Induction of chromosomal instability in colonic cells by the human polyomavirus JC virus

Most colorectal cancers display chromosomal instability, which is characterized by gross chromosomal rearrangements, loss of heterozygosity and aneuploidy. We have previously demonstrated a link between JC virus strains Mad-1 and Delta98 and colorectal cancer. Others have also associated the virus to the induction of colon cancer and aneuploid brain tumors by producing a highly tumorigenic protein named T antigen (TAg), which binds to beta-catenin and inactivates key proteins such as p53. The aim is to demonstrate that JC virus is capable of inducing chromosomal instability in colonic cells. We used the human colon cancer cell line RKO as a model. The cell line has wild-type p53, wild-type beta-catenin and APC and is diploid. Neuroblastoma JCI cells, which are infected with the virus, VA13 fibroblasts, which are transformed by the SV40 TAg, were used as positive controls. HCT116, which has mutated beta-catenin, and SW480, which is a model of CIN, were also used as controls. The genomes of the Mad-1 and Delta98 strains were transfected into cells. As negative controls we used pUC or no plasmids. Cells were collected at 0, 7, 14, and 21 days after transfection. PCR was used for the detection of TAg and the regulatory region DNA sequences at different time frames and Southern blot of whole genomic extracts for viral DNA integration into the host genome. Immunofluorescence and Western blot were performed for TAg, viral capsid proteins, and nuclear beta-catenin expressions, whereas coimmunoprecipitation was used to detect protein interactions. Karyotype analysis and electron microscopy were performed to seek chromosomal instability and cell abnormalities, respectively. Retention of viral sequences was observed for Mad-1- and Delta98-transfected RKO cells at all time frames with PCR only, whereas Southern blot analysis showed nonintegrated sequences at T7 alone. TAg and capsid protein expressions, as well as increased p53 and nuclear beta-catenin, were observed between T0 and T7 for Mad-1 and Delta98 alone. Also, interaction between TAg and both p53 and beta-catenin was also observed between T0 and T7. Chromosomal instability, characterized by chromosomal breakage, dicentric chromosomes, and increasing ploidy, was observed at all time frames for Mad-1 and Delta98, as well as cell abnormalities. In conclusion, we demonstrate that JC virus Mad-1 and Delta98 are able to induce chromosomal instability in colonic cells with a hit and run mechanism that involves an early interaction with beta-catenin and p53.     

Clinical evaluation of a new recombinant antigen-based cytomegalovirus immunoglobulin M immunoassay in liver transplant recipients

BACKGROUND: Human cytomegalovirus (CMV) is a significant cause of morbidity and mortality among transplant recipients. Monitoring transplant recipients by CMV IgM serology has been questioned by several studies due to the reported insensitivity of serologic tests relative to antigen detection methods. METHODS: In this retrospective study, we have evaluated the performance of the new recombinant antigen-based Abbott AxSYM CMV IgM assay and compared it with CMV culture technique in a cohort of 40 liver transplant recipients who did not receive antiviral prophylaxis. RESULTS: The sensitivity, specificity, and positive and negative predictive values for detection of CMV disease by the AxSYM CMV IgM assay were 90.0%, 60.0%, 69.2%, and 85.7%, respectively, and by culture the values were 100%, 55.0%, 69.0%, and 100%, respectively. Detection of CMV IgM occurred before or at the time of CMV disease in only R+ recipients. CONCLUSION: Although this assay is a sensitive test for CMV-specific IgM, detection of CMV IgM preceded detection of virus by culture in patients only when the liver transplant recipient was CMV immune before transplantation (R+).     

Distinct epitopes on formiminotransferase cyclodeaminase induce autoimmune liver cytosol antibody type 1

Liver cytosol antibody type 1 (LC1) is regarded as a serologic marker of type 2 autoimmune hepatitis, in addition to liver kidney microsomal antibody type 1. Among 38 patients with type 2 autoimmune hepatitis, 23 were positive for LC1 antibodies. The antigen recognized by LC1 has been identified as a liver-specific 58-kd metabolic enzyme named formiminotransferase cyclodeaminase (FTCD). All 23 LC1-positive sera immunoprecipitated rat FTCD, and 22 gave an identity reaction with rat FTCD by immunodiffusion. No reaction was observed with sera from 10 patients with type 1 autoimmune hepatitis, 10 with primary biliary cirrhosis, 10 with chronic hepatitis C, and 10 healthy controls. By Western immunoblotting all 23 LC1-positive sera and all the controls tested negative, suggesting that all the antigenic epitopes were destroyed by denaturation. FTCD is a bifunctional protein composed of distinct globular FT and CD domains connected by a short linker. To identify epitopes that trigger the LC1 autoimmune response, we tested LC1 antibodies against FTCD constructs encoding the N-terminal FT domain (amino acids 1-339), or the C-terminal CD domain (amino acids 332-541). Of 20 sera positive against full-length FTCD, 8 (40%) recognized the FT domain and the CD domain, 7 (35%) recognized only the FT domain, and 5 (25%) did not recognize either construct. No sera reacted with only the CD domain. These data indicate that multiple regions of FTCD trigger the LC1 autoimmune response, and that LC1 reactivity is mainly directed to conformation-sensitive epitopes located in the FT region of FTCD.     

pUL25 immunolocalization in human cytomegalovirus-infected and gene-transfected cells

Summary.  By means of confocal and electron microscope immunocytochemistry we have studied the localization of a recently described structural protein (pUL25) of human cytomegalovirus, in both infected cells and in cells transiently transfected with UL25. pUL25 localization in infected cells was observed in typical cytoplasmic structures characterized by a very electrondense texture previously reported to accumulate other tegument proteins. At the virion level pUL25 seems to localize at the interface between the tegument and the capsid of both intracytoplasmic and extracellular virions. In UL-25-transfected cells, pUL25 has been found in characteristic para-crystalline cytoplasmic aggregates, suggesting its intrinsic ability to aggregate in a regular subunit pattern. Received August 6, 1999/Accepted November 3, 1999     

Prenatal diagnosis of symptomatic congenital cytomegalovirus infection

OBJECTIVE: The aim of this study was to evaluate whether the amniotic viral load of mothers with primary cytomegalovirus infection correlate with fetal or neonatal outcomes. STUDY DESIGN: Sixty-eight of 138 pregnant women with primary infection defined by immunoglobulin G seroconversion or the presence of immunoglobulin M with low immunoglobulin G avidity accepted amniocentesis. Polymerase chain reaction and quantitative polymerase chain reaction were used to detect amniotic fluid cytomegalovirus. Cytomegalovirus infection in neonates was determined by means of urinary viral isolation during the first week after birth or the histologic examination of tissue from aborted fetuses. RESULTS: Cytomegalovirus infection was found in 16 fetuses and neonates (23%), 5 of whom had symptoms. Quantitative polymerase chain reaction showed that the presence of >/=10(3) genome equivalents predicted mother-child infection with 100% probability; >/=10(5) genome equivalents predicted the development of a symptomatic infection. CONCLUSION: Fewer than expected cytomegalovirus-infected fetuses are at risk for development of cytomegaloviral disease, and this fact may be useful in counseling pregnant women with primary cytomegalovirus infection.     

Chronic Arsenic Poisoning From Burning High-Arsenic-Containing Coal In Guizhou, China

Optimization of a previously disclosed sorbitol dehydrogenase inhibitor (SDI, II) for potency and duration of action was achieved by replacing the metabolically labile N,N-dimethylsulfamoyl group with a variety of heterocycles. Specifically, this effort led to a series of novel, in vitro potent SDIs with longer serum half-lives and acceptable in vivo activity in acutely diabetic rats (e.g., 62, 67, and 69). However, the desired in vivo potency in chronically diabetic rats, ED90 less than or equal to 5 mg/kg/day, was achieved only through further modification of the piperazine linker. Several members of this family, including 86, showed better than the targeted potency with ED90 values of 1-2 mg/kg/day. Compound 86 was further profiled and found to be a selective inhibitor of sorbitol dehydrogenase, with excellent pharmacodynamic/pharmacokinetic properties, demonstrating normalization of sciatic nerve fructose in a chronically diabetic rat model for approximately 17 h, when administered orally at a single dose of 2 mg/kg/day.     

冠状动脉粥样硬化ADAMTS7新位点的发现和冠状动脉粥样硬化中ABO位点与心肌梗死的相关性:两项全基因组关联研究

<正>背景:该研究旨在评价在现有的冠状动脉粥样硬化的病变中,遗传因素对粥样硬化斑块进展及特异性心肌梗死是否存在显著作用。方法:对欧洲后裔参与者冠状动脉造影表型进行了两项全基因组关联研究(GWAS),为寻找冠状动脉疾病(CAD)易感性基因位点,研究比较了有异常(n=12393)和无异常的(对照组n=7383)个体;为寻找心肌梗死易感性基因位点,也同时比较了造影证实存在CAD并且有心肌梗死的个体(n=5783)与虽有CAD但无心肌梗死的个体(n=3644)。     

Naphthalimido derivatives as antifolate thymidylate synthase inhibitors

A new series of N-(substituted)benzyl-1,8-naphthalimides 4, structurally related to the previously reported thymidylate synthase (TS) inhibitor naphthaleins 3, were synthesized and compounds tested for their inhibition of several species of TS. Moreover, their in vitro cytotoxicity together with antimycotic and antibacterial properties were assayed. While no activity was detected in the antibacterial tests, the m-nitro (4ae) and the p-nitro (4af) derivatives were found able to partially inhibit TS at low micromolar concentrations. Introduction of nitro or (substituted)-amino groups in position 4 of the naphthalic ring always led to less active compounds.