Ｐ＜００５）。结论：ｃ－ｅｒｂＢ－２及ｂｃｌ－２基因蛋白与反映肺癌侵袭力的某些影像征象之间可能存在密切的关系。     

E1A激活基因阻遏子在不同表型血管平滑肌细胞中的表达

目的 探讨血管平滑肌细胞表型转换、分化过程中E1A激活基因阻遏子(cellular repressor of E1A-stimulated genes,CREG)的表达变化及其相关机制。方法 将PCR扩增的CREG片段插入pGEX-4T-1原核表达载体,纯化的CREG融合蛋白免疫家兔以制备多克隆抗体。以人胸廓内动脉平滑肌细胞(human internal thoracic artery smooth muscle cells,HITASY)为模型,[^3H]标记脱氧胸苷掺入法测定去血清培养HITASY细胞DNA合成变化。以Western印迹观察HITASY细胞在去血清培养过程中SM α-肌动蛋白、肌钙结合蛋白(calponin)的表达,RT-PCR和Western印迹分析去血清培养诱导CREG mRNA和蛋白表达变化;免疫组化法观察CREG在细胞内的表达及定位。结果 构建载体并成功得到抗CREG多克隆抗体。去血清培养可使HITASY细胞DNA合成明显降低。随去血清培养时间延长,除SM α-肌动蛋白和肌钙结合蛋白表达逐渐上调之外,CREG mRNA转录活性和蛋白翻译合成亦上调。免疫组化显示去血清培养后,CREG蛋白在细胞内表达并主要位于细胞核周。结论 去血清能促使HITASY细胞由合成表型向收缩表型转换,同时伴CREG mRNA和蛋白表达上调。     

多焦点人工晶状体植入的研究进展

白内障是导致患者视力下降乃至失明的主要原因之一,手术是治疗白内障确切有效的手段。随着科学技术的进步,传统的复明性白内障手术逐渐过渡到屈光性白内障手术。各种屈光性人工晶状体也应运而生,传统的单焦点人工晶状体(single focus intraocular lens, SIOL)不再是患者的唯一选择,多焦点人工晶状体(multifocal intraocular lens, MIOL)越来越被患者接受和认可。本文对现有的多焦点人工晶状体进行了归纳总结,通过分类列举,简述不同类型多焦点人工晶状体的多种特点及评估患者术后临床效果的方法,以期为眼科医生提供参考。     

Disruption of Bombyx mori nucleopolyhedrovirus ORF71 (Bm71) results in inefficient budded virus production and decreased virulence in host larvae

The Bombyx mori nucleopolyhedrovirus (BmNPV) is a baculovirus that selectively infects domestic silkworm. BmNPV ORF71 (Bm71) is not a core set gene in baculovirus and shares 92 % amino acid sequence identity with Autographa californica multinucleocapsid NPV ORF88 (Ac88/cg30). Previously, it has been reported that virus lacking Ac88 had no striking phenotypes in cell lines or host larvae. However, the exact role of Bm71 during BmNPV life cycle remains unknown. In the present study, we constructed a Bm71-disrupted (Bm71-D) virus and assessed the effect of the Bm71 disruption on viral replication and viral phenotype throughout the viral life cycle. Results showed that the Bm71-D bacmid could successfully transfect Bm5 cell lines and produce infectious budded virus (BV). But the BV titer was 10- to 100-fold lower than that of the wild-type (WT) virus during infection, and the decreased BV titer was rescued by Bm71 gene repair virus (Bm71-R). A larval bioassay showed that Bm71-D virus took 7.5 h longer than the WT to kill Bombyx mori larvae. Transmission electron microscopy analysis indicated that the Bm71-D virus-infected cells had typical virogenic stroma, bundles of nucleocapsids and polyhedra. Taken together, these results suggest that Bm71 has important implications for determining BV yield and virulence in viral life cycle even though it is not an essential gene for replication of BmNPV.