Translocation of Enterococcus faecalis strains across a monolayer of polarized human enterocyte-like T84 cells

We used a two-chamber system to study transcytosis of Enterococcus faecalis across monolayers of human colon carcinoma-derived T84 cells, which show structural resemblance to the native intestine. Among 16 E. faecalis isolates from different sources, the well-characterized strain OG1RF and 8 other isolates (2 endocarditis isolates, 1 urine isolate, and all 5 fecal isolates) showed translocation in this assay, while 6 clinical isolates (3 endocarditis and 3 urine isolates), the recipient strain JH2-2, and the control, Escherichia coli DH5alpha, had no detectable translocation. Of two OG1RF mutants involving the previously studied epa (enterococcal polysaccharide antigen) gene cluster, known to be needed for virulence and resistance to killing by polymorphonuclear leukocytes, one epa mutant (TX5179) was unable to translocate, while TX5180, with an epa disruption farther downstream, showed a moderate decrease in translocation relative to that of the wild-type strain OG1RF (P < 0.01), indicating that the epa gene cluster is important for translocation across a T84 monolayer. This observation was confirmed by complementation of the epa mutant (TX5179) with epa genes and restoration of its translocation ability. In conclusion, we have demonstrated translocation of at least some strains of E. faecalis across T84 monolayers, although strains differ considerably in this ability, and we have demonstrated that epa mutations can cause marked changes in successful translocation. These results suggest that this model may be a useful in vitro system for studying the process of translocation from the intestinal tract.     

Detection of reactive oxygen species (ROS) and apoptosis in human fragmented embryos

In human in-vitro fertilization (IVF)-embryo transfer, the in-vitro culture environment differs from in-vivo conditions in that the oxygen concentration is higher, and in such conditions the mouse embryos show a higher concentration of reactive oxygen species (ROS) in simple culture media. ROS are believed to cause damage to cell membranes and DNA fragmentation in somatic cells. This study was conducted to ascertain the level of H2O2 concentration within embryos and the morphological features of cell damage induced by H2O2. A total of 62 human oocytes and embryos (31 fragmented, 15 non-fragmented embryos, 16 unfertilized oocytes) was obtained from the IVF-embryo transfer programme. The relative intensity of H2O2 concentrations within embryos was measured using 2',7'-dichlorodihydrofluorescein diacetate by Quanti cell 500 fluorescence imaging and DNA fragmentation was observed with transmission electron microscopy and an in-situ apoptosis detection kit. The H2O2 concentrations were significantly higher in fragmented embryos (72.21 +/- 9.62, mean +/- SEM) compared to non-fragmented embryos (31.30 +/- 3.50, P < 0.05) and unfertilized oocytes (30.75 +/- 2.67, P < 0.05). Apoptosis was observed only in fragmented embryos, and was absent in non-fragmented embryos. Electron microscopic findings confirmed apoptotic bodies and cytoplasmic condensation in the fragmented blastomeres. We conclude that there is a direct relationship between increased H2O2 concentration and apoptosis, and that further studies should be undertaken to confirm these findings.      

Cultivation and characterisation of avian lymphocytes with natural killer cell activity

Conditioned medium (CM), as a presumed source of lymphokines including interleukin-2, was prepared from chicken spleen cell cultures stimulated with concanavalin A (con A). When CM was used to cultivate spleen cells from 6- to 8-week-old P-2 chickens, eight of nine spleens yielded cell lines which grew continuously for at least 50 days. Six of the cultures were tested for natural killer (NK) cell activity against LSCC-RP9, a lymphoblastoid cell known to be susceptible to NK cells, and against several Marek's disease lymphoblastoid cell lines (MDCC-CU2, -CU36 and -MSB-1). All six cultures lysed the RP9 cells in a chromium release assay with high levels of specific release (30 to 50%) at effector cell to target cell ratios of 5:1 or 10:1. CU2 and CU36, which are NK-cell resistant, were not lysed, while there was a low level of specific activity against MSB-1. The cells were characterised for surface and internal antigens with monoclonal antibodies and were negative for thymocyte antigen, IgM, a T cell antigen also present on granulocytes and red blood cells, and two antigens found in macrophages. Two of the six lines examined had a low number of cells expressing la antigen, while the other four were negative. An antigen present on circulating T cells and a macrophage (sub)population was present on all lines. The majority of the cells had the morphological appearance of mammalian large granular lymphocytes (LGL) with a rather small kidney-shaped nucleus. Granules and vacuoles were present in the cytoplasm. However, there was a variable percentage of lymphoblastoid (LB) cells also present. Cell lines established with CM from con A-stimulated spleen lymphocytes in other studies were also shown to have high levels of NK activity regardless of the relative proportions of cells with the morphological appearance of LGLs or LBs, or the relative frequency of expression of the two T cell markers or la antigen, all of which varied markedly.     

Cloning and expression of the gene involved in Sanfilippo B syndrome (mucopolysaccharidosis III B)

Sanfilippo B syndrome is caused by a deficiency of alpha-N- acetylglucosaminidase, a lysosomal enzyme involved in the degradation of heparan sulphate. Accumulation of the substrate in lysosomes results in degeneration of the central nervous system with progressive dementia often combined with hyperactivity and aggressive behaviour. In order to clone the deficient gene, we purified the enzyme from human placenta and obtained amino acid sequence information. Alignment of one of the CNBr generated internal peptides to sequence from the database revealed the chromosomal location of the gene in the 5' upstream flanking region of the gene for 17-beta-hydroxysteroid-dehydrogenase at 17q21.1. The available DNA sequence was used to clone the cDNA coding for alpha-N- acetylglucosaminidase and analyse its gene structure. The gene is fully contained in the 5' upstream flanking region of the gene for 17-beta- hydroxysteroid-dehydrogenase and interrupted by five introns. The cDNA clone has a length of 2575 bp and encodes a protein of 743 amino acids. Chinese hamster ovary cells transfected with the cDNA construct show alpha-N-acetylglucosaminidase activity about 17-fold over background. This will allow correction studies with NAG deficient Sanfilippo B cell lines and facilitate the development of enzyme replacement therapy for these patients.      

Interleukin-4 receptor alpha chain and STAT6 signaling inhibit gamma interferon but not Th2 cytokine expression within schistosome granulomas

Compared to wild-type (WT) mice, schistosome granulomas in Stat6 knockout (KO) mice lacked eosinophils and had Th1 features. Interleukin-4 (IL-4) acts through Stat6 in assisting Th2 cell development. The importance of Stat6 for Th2-cell development within schistosome granulomas had not been explored. Therefore we studied gamma interferon (IFN-gamma), IL-4, and IL-5 production in granulomas from Stat6 KO and WT mice. Dispersed granuloma cells from Stat6 KO and WT mice made similar amounts of IL-4 and IL-5. Only Stat6 KO granuloma cells released IFN-gamma. Granuloma T cells contained most of the IL-4, IL-5, and IFN-gamma mRNA and secreted these cytokines. In Stat6 KO mice, 16.6% of the granuloma cells were CD4(+). Of these, 10.7% stained for IFN-gamma and/or IL-4 by intracytoplasmic flow analysis. Few CD4(-) T cells stained positively. The IL-4-producing T cells did not stain for DX5 or with labeled alpha-GalCer CD1d tetramer, suggesting an absence of NK T cells. Thus, conventional Th cells in Stat6 KO granulomas produce IFN-gamma and Th2 cytokines. Stat6 limits IFN-gamma production but is unnecessary for Th2-cell development or localization within the granuloma.     

玻璃体切除术治疗外伤性眼内炎62例

0　引言　外伤性眼内炎是眼外伤的常见并发症 ,对视力及眼球威协极大 .由于病原体随致伤物直接进入眼内 ,在眼内和玻璃体繁殖 ,产生剧烈炎症反应 ,对眼内组织造成严重破坏 .如不及时而有效的控制 ,终使视力丧失 ,眼球萎缩 .近年随着现代玻璃体手术的发展 ,外伤性眼内炎的治愈率明显提高 .现将我科收治的 6 2例报告如下 .1　对象和方法1 .1　对象 1 996 - 0 6 / 1 998- 0 6我院连续收治 6 2例外伤性眼内炎 ,其中男 5 0例 ,女 1 2例 ,年龄 2～ 45岁 ,平均 1 6 .5岁 .1 4岁以下儿童 38例 ,占 6 1 .3% .以致伤原因分类 :角膜穿通伤30例 ,巩膜穿…     

CLINICO-EPIDEMIOLOGICAL ALGORITHM FOR PREDICTING SYSTEMIC ARTERIAL HYPERTENSION AT HIGH ALTITUDE THROUGH MATHEMATICAL MODELLING

A population based hybrid design combining element of cohort and cross-sectional approach was used to develop a simple clinical algorithm to predict individual probability of developing hypertension (systolic BP > 140 mm Hg and/or diastolic BP > 90 mmHg). 3615 soldiers initially normotensive at the time of induction into high altitude, were studied by systematic random sampling. Multiple logistic regression analysis showed a high significant association between hypertension and age, body mass index (BMI), tobacco smoking and alcohol consumption. Using the constant/coefficient values obtained from the logistic model and the receiver operating characteristics (ROC) curve analysis, the following predictive rule was developed – To the age in years, add (BMIx 3.86); also add 5.53 if he is a smoker; and add 19.81 if he consumes alcohol. If the total exceeds 142, the individual is at high risk of developing hypertension. This algorithm carries a sensitivity of 68.2% and specificity of 78.5%.KEY WORDS: Hypertension, High altitude     

Risk factors for basal cell carcinoma of the skin in men: results from the health professionals follow-up study.

The authors examined the relation of constitutional factors and sun exposure to risk of basal cell carcinoma of the skin (BCC) in a prospective cohort of 44,591 predominantly Caucasian US male health professionals, 40-75 years of age and free of cancer at enrollment in 1986. During 8 years of follow-up, 3,273 cases of self-reported BCC were documented. The following variables were each associated with an elevated risk of BCC: having red hair; green, hazel, or blue eyes; a tendency to sunburn; and north European ancestry. The lifetime number of blistering sunburns was also positively associated with BCC risk (p trend < 0.0001). Compared with men who as teenagers had been outside less than once a week, men who had been outside weekly (relative risk (RR) = 1.30; 95% confidence interval (CI): 1.14, 1.47) and daily (RR = 1.42; 95% CI: 1.24, 1.63) had an elevated risk of BCC. Living in a region of residence with high solar radiation as an adult was also associated with an increased risk of BCC (RR = 1.48; 95% CI: 1.36, 1.60), whereas living in such a region only in childhood did not increase BCC risk. These results confirm the role of constitutional factors and suggest that adult sun exposure increases BCC risk.     

Seroconversion rates in healthcare workers using a latex agglutination assay after varicella virus vaccination.

OBJECTIVES: To determine the seroconversion rate after varicella immunization of healthcare workers (HCWs) and the effect of seroconversion rate on current cost-based recommendations for universal vaccination. METHODS: A voluntary vaccination program for HCWs was performed at a tertiary-care cancer center in New York City. A commercial latex agglutination assay was used to test postvaccination antibody response. Costs for vaccination and postvaccination serological testing were compared to potential costs of postexposure employee furloughs. RESULTS: Of 263 seronegative HCWs, 96 (36.5%) began the vaccine program. Thirty-nine HCWs received only one dose of vaccine. Seven returned for follow-up antibody testing, of whom 4 were seropositive. Of the 57 HCWs who received two doses, 38 returned for follow-up serology. Thirty-one (81.6%) HCWs were seropositive for varicella-zoster virus antibodies, and seven HCWs (18.4%) remained seronegative. Total cost of vaccination for all 263 seronegative HCWs was estimated and compared to the cost of varicella-related furloughs at our institution. CONCLUSIONS: We found a considerably lower rate of vaccine-induced seroconversion at our hospital compared to that of the published literature. Despite this finding, universal varicella vaccination remained an extremely cost-effective alternative to the furloughing of exposed, seronegative HCWs. Projected hospital savings exceeded $53,000 in the first year after vaccination alone.