人乳头瘤病毒多肽与人免疫球蛋白G融合表达

目的 将人乳头瘤病毒16型(Human papillomavirus type 16，HPV-16)的晚期表达蛋白E7上的抗原24肽(从第38位氨基酸到第61位氨基6病毒感染防治酸)与人免疫球蛋白G的重链恒定区融合表达，并以此融合蛋白作为抗原，可能为HPV-1提供免疫治疗方法。方法 利用PCR方法分别扩增HPV-16 E7(38-61)24肽的DNA片段和人免疫球蛋白G的重链恒定区DNA片段，并构建到pEV21a表达载体上，转化入E．coli中表达，利用SDS-聚丙烯酰胺凝胶电泳(SDS-PAGE)和蛋白质免疫印迹(Western-blotting)的方法对表达结果进行鉴定。结果 构建的表达载体HPV16E7e／hIgGHCCR-pET21a经酶切鉴定和测序显示序列正确；通过SDS-PAGE和Western-blotting的鉴定，重组融合蛋白Mr约40000，表达量可占菌体蛋白的20％左右。结论 成功构建HPV16-E7的抗原多肽片段和人免疫球蛋白G的重链恒定区的融合蛋白，并可在E．coli中高效表达。     

IL-2对垂体瘤细胞系RC-4B/C细胞分泌ACTH的影响

目的 :探讨白细胞介素 2 (IL 2 )对垂体瘤细胞系RC 4B/C细胞ACTH分泌的调控作用及其影响因素 .方法 :以放射免疫方法测定培养的垂体瘤细胞系RC 4B/C细胞的培养液中的ACTH浓度 .结果 :IL 2 (1× 10 4～ 5× 10 5U·L-1)促进RC 4B/C细胞分泌ACTH ;蛋白激酶A的抑制剂H 9(1μmol·L-1)和酪氨酸蛋白激酶的抑制剂tyrphostin2 3 (1μmol·L-1)均可显著性抑制IL 2的促ACTH分泌作用 .结论 :IL 2可促进RC 4B/C细胞分泌ACTH ,该作用与蛋白激酶A和酪氨酸蛋白激酶信号转导途径紧密相关     

大豆蛋白影响肾损害模型钙摄入和钙排出的实验研究

[目的]研究大豆蛋白对肾损害大鼠钙平衡的影响。[方法]选择3月龄断乳雄性SD大鼠40只．按体质量从小到大排序，采用完全随机化原则分为4组。每组10只。标准饲料对照组：喂食含有14％酪蛋白饲料；大豆蛋白饲料组：喂食含有14％大豆分离蛋白饲料；混合饲料1组：喂食含有7％酪蛋白加7％大豆分离蛋白饲料；混合饲料2组：喂食含有7％酪蛋白加14％大豆分离蛋白饲料。实验期用腺嘌呤灌胃共21d，建立肾损害大鼠模型，各组大鼠喂养相应饲料6周，测饲料消耗量、24h尿蛋白、尿钙、钙表观吸收率、储留钙量。[结果]4组实验大鼠饲料摄入量1d～30d无统计学差异，大豆蛋白饲料组和混合饲料1组实验结束时与另外两组相比，尿钙排泄量低，钙表观吸收率和储留钙量高，差异有统计学意义（P〈0．05）。[结论]蛋白质含量水平在14％条件下，含大豆蛋白的两种饲料对肾损害大鼠钙吸收、钙排泄的影响无统计学差异，有利于促进钙吸收，减少钙排泄。     

恶性肿瘤局部淋巴结树突状细胞免疫组化研究

本文采用S-100蛋白为免疫学标记，对34例恶性肿瘤局部淋巴结内树突状细胞进行免疫组化定量研究。结果按S-100蛋白阳性细胞数目多少分为增多（7例）、减少（20例）及正常（7例）3组，统计学分析增多组均值（164.4个/mm^2)明显高于对照组（58.3个/mm^2)；减少组均值（16.5个/mm^2)组显低于对照组；而正常组均值（69.8个/mm^2)与对照组无显著差异。     

Intrathecal synthesis of anti-myelin basic protein IgG in HIV-1+ patients

Human immunodeficiency virus type 1 (HIV-1)-infected individuals frequently develop a broad spectrum of neurological syndromes, classified as HIV-1-associated cognitive/motor complex. Diffuse demyelination of hemispheric white matter is a commonly observed in HIV-1 infected brain, but the events leading to myelin destruction are still obscure. Since oligodendrocyte infection by HIV-1 is not proven as yet, myelin damage in HIV-1 infection may result from indirect mechanisms such as the excessive release of myelinotoxic substances or the triggering of autoimmune responses directed to myelin constituents. To verify the latter hypothesis, we searched for elevated anti-myelin basic protein (MBP) IgG levels in the cerebrospinal fluid (CSF) and serum of 25 patients with HIV-1 infection, 12 with multiple sclerosis (MS), and 9 with non-inflammatory neurological diseases (NIND). CSF, but not serum, anti-MBP IgG levels were more frequently elevated in HIV-1⁺ (16/25, 64%) than in MS (3/12, 25%) or NIND (0/9) patients. By using the anti-MBP IgG index, the anti-MBP IgG antibody specificity index (ASI), and the search for anti-MBP oligoclonal IgG, we ascertained that anti-MBP IgG were produced within the CNS in 13 of 25 (52%) HIV-1⁺, in 6 of 12 (50%) MS, and in none of NIND patients. The incidence of increased CSF anti-MBP IgG levels was higher among HIV-1⁺ patients at stage II–III (4/4, 100%) or at stage IV B (7/9, 78%) than among those at stage IV C–IV D (5/12, 42%). Although our data indicate that intrathecal anti-MBP IgG may occur early during HIV-1 infection and that they are more common in patients with HIV-1-associated cognitive/motor complex, the possible demyelinating role of these antibodies remains to be demonstrated.     

Astrocyte RNA in relation to neuronal RNA depletion in Alzheimer's disease

Summary A new double-staining procedure, in which the techniques of immunocytochemistry of glial fibrillary acidic protein (GFAP) and quantitative microdensitometry of azure B-RNA were combined, was used to study nucleic acid alterations in fibrous astrocytes in Alzheimer's disease (AD). RNA contents of GFAP-positive cells of the hippocampal endplate (Rose's H₃-H₅ fields) and the dentate gyrus molecular layer were determined in ten autopsy-proven AD patients (ages 51–88) and ten age-matched, non-demented control. In addition, RNA contents of pyramidal neurons of the endplate were examined. While there were no differences in RNA contents of astrocytes of either region between AD patients and controls, neuronal RNA was markedly depleted. These data suggest that astrocytes maintain protein synthetic capabilities in AD and that RNA loss is limited to the neuronal compartment.Supported by Grants 1P01-AG05119 and 1P50-AG05144 from the National Institutes of Health and by a Small Research Project Award from the University of Kentucky Medical Center     

Axial tissue diffusion can account for the disparity between current models of hepatic elimination for lipophilic drugs

An assumption of previous models of hepatic elimination is that there is negligible axial diffusion in the liver. We show, by construction of a stochastic model and analysis of published data, that compounds which are readily diffusible and partitioned into hepatocytes may undergo axial tissue diffusion. The compounds most likely to be affected by axial tissue diffusion are the lipophilic drugs for which the cell membranes provide little resistance and which are highly extracted, thereby creating steep concentration gradients along the sinusoid at steady state. This phenomenon greatly modifies the availability of the compound under conditions of altered hepatic blood flow and protein binding. For moderately diffusible compounds, these relationships are similar to those predicted by the simplistic venous-equilibrium model. Hence, the paradoxical ability of the venous-equilibrium model to describe the steady-state kinetics of lipophilic drugs such as lidocaine, meperidine, and propranolol may be finally resolved. The effects of axial tissue diffusion and vascular dispersion on hepatic availability of drugs are compared. Vascular dispersion is of major importance to the availability of poorly diffusible compounds, whereas axial tissue diffusion becomes increasingly dominant for highly diffusive and partitioned substances.This study was supported by the National Health and Medical Research Council of Australia.     

Both phenolic and acyl glucuronidation pathways of diflunisal are impaired in liver cirrhosis

Summary The pharmacokinetics of diflunisal, a salicylate derivative that undergoes phenolic and acyl glucuronidation as well as sulphate conjugation, has been studied after a single oral dose (250 mg) in patients with cirrhosis (n=5) and in healthy controls (n=5).The plasma clearance of total (bound + unbound) diflunisal was 10.2 ml · min^–1 in the control subjects and it was not affected by cirrhosis (10.9 ml · min^–1). The plasma protein binding of diflunisal was significantly reduced in cirrhosis; the percentage of unbound diflunisal in plasma was 0.089 in the controls and 0.147 in the patients with cirrhosis. Plasma clearance of unbound diflunisal was significantly impaired in cirrhosis: 11.51 · min^–1 in control subjects vs 7.41 · min^–1 in cirrhotics.In cirrhotic patients, the unbound partial clearances to the phenolic and acyl glucuronides were both significantly reduced, by approximately 38%. The unbound partial clearance to the sulphate conjugate was not significantly affected by cirrhosis.The results show that both the phenolic and acyl glucuronidation pathways of diflunisal are equally susceptible to the effects of liver cirrhosis.     

Evidence for negative control in protein kinase C substrate specificity

The peptide Leu-Asp-Asp-Ser-Lys-Arg-Val-Ala-Lys-Arg-Lys-Leu-Ile-Glu, which corresponds to sequence 124 to 137 of c-erb-A protein, was synthesized and tested as substrate for protein kinase C (PKC). Although a typical recognition sequence for PKC, consisting of a cluster of basic residues, is found on the C-terminus side of serine, its phosphorylation was totally prevented by the presence of the two acidic residues on the amino-terminus side. Three analogs in which aspartyl residues were successively replaced with alanine were studied and the influence of the acidic side chain in modulating phosphorylation by PKC was thus possible to determine. The results show that the presence of a single aspartyl residue located in positions i-1 or i-2 with respect to the phosphorylable residue can almost totally abolish the positive effect of a highly favorable cluster of basic residues. These observations highlight the role of negative substrate specificity determinants in settling the protein substrate profile of protein kinase C.