甲氨喋呤对胶原诱导性关节炎大鼠滑膜骨桥蛋白和整合素αVβ3表达的影响

目的 探讨甲氨喋呤对胶原诱导性关节炎(collagen-induced arthritis,CIA)大鼠滑膜骨桥蛋白(osteopontin,OPN)及其受体整合素αvβ3表达的影响及其作用机制.方法 建立CIA大鼠模型,分为模型组和甲氨喋呤(MTX)组,后者用MTX进行干预治疗;采用免疫组织化学染色技术检测滑膜组织OPN和整合素αvβ3的表达,ELISA方法检测血清TNF-α水平.结果 ① CIA模型组和MTX组大鼠的滑膜OPN、整合素αvβ3表达均明显高于正常大鼠对照组(均为P<0.01);与模型组比较,MTX组OPN和整合素αVβ3的表达减少(均为P<0.01). ②正常对照组和MTX组大鼠血清TNF-α水平显著性低于模型组大鼠(均为P<0.01).结论 滑膜OPN和整合素αvβ3异常表达在CIA发病中具有重要意义.MTX通过下调滑膜OPN和整合素αvβ3的表达,降低血清TNF-α水平从而达到治疗CIA的作用.     

肾上腺髓质素对豚鼠心室肌细胞L-型钙通道的作用及其信号转导机制

目的: 研究肾上腺髓质素( adrenomedullin, ADM)抑制豚鼠心室肌细胞L-型钙通道的信号转导机制。方法: 应用全细胞膜片钳技术,记录应用ADM(1-100 nmol·L^-1)前后L-型钙电流(I_Ca,L),以及分别记录应用ADM特异性受体拮抗剂ADM_22-52(100 nmol·L^-1)+ADM(100 nmol·L^-1)、蛋白激酶A (PKA) 特异性拮抗剂H-89(10 μmol·L^-1) + ADM(100 nmol·L^-1)、蛋白激酶C (PKC) 特异性拮抗剂PKC_19-36(10 μmol·L^-1)+ADM(100 nmol·L^-1)、PKC特异性激动剂PMA(1 μmol·L^-1)前后I_Ca,L。结果: ADM(1-100 nmol·L^-1)浓度依赖性地抑制豚鼠心室肌细胞I_Ca,L,并可被ADM_22-52(100 nmol·L^-1)完全阻断;H-89(10 μmol·L^-1)对ADM抑制I_Ca,L的作用无影响。PKC_19-36(10 μmol·L^-1)可完全阻断ADM 对I_Ca,L的抑制效应,且PMA(1 μmol·L^-1)可模拟ADM 对I_Ca,L的抑制效应。结论: ADM作用于特异性ADM受体可浓度依赖性地抑制豚鼠心室肌细胞I_Ca,L,此作用有可能与PKC激活相关。     

Comparison of radioimmunoassay and ELISA methods for detection of antibodies to chromatin components

A solid phase radioimmunoassay has been compared with an enzyme-linked immunosorbent assay (ELISA) for efficacy in measuring anti-chromatin antibodies. The low backgrounds achieved with the radioimmunoassay method produced a high signal-to-noise ratio and enabled detection of the human test antiserum at a dilution of 1:102,400. By contrast, the ELISA could detect the same antiserum only at a dilution of 1:3200 and above. The radioimmunoassay was consistently more sensitive than the ELISA for detection of anti-chromatin antibodies in a number of human and mouse sera and ascites fluid containing a monoclonal antibody. Factors affecting sensitivity in both assays are discussed.     

Mechanical properties of abdominal aortic aneurysm wall

There is a need to understand why and where the abdominal aortic aneurysm may rupture. Our goal therefore is to investigate whether the mechanical properties are different in different regions of the aneurysm. Aorta samples from five freshly excised whole aneurysms, > or = 5 cm in diameter, from five patients, average age 71 +/- 10 years, were subjected to uniaxial testing. We report the wall thickness, yield stress and strain, and parameters that describe nonlinear stress-strain curves for the anterior, lateral and posterior regions of the aneurysm. The posterior region was thicker than the anterior region (2.73 +/- 0.46 mm versus 2.09 +/- 0.51 mm). The stress-strain curves were described by sigma = a epsilon(b), where sigma is true stress and epsilon is engineering strain. In the circumferential direction, the wall stiffness increased from posterior to anterior to lateral. In the longitudinal direction, the lateral and anterior regions showed greater wall stiffness than the posterior region. The wall stiffness was greater in the circumferential than longitudinal direction. The anterior region was the weakest, especially in the longitudinal direction (yield stress sigmaY = 0.38 +/- 0.18 N mm(-2)). For a less complex model the aneurysmal wall could be considered orthotropic with sigma = 12.89epsilon(2.92) and 4.95epsilon(2.84) in the circumferential and longitudinal directions. For the isotropic model, sigma =7.89epsilon(2.88). In conclusion, different regions of the aneurysm have different yield stress, yield strains, and other mechanical properties, and this must be considered in understanding where the rupture might occur.     

The transplacental toxicity of methyl mercury to fetal rat liver mitochondria. Morphometric and biochemical studies.

Ultrastructural morphometric and biochemical changes in liver mitochondria of fetal rats whose mothers were exposed to methyl mercury hydroxide in their drinking water at concentrations of 0, 3, 5, or 10 p.p.m. for 4 weeks prior to mating and through day 19 of pregnancy are described. A dose-related decrease in the volume density of mitochondria was observed in fetal hepatocytes of dams given the 5- and 10-p.p.m. dose levels. This finding was associated with decreased mitochondrial protein synthesis, which appeared to result primarily from decreased synthesis of mitochondrial structural proteins. Loss of respiratory control was observed in mitochondria from animals in the 3-p.p.m. dose group whereas state 3 respiration was abolished in animals exposed to the 5- and 10-p.p.m. dose levels. The specific activities of monoamine oxidase and cytochrome oxidase, outer and inner mitochondrial membrane marker enzymes respectively, showed dose-related decreases of up to 62 and 78 per cent of control, respectively. delta-Aminolevulinic acid synthetase, which is loosely bound to the inner mitochondrial membrane, also showed dose-related decreases of up to 68 per cent of control. Malate dehydrogenase, a mitochondrial matrix marker enzyme, showed no change in activity at any dose level tested. These observations were correlated with dose-related tissue concentrations of methyl and inormpaired mitochondrial biogenesis and functional development is one basis for explaining the sensitivity of fetal animals to methyl mercury toxicity.     

Performance characteristics of HIV-1 culture and HIV-1 DNA and RNA amplification assays for early diagnosis of perinatal HIV-1 infection

A plasma HIV-1 RNA amplification assay (RNA assay), a quantitative peripheral blood mononuclear cell (PBMC) microculture (culture), and a PBMC HIV-1 DNA amplification assay (DNA assay) were compared for diagnosis of HIV-1 infection in infants receiving zidovudine in Pediatric AIDS Clinical Trials Group protocol 185; assays were performed for all 24 infected and 100 uninfected infants. HIV-1 infection was defined as >or=2 positive cultures or positive antibody to HIV-1 at >or=18 months. Cultures were performed at birth and 6 and 24 weeks of age; DNA and RNA assays were performed on cryopreserved specimens. The sensitivity of culture and DNA and RNA assays at birth was 20.8%, 10.5%, and 26.7%, respectively. At older ages, sensitivity typically exceeded 80%, remaining highest for the RNA assay (>85%). Assay specificity was >99%. Positive predictive values exceeded 93% for each assay at each age; negative predictive values were highest (>90%) for the RNA assay. At birth (P < 0.005) and age 6 weeks (P < 0.001), a significantly larger proportion of infected infants were identified by means of the RNA assay than by the other assays. The diagnostic performance of the RNA assay matched or exceeded that of culture and the DNA assay. Given that RNA assays require less blood volume and yield rapid results, our study adds to existing data suggesting that RNA assays may be used for early diagnosis of HIV-1 infection in infants.     

Physiological basis of the low calcium response in Yersinia pestis.

It is established that duplication in vitro of that amount of Ca2+ (2.5 mM) and Mg2+ (1.5 mM) present in blood permits vegetative growth of Yersinia pestis with repression of virulence factors encoded by the Lcr plasmid (Lcr+); similar simulation of intracellular fluid (no Ca2+ and 20 mM Mg2+) promotes bacteriostasis with induction of these virulence determinants. However, proliferation of yersiniae in mice occurs primarily within necrotic focal lesions (supplied by Ca(2+)-deficient host cell cytoplasm) within visceral organs rather than in Ca(2+)-sufficient blood. The present study addressed this enigma by defining conditions necessary for achieving vegetative growth of Lcr+ yersiniae at 37 degrees C in simulated intracellular fluid. Maximum optical densities were increased by substitution of K+ for Na+ and elimination of Cl-; the combination of Na+ plus L-glutamate was selectively toxic to Lcr+ cells. This phenomenon was attributed in part to the absence of aspartase in Y. pestis (a lesion known to facilitate massive accumulation of L-aspartate via transamination of the oxalacetate pool by L-glutamate). Replacement of L-glutamate by exogenous L-aspartate or alpha-ketoglutarate reversed this toxicity by favoring retention of oxalacetate. Proliferation of Lcr+ cells in a medium containing K+ and L-aspartate but lacking added Ca2+ and Na+ was markedly enhanced by increasing the concentration of fermentable carbohydrate. Accordingly, in the worst-case scenario (i.e., added Na+, Cl-, and L-glutamate), Lcr+ yersiniae underwent restriction of growth after one doubling, and in the best-case scenario (i.e., added K+ and L-aspartate), the organisms completed more than five doublings, thereby achieving full-scale growth. Both of these Ca(2+)-deficient media promoted maximum induction of Mg(2+)-induced V antigen, a virulence factor encoded by the Lcr plasmid.     

Preservation of 5-hydroxytryptamine1A receptor-G protein interactions in the cerebral cortex of patients with Alzheimer's disease.

The coupling of 5-hydroxytryptamine1A (5-HT1A) receptors to guanine nucleotide binding (G) proteins was investigated in membranes prepared from frontal and parietal cortices of control and Alzheimer's disease brains by characterising the effect of guanosine 5'-[beta gamma-imido]diphosphate (Gpp[NH]p) on [3H]8-hydroxy-2-(di-n-propylamino)-tetralin ([3H]8-OH-DPAT) binding parameters. In the absence of guanine nucleotides, [3H]8-OH-DPAT bound to a single high affinity binding site in all membrane types. The number of [3H]8-OH-DPAT binding sites was significantly decreased in the parietal cortex of Alzheimer's disease samples compared with controls, whereas in the frontal cortex the number of binding sites remained unchanged. Gpp[NH]p reduced the [3H]8-OH-DPAT binding affinity and the number of binding sites to the same degree in both regions in control and Alzheimer's disease cases. [3H]8-OH-DPAT binding was inhibited in a concentration dependent manner with an IC50 value of approximately 1 microM in all cases. These results suggest that the 5-HT1A receptor-G protein complex is functionally intact in these regions in Alzheimer's disease brain.     

B cell chronic lymphocytic leukaemia cells have reduced capacity to upregulate expression of MHC class I in response to interferon-gamma

AIMS: An important consideration in the design of a tumour vaccine is the ability of tumour-specific cytotoxic T lymphocytes (CTL) to recognise unmanipulated tumour cells in vivo. To determine whether B-CLL might use an escape strategy, the current studies compared B-CLL and normal B cell MHC class I expression. METHODS: Flow cytometry, TAP allele PCR and MHC class I PCR were used. RESULTS: While baseline expression of MHC class I did not differ, upregulation of MHC class I expression by B-CLL cells in response to IFN-gamma was reduced. No deletions or mutations of TAP 1 or 2 genes were detected. B-CLL cells upregulated TAP protein expression in response to IFN-gamma. Responsiveness of B-CLL MHC class I mRNA to IFN-gamma was not impaired. CONCLUSIONS: The data suggest that MHC class I molecules might be less stable at the cell surface in B-CLL than normal B cells, as a result of the described release of beta(2)m and beta(2)m-free class I heavy chains from the membrane. This relative MHC class I expression defect of B-CLL cells may reduce their susceptibility to CTL lysis in response to immunotherapeutic approaches.     

人核受体hLRH-1不同变异体在多种肿瘤组织和细胞系中的表达

目的:初步分析人核受体hLRH-1不同变异体在多种肿瘤组织和细胞系中的表达。方法:常规RT-PCR法分析hLRH-1v1和hLRH-1在肝癌等多种肿瘤组织和HepG2等肿瘤细胞系中的表达情况，实时定量PCR法分析hLRH-1v1，hLRH-1和hOct4在HepG2，SMMC-7721和BEL-7402三株肝癌细胞系的表达水平。结果:hLRH-1v1的表达见于所有检测的12种类型肿瘤和8种恶性肿瘤细胞系，hLRH-1的表达则仅在12种类型肿瘤中的6种肿瘤组织中可检测到，但8种恶性肿瘤细胞系均为hLRH-1阳性表达；在HepG2，SMMC-7721和BEL-7402三株肝癌细胞系中hLRH-1v1和hOct4的表达呈正相关。结论:hLRH-1v1在肿瘤中广泛表达，可能在维持肿瘤干细胞的自我更新中发挥重要作用。