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Biomedical subjects

C Y Ou

Publications and source records attributed to C Y Ou.

At least 55 records · Page 3Linked to original sources
PubMed1992

Zidovudine-interferon-alpha combination therapy in patients with advanced human immunodeficiency virus type 1 infection: biphasic response of p24 antigen and quantitative polymerase chain reaction.

In an open-label dose-ranging pilot trial, 13 homosexual men with human immunodeficiency virus type 1 (HIV-1) p24 antigenemia after at least 6 weeks of zidovudine monotherapy were continued on zidovudine and given interferon-alpha, 1.25-7.5 x 10(6) units/m2 subcutaneously three times/week. Plasma p24 antigen levels demonstrated a biphasic response, falling initially in 11 patients by a mean of 50% (95% confidence interval, 36%-64%; P = .001) at a median of 11 weeks, but rising steadily thereafter (P = .001). CD4+ cell counts fell by a mean of 7.1 cells/mm3/week (P = .01). Higher initial CD4+ counts predicted greater p24 antigen reductions. At higher interferon doses no greater reductions in p24 antigen occurred, but side effects were more severe and CD4+ lymphocyte counts fell faster. Polymerase chain reaction quantification of HIV-1 DNA in 3 patients showed a biphasic pattern paralleling the p24 antigen response. In sum, although evidence of short-term effects was found, the combination showed no evidence of lasting antiviral activity beyond that achieved with zidovudine alone in patients with advanced HIV-1 infection.

AIDS-Related Complex↗
PubMed1992

Thermostable, salt tolerant, wide pH range novel chitobiase from Vibrio parahemolyticus: isolation, characterization, molecular cloning, and expression.

A chitobiase gene from Vibrio parahemolyticus was cloned into plasmid pUC18 in Escherichia coli strain DH5 alpha. The plasmid construct, pC120, contained a 6.4 kb Vibrio DNA insert. The recombinant gene expressed chitobiase [EC 3.2.1.30] activity similar to that found in the native Vibrio. The enzyme was purified by ion exchange, hydroxylapatite and gel permeation chromatographies, and exhibited an apparent molecular weight of 80 kDa on SDS-polyacrylamide gel electrophoresis. Chitobiose and 6 more substrates, including beta-N-acetyl galactosamine glycosides, were hydrolyzed by the recombinant chitobiase, indicating its putative classification as an hexosaminidase [EC 3.2.1.52]. The enzyme was resistant to denaturation by 2 M NaCl, thermostable at 45 degrees C and active over a very unusual (for glycosyl hydrolases) pH range, from 4 to 10. The purified cloned chitobiase gave 4 closely focussed bands on an isoelectric focusing gel, at pH 4 to 6.5. The N-terminal 43 amino acid sequence shows no homology with other proteins in commercial databanks or in the literature, and from its N-terminal sequence, appears to be a novel protein, unrelated in sequence to chitobiases from other Vibrios reported and unrelated to hexosaminidases from other organisms.

Acetylglucosaminidase↗
PubMed1991

Human chromosome-dependent and -independent pathways for HIV-2 trans-activation.

Human immunodeficiency virus (HIV types 1 and 2) replication is controlled by the interaction of viral-encoded regulatory proteins and host cellular proteins with the viral long terminal repeat (LTR). The presence of HIV-1 and HIV-2 trans-activator proteins, tat1 and tat2, respectively, greatly increases viral gene expression from their homologous LTRs. It is unclear if the cellular factors that support tat1-directed trans-activation of the HIV-1 LTR are the same for tat2 trans-activation of the HIV-2 LTR. Human-Chinese hamster ovary hybrid cell clones were used to probe for human chromosomes involved in regulating HIV-1 and HIV-2 tat-directed transactivation. DNA transfection experiments showed that the presence of human chromosome 12 in human-hamster hybrid clones was necessary for high-level tat-directed trans-activation of the HIV-1 and -2 LTR. Cross-trans-activation of the HIV-2 LTR by tat1 was found to be chromosome 12 independent. In addition, chromosome 12 did not support trans-activation of another human retrovirus (human T-cell leukemia virus type I). Our results suggest that HIV-1 and -2 have evolved to employ a cellular pathway(s) encoded on human chromosome 12 for supporting homologous tat-directed trans-activation. Trans-activation of the HIV-2 LTR by tat1 in chromosome 12-minus cells suggests that multiple cellular pathways can be recruited to trans-activate the HIV-2 LTR and that these pathways may have been important in an HIV-like progenitor virus.

Animals↗
PubMed1991

Identification of toxigenic Clostridium difficile by the polymerase chain reaction.

Toxigenic strains of Clostridium difficile are causative agents of pseudomembranous colitis and antimicrobial agent-associated diarrhea and colitis. The toxigenicity is routinely assayed by using highly sensitive cell cultures. We used a simple and rapid polymerase chain reaction (PCR) assay to differentiate toxigenic and nontoxigenic strains of C. difficile. Two sets of oligonucleotide primer pairs derived from nonrepeating sequences of the toxin A gene were used to amplify 546- and 252-bp DNA fragments. A primer pair derived from repeating sequences of the toxin A gene was used to amplify a 1,266-bp DNA product. Amplified products were visualized by polyacrylamide gel electrophoresis followed by ethidium bromide staining. All 35 cytotoxic strains of C. difficile tested generated the expected amplified DNA. In contrast, none of the 26 noncytotoxic strains tested gave positive results. Although the toxins of C. difficile have been demonstrated to cross-react serologically with the toxins of Clostridium sordellii, we did not detect any amplified DNA in two cytotoxic strains or seven noncytotoxic strains of C. sordellii. PCR was negative in all 30 strains of 20 other Clostridium species. Southern hybridization of HindIII-digested genomic DNA by use of subgenomic probes showed a single hybridization band in toxigenic strains but not in nontoxigenic strains. PCR appears to be a sensitive and specific assay for the rapid identification of toxigenic C. difficile. Nontoxigenic C. difficile appeared to lack the C. difficile toxin A gene.

Bacterial Proteins↗
PubMed1991

Use of UV irradiation to reduce false positivity in polymerase chain reaction.

UV irradiation provides a simple and efficient way to minimize contamination or false positivity which often occurs in laboratories performing routine PCR tests. Here, we characterize several parameters of the effect of UV irradiation on DNA template, primers, deoxynucleoside triphosphate and Taq polymerase. UV irradiation of DNA results in the formation of pyrimidine dimers and thus prevents them from being effective templates in subsequent PCR. Reduction of the HIV DNA templates in polypropylene microcentrifuge tubes by more than 1000-fold can be achieved by UV irradiation. The sensitivity of the primers is sequence- and concentration-dependent. Oligonucleotides with neighboring thymine bases are more susceptible to UV than those without. Taq polymerase is highly UV sensitive, whereas deoxynucleotide triphosphate is relatively UV resistant.

Base Sequence↗
PubMed1991

A prospective study of infants of human immunodeficiency virus seropositive and seronegative women with a history of intravenous drug use or of intravenous drug-using sex partners, in the Bronx, New York City.

A prospective study was conducted in the Bronx, New York, of 70 infants of human immunodeficiency virus (HIV)-infected (n = 33) and uninfected (n = 37) mothers who had a history of intravenous drug use or of intravenous drug-using sex partners. Infants were observed from birth to a median age of 23 months (range 3 to 54 months). HIV infection was confirmed in seven infants (21%) of seropositive mothers; six developed HIV disease, with symptoms observed in the first year. Of these, three died (3, 9, and 36 months) of HIV-related causes; 3 of 4 survivors were greater than 25 months of age. HIV symptoms preceded or were concurrent with abnormalities in T-lymphocyte subsets; postneonatal polymerase chain reaction confirmed HIV infection in five infants with symptoms and one without symptoms. Among infants of seropositive mothers, seven without laboratory evidence of HIV (including polymerase chain reaction) had findings suggestive of HIV infection, including persistent generalized lymphadenopathy, hepatosplenomegaly, oral candidiasis, parotitis, and inverted T-lymphocyte ratios. These findings were not observed in infants of seronegative mothers. Although the presence of HIV proviral sequences was associated with HIV disease, the observation of indeterminate symptoms in at-risk infants indicates the importance of long-term clinical follow-up to exclude HIV infection. Disease manifestations in comparable infants of seronegative mothers are important for assessment of the impact of maternal drug use, development of specific clinical criteria for early diagnosis of HIV and eligibility for antiretroviral therapy.

Acquired Immunodeficiency Syndrome↗
PubMed1990

Rapid and quantitative detection of enzymatically amplified HIV-1 DNA using chemiluminescent oligonucleotide probes.

A hybridization protection assay (HPA) that uses acridinium ester (AE) labeled oligonucleotide probes which are specific for a conserved gag gene region of human immunodeficiency virus type 1 (HIV-1) was developed to measure the amount of HIV-1 nucleic acid. Hybridization of the single-stranded probes with their target HIV-1 sequences protected the chemiluminescent AE group from subsequent alkaline hydrolysis. The chemiluminescence from the residual AE could be easily quantitated in a luminometer. The entire process comprising template dissociation, hybridization, alkaline hydrolysis, and chemiluminescence measurement can be completed in less than one hour and does not require the separation of hybridized probe from unhybridized probe. We demonstrated that HPA could quantitatively measure the amount of DNA amplified by polymerase chain reaction. A comparative study using amplified DNA from the peripheral blood mononuclear cells (PBMC) of HIV seropositive and seronegative persons showed that HPA was as sensitive as the previous methods using 32P-labeled DNA probes.

Acridines↗
PubMed1990

Concordance of polymerase chain reaction with human immunodeficiency virus antibody detection.

To evaluate the correlation of detection of human immunodeficiency virus (HIV) by polymerase chain reaction (PCR) with detection of HIV antibody, 271 simultaneous serum and peripheral blood mononuclear cell samples were examined from 242 persons whose activities placed them at increased risk for HIV infection: 142 from homosexual men, 86 from hemophilic men, and 43 from heterosexual partners of HIV-infected persons. PCR was performed using the gag region primer pair SK38/39 and the env region primer pairs SK68/69 and CO71/72. Amplified HIV DNA was detected using specific oligomer probes. Of 63 HIV antibody-positive samples, 58 (92%) had HIV DNA by PCR. Of 208 HIV antibody-negative samples, 7 (3.4%) had HIV DNA by PCR. On follow-up, 4 of the latter persons were seropositive when next tested; 2 were well and antibody- and PCR-negative; 1 had died of a stroke before retesting. Thus, PCR detects HIV in most antibody-positive persons; detection is increased by use of multiple primer pairs. PCR-positive antibody-negative specimens may indicate HIV infection in which antibody has not yet developed or may be false-positive PCR results. When PCR is discordant with HIV antibody, testing of additional specimens and clinical follow-up are necessary to assess HIV infection status.

Bisexuality↗
PubMed1990

Analysis of lymphocytes, monocytes, and neutrophils from human immunodeficiency virus (HIV)-infected persons for HIV DNA.

The human immunodeficiency virus (HIV) infects and depletes or alters the function of cells involved in immune responsiveness. While both T helper lymphocytes and monocyte/macrophages can be infected via cell-surface CD4 in vitro, previous studies showed that few blood cells express HIV RNA in vivo. This study used DNA amplification to determine the levels of HIV DNA in purified lymphocytes, monocytes, and neutrophils from HIV-infected asymptomatic individuals and persons with AIDS. The average numbers of HIV DNA copies in lymphocytes from AIDS patients and asymptomatic individuals were similar (approximately 100-140 copies/150,000 cells). However, when expressed on the basis of numbers of CD4+ T cells, AIDS patients' cells contained approximately 2.5 times more HIV DNA. While HIV DNA was present in lymphocytes from all 27 subjects, little or no HIV DNA was observed in monocytes or neutrophils. Only 1 asymptomatic person contained levels of HIV DNA in monocytes (125 proviral copies/150,000 cells) that were comparable to levels expressed in lymphocytes (160/150,000). While expression of monocyte HIV DNA in this person was persistent over at least 8 months, it was not observed in neutrophils, suggesting that monocyte HIV DNA did not originate in myeloid precursors. This study shows that in AIDS or asymptomatic HIV infection, lymphocytes are the predominant infected cell found in blood.

Acquired Immunodeficiency Syndrome↗
PubMed1989

Human chromosome 12 is required for elevated HIV-1 expression in human-hamster hybrid cells.

Host cell factors act together with regulatory genes of the human immunodeficiency virus (HIV) to control virus production. Human-Chinese hamster ovary hybrid cell clones were used to probe for human chromosomes involved in regulating HIV gene expression. DNA transfection experiments showed that 4 of 18 clones had high levels of HIV gene expression measured by both extracellular virus production and transactivation of the HIV long terminal repeat in the presence of the trans-activator (tat) gene. Karyotype analyses revealed a 94% concordance (17/18) between human chromosome 12 and HIV gene expression. Other chromosomes had an 11 to 72% concordance with virus production.

Animals↗
PubMed1989

Duration of human immunodeficiency virus infection before detection of antibody.

To estimate the duration and frequency of the period of HIV infection without detectable antibody, modelling techniques were applied to results of detection of HIV DNA by means of the polymerase chain reaction (PCR) and to data from cases in published reports. PCR was carried out with gag and env region primers on samples from 27 homosexual and 12 haemophilic men for whom stored samples were available from before and after seroconversion; serum was also tested for p24 antigen by antigen-capture enzyme immunoassay. HIV DNA was detectable before seroconversion in 4 men; in all 4 PCR was positive only in the seronegative sample taken closest to the time of seroconversion. In 3 men antigen was detected before seroconversion; in each case HIV DNA was also detected. By a Markov model, the time from infection with HIV (as assessed by detection of HIV DNA) to first detection of HIV antibody was estimated to be 2.4 (SE 2.1) months for the median individual. Modelling of cases of HIV infection with known exposure in published reports gave a median estimate of 2.1 (0.1) months from exposure to antibody detection, and 95% of cases would be expected to seroconvert within 5.8 (0.6) months. HIV infection for longer than 6 months without detectable antibody seems uncommon.

Acquired Immunodeficiency Syndrome↗
PubMed1989

Use of the polymerase chain reaction for early detection of the proviral sequences of human immunodeficiency virus in infants born to seropositive mothers. New York City Collaborative Study of Maternal HIV Transmission and Montefiore Medical Center HIV Perinatal Transmission Study Group.

The early diagnosis of infection with human immunodeficiency virus (HIV) in infants born to infected mothers is essential for early treatment, but current tests cannot detect HIV infection in newborns because of the presence of maternal antibodies. We used the polymerase chain reaction, a new technique that amplifies proviral sequences of HIV within DNA, to detect HIV infection in peripheral-blood mononuclear cells obtained from infants of seropositive women during the neonatal (age less than 28 days) and postneonatal periods. In blood obtained during the neonatal period, the polymerase chain reaction was positive in five of seven infants in whom the acquired immunodeficiency syndrome (AIDS) later developed (a mean of 9.8 months after the test). The test was also positive in one of eight newborns who later had nonspecific signs and symptoms suggestive of HIV infection (mean follow-up, 12 months). No proviral sequences were detected in neonatal samples from nine infants who remained well (mean follow-up, 16 months). HIV proviral sequences were detected in samples obtained during the postneonatal period (median age, five months) in all of 6 infants tested who later had AIDS and in 4 of 14 infants with nonspecific findings suggestive of HIV infection. No proviral sequences were detected in 25 infants who remained well (mean follow-up, 17 months) after being born to HIV-seropositive mothers, or in 15 infants born to HIV-seronegative mothers. We conclude that the polymerase chain reaction will be a useful technique to diagnose HIV infection in newborns and to predict the subsequent development of AIDS. However, larger studies will be required to determine the sensitivity and specificity of the test.

Acquired Immunodeficiency Syndrome↗
PubMed1989

Molecular characterization of HIV-1 isolated from a serum collected in 1976: nucleotide sequence comparison to recent isolates and generation of hybrid HIV.

Human immunodeficiency virus type 1 (Z321 designate, HIV-1Z321), the oldest known HIV, was isolated from a serum sample collected in Zaire in 1976 and was molecularly cloned. Restriction enzyme analysis of unintegrated viral DNA revealed the presence of conserved restriction enzyme cleavage sites in the long terminal repeat sequences. Nucleotide sequence analysis of the 3' end of the viral DNA revealed a pattern similar to other HIV-1 isolates described. However, some of the common restriction sites present in other isolates were absent in HIV-1Z321. The extent of differences between HIV-1Z321 and recent isolates from North America and Zaire was 17.86-18.36% on the nucleotide sequence level and 26.5-33.2% difference in the predicted amino acid sequence in the envelope gene. Differences were also noted in 3'-orf (nef: according to HIV gene nomenclature; see Ref. 42) gene and U3 region of the long terminal repeat sequences of HIV-1Z321 and other isolates. Nucleotide sequence of a HIV-1 isolate, 12 years apart from the present isolates, will provide an important time calibration point for the evolutionary divergence of HIV isolates. Hybrid HIV was also generated by transfecting HIV-1Z321 and HIV-1HTLV-III viral DNAs into cells.

Amino Acid Sequence↗
PubMed1989

Prevalence of human immunodeficiency virus type 1 DNA in hemophilic men and their sex partners. Hemophilia-AIDS Collaborative Study Group.

Polymerase chain reaction (PCR) was used to detect human immunodeficiency virus (HIV)-1 DNA in peripheral blood mononuclear cells to assess in hemophilic men whether any were HIV-seropositive but uninfected or seronegative but infected and in seronegative sex partners of seropositive hemophilic men whether any were infected. Of 40 seropositive men, 38 (95%) were PCR-positive; one was PCR-indeterminate and one PCR-negative. None of 41 seronegative men who used only donor-screened, virus-inactivated coagulation factor products were PCR-positive. However, two of six who received noninactivated products were PCR-positive; one had low T-helper cell counts and died of unrelated causes and the other had seroconverted 11 mo later. PCR with a second primer pair also detected HIV-1 DNA in these two men. None of 25 seronegative female sex partners of seropositive men, including six men with AIDS and seven with AIDS-related symptoms, were PCR-positive. These data suggest that most seropositive hemophilic men are HIV-infected; whether some are infected with defective virus remains to be resolved as does the infection status of seropositive PCR-negative men. Identification of two seronegative PCR-positive men supports the possibility that HIV-1 DNA can be detected before seroconversion.

DNA, Viral↗
PubMed1989

Potential for insect transmission of HIV: experimental exposure of Cimex hemipterus and Toxorhynchites amboinensis to human immunodeficiency virus.

Human immunodeficiency virus (HIV) was detected in bedbugs (Cimex hemipterus) up to 8 d after oral exposure to highly concentrated virus in blood meals, but no virus replication was observed. HIV did not replicate in either intraabdominally inoculated bedbugs or intrathoracically inoculated mosquitoes (Toxorhynchites amboinensis). The virus was not detected in bedbug feces. Mechanical transmission of HIV by bedbugs could not be demonstrated in an in vitro model. The persistence of HIV in an insect or on its mouthparts is one of many factors necessary for mechanical transmission in nature. The risk of insect transmission of HIV appears to be extremely low or nonexistent.

Animals↗