Prices of CD4 assays and viral load tests must be reduced for developing countries.
Explore the source record for details and available documents.
CD4 Lymphocyte Count↗
Biomedical subjects
C Bartholomew
Publications and source records attributed to C Bartholomew.
Immunologic and virologic analyses of an acutely HIV type 1-infected patient with extremely rapid disease progression.
The immunologic and virologic factors that impact on the rate of disease progression after acute infection with human immunodeficiency virus (HIV) type 1 are poorly understood. A patient with an extraordinarily rapid disease course leading to AIDS-associated death within 6 months of infection was studied intensively for the presence of anti-HIV immune reactivities as well as changes in the genetic and biologic properties of virus isolates. Although altered humoral responses were evident, the most distinctive immunologic feature was a nearly complete absence of detectable HIV-specific CTL responses. In addition to a rapid decline in CD3+CD4+ cells, elevated percentages of CD8+CD45RA+ and CD8+CD57+ cells and diminished CD8+CD45R0+ and CD8+CD28+ cells were evident. Primary viral isolates recovered throughout the course of infection exhibited limited sequence diversity. Cloned viral envelopes were found to have unusually broad patterns of coreceptor usage for cell-cell fusion, although infectivity studies yielded no evidence of infection via these alternative receptors. The infectivity studies demonstrated that these isolates and their envelopes maintained an R5 phenotype throughout the course of disease. The absence of demonstrable anti-HIV CTL reactivities, coupled with a protracted course of seroconversion, highlights the importance of robust HIV-specific immune responses in the control of disease progression.
Acute Disease↗
Detection of functional PTEN lipid phosphatase protein and enzyme activity in squamous cell carcinomas of the head and neck, despite loss of heterozygosity at this locus.
The human tumour suppressor gene PTEN located at 10q23 is mutated in a variety of tumour types particularly metastatic cases and in the germline of some individuals with Cowdens cancer predisposition syndrome. We have assessed the status of PTEN and associated pathways in cell lines derived from 19 squamous cell carcinomas of the head and neck. Loss of heterozygosity is evident at, or close to the PTEN gene in 5 cases, however there were no mutations in the remaining alleles. Furthermore by Western analysis PTEN protein levels are normal in all of these SCC-HN tumours and cell lines. To assess the possibility that PTEN may be inactivated by another mechanism, we characterized lipid phosphatase levels and from a specific PIP3 biochemical assay it is clear that PTEN is functionally active in all 19 human SCCs. Our data strongly suggest the possibility that a tumour suppressor gene associated with development of SCC-HN, other than PTEN, is located in this chromosomal region. This gene does not appear to be MXI-1, which has been implicated in some other human tumour types. PTEN is an important negative regulator of PI3Kinase, of which subunit alpha is frequently amplified in SCC-HN. To examine the possibility that PI3K is upregulated by amplification in this tumour set we assessed the phosphorylation status of Akt, a downstream target of PI3K. In all cases there is no detectable increase in Akt phosphorylation. Therefore there is no detectable defect in the PI3K pathway in SCC-HN suggesting that the reason for 3q26.3 over-representation may be due to genes other than PI3K110alpha.
Blotting, Western↗
Evi-1 transforming and repressor activities are mediated by CtBP co-repressor proteins.
Ectopic production of the EVI1 transcriptional repressor zinc finger protein is seen in 4--6% of human acute myeloid leukemias. Overexpression also transforms Rat1 fibroblasts by an unknown mechanism, which is likely to be related to its role in leukemia and which depends upon its repressor activity. We show here that mutant murine Evi-1 proteins, lacking either the N-terminal zinc finger DNA binding domain or both DNA binding zinc finger clusters, function as dominant negative mutants by reverting the transformed phenotype of Evi-1 transformed Rat1 fibroblasts. The dominant negative activity of the non-DNA binding mutants suggests sequestration of transformation-specific cofactors and that recruitment of these cellular factors might mediate Evi-1 transforming activity. C-terminal binding protein (CtBP) co-repressor family proteins bind PLDLS-like motifs. We show that the murine Evi-1 repressor domain has two such sites, PFDLT (site a, amino acids 553--559) and PLDLS (site b, amino acids 584--590), which independently can bind CtBP family co-repressor proteins, with site b binding with higher affinity than site a. Functional analysis of specific CtBP binding mutants show site b is absolutely required to mediate both transformation of Rat1 fibroblasts and transcriptional repressor activity. This is the first demonstration that the biological activity of a mammalian cellular transcriptional repressor protein is mediated by CtBPs. Furthermore, it suggests that CtBP proteins are involved in the development of some acute leukemias and that blocking their ability to specifically interact with EVI1 might provide a target for the development of pharmacological therapeutic agents.
Animals↗
Fourth-generation enzyme-linked immunosorbent assay for the simultaneous detection of human immunodeficiency virus antigen and antibody.
The VIDAS HIV DUO Ultra, a fourth-generation immunoassay under development for the simultaneous detection of human immunodeficiency virus type 1 (HIV-1) p24 antigen and antibodies to HIV-1 and HIV-2, was evaluated. The enzyme-linked fluorescence immunoassay, performed on the automated VIDAS instrument, is claimed to detect early and established HIV infection. The assay was challenged with a total of 2,847 samples that included 74 members of 10 seroconversion panels, 9 p24 antigen-only-reactive members of a panel of group M clades, 503 consecutively collected samples from individuals seeking care in the University of Maryland Medical System, 1,010 samples from U.S. blood donors, 1,141 samples from patients in a high-incidence population in Trinidad, 83 samples from a clinic for sexually transmitted diseases in the Bahamas, 10 confirmed HIV-1 group O samples, and 16 confirmed HIV-2 samples from the Cote d'Ivoire. Reference tests were U.S. Food and Drug Administration-licensed HIV antibody screening, p24 antigen tests, HIV confirmatory assays, and the Roche Diagnostics Amplicor HIV-1 Monitor. The VIDAS HIV DUO Ultra demonstrated 100% sensitivity and 99.5% specificity overall, with a 99.7% specificity in low-risk individuals. The analytical sensitivity, as assessed by seroconversion panels and p24 antigen in samples, was equivalent to the sensitivity of the reference assays used to characterize these panels. The VIDAS HIV DUO Ultra is accurate, offers potential advantages over conventional HIV testing for time and cost savings, has walk-away capability, and correctly identifies both early and established HIV infections.
Enzyme-Linked Immunosorbent Assay↗
A distinctive clade B HIV type 1 is heterosexually transmitted in Trinidad and Tobago.
HIV-1 transmission worldwide is predominantly associated with heterosexual activity, and non-clade B viruses account for the most spread. The HIV-1 epidemic in Trinidad/Tobago and the Caribbean shares many features with such heterosexual epidemics, including a prominent role for coincident sexually transmitted diseases. This study evaluates the molecular epidemiology of HIV-1 in Trinidad/Tobago during a period when abrupt transition from homosexual to heterosexual transmission occurred in the absence of injecting drug use, concomitant with a rapid rise in HIV-1 prevalence in the heterosexual population. Of 31 viral isolates studied during 1987-1995, all cluster with subtype B reference strains. In the analysis of full env genes from 22 early seroconverters, the Trinidad isolates constitute a significant subcluster within the B subtype. The Trinidad V3 consensus sequence differs by a single amino acid from the prototype B V3 consensus and demonstrates stability over the decade of this study. In the majority of isolates, the V3 loop of env contains a signature threonine deletion that marks the lineage of the Trinidad HIV-1 clade B epidemic from pre-1984. No phenotypic features, including syncitium induction, neutralization profiles, and chemokine receptor usage, distinguish this virus population from other subtype B viruses. Thus, although the subtype B HIV-1 viruses being transmitted in Trinidad are genetically distinguishable from other subtype B viruses, this is probably the result of a strong founder effect in a geographically circumscribed population rather than genetic selection for heterosexual transmission. These results demonstrate that canonical clade B HIV-1 can generate a typical heterosexual epidemic.
Amino Acid Sequence↗
A study of HTLV-I infection and breast cancers in Trinidad and Tobago.
Explore the source record for details and available documents.
Adult↗
Loss of cell cycle control by deregulation of cyclin-dependent kinase 2 kinase activity in Evi-1 transformed fibroblasts.
The Evi-1 transcriptional repressor protein has two distinct zinc finger DNA binding domains designated ZF1 and ZF2 and is implicated in the progression of human and murine leukemias, in which it is abnormally expressed. In this report, we show that Evi-1-expressing Rat1 fibroblasts are anchorage independent, have an abbreviated G1 phase of the cell cycle, and have a reduced requirement for serum mitogens for S-phase entry. These biological changes are accompanied by a moderately increased production of cell cycle-regulatory proteins cyclin A and cyclin-dependent kinase (Cdk) 2, a dramatic deregulation of Cdk2 kinase activity, and a corresponding increase in the levels of hyperphosphorylated retinoblastoma protein (pRb). We show that the elevated cyclin A-Cdk2 activity is due to the combination of increased accumulation and stabilization of cyclin A bound to a faster-migrating species of Cdk2 believed to be the active threonine 160 phosphorylated form and a substantial reduction in complexed p27. Cyclin E kinase activity is also elevated due to a reduction in p27. A significant reduction in total cellular p27 protein levels and a moderate reduction in p27 mRNA are observed, but no changes in Cdk regulatory kinases and phosphatases occur. The Evi-1 transcriptional repressor domain and the ZF1 DNA binding domain are required for both cell transformation and induction of Cdk2 catalytic activity. We propose that one consequence of Evi-1 expression is to repress the transcription of target genes, which may include p27, that deregulate the normal control of the G1 phase of the cell cycle, providing a cellular proliferative advantage that contributes to transformation in vitro and leukemogenesis in vivo.
Animals↗
An activating mutation in the kit receptor abolishes the stroma requirement for growth of ELM erythroleukemia cells, but does not prevent their differentiation in response to erythropoietin.
We have previously shown that murine ELM erythroleukemia cells can only be grown in vitro in the presence of a stromal feeder layer, or alternatively stem cell factor (SCF), without which they differentiate. When grown in the presence of SCF, ELM cells can still differentiate in response to erythropoietin (Epo), but growth on stroma prevents this. We previously isolated a stroma-independent ELM variant, ELM-I-1, that is also defective in Epo-induced differentiation. We show here that this variant has an activating mutation in the Kit receptor, converting aspartic acid 814 to histidine. Expression of the mutant receptor in stroma-dependent ELM-D cells causes growth factor-independent proliferation and also gives the cells a selective advantage, in terms of proliferation rate and clonegenicity, compared with ELM-D cells grown in optimal amounts of SCF. Expression of the mutant receptor in ELM-D cells also prevents spontaneous differentiation, but not differentiation induced by Epo. Analysis of mitogenic signaling pathways in these cells shows that the mutant receptor induces constitutive activation of p42/p44 mitogen-activated protein kinases. It also selectively inhibits the expression of p66Shc but not the p46/p52 Shc isoforms (as did treatment of ELM cells with SCF), which is of interest, because p66Shc is known to play an inhibitory role in growth factor signaling.
Adaptor Proteins, Signal Transducing↗
Direct and indirect estimates of HIV-1 incidence in a high-prevalence population.
While the worldwide AIDS epidemic continues to expand, directly measured incidence data are difficult to obtain. Methods to reliably estimate human immunodeficiency virus type 1 (HIV-1) incidence from more easily available data are particularly relevant in those parts of the world where prevalence is rising in heterosexually exposed populations. The authors set out to estimate HIV-1 incidence in a population of heterosexual sexually transmitted disease clinic attendees in Trinidad who had a known high prevalence of HIV-1 subtype B. Over the period 1987-1995, HIV-1 incidence estimates from serial cross-sectional studies of HIV-1 prevalence, passive follow-up of clinic recidivists, modeling of early markers of HIV-1 infection (p24 antigen screening), and a cohort study of seronegative genital ulcer disease cases were compared. Measuring incidence density in the genital ulcer disease cases directly gave the highest estimate, 6.9% per annum. Screening for the detection of early HIV-1 markers yielded an incidence of 5.0% per annum, while estimating incidence from serial cross-sectional prevalence data and clinic recidivists gave estimates of 3.5% and 4.5% per annum, respectively. These results were found to be internally consistent. Indirect estimates of incidence based on prevalence data can give accurate surrogates of true incidence. Within limitations, even crude measures of incidence are robust enough for health planning and evaluation purposes. For planning vaccine efficacy trials, consistent conservative estimates may be used to evaluate populations before targeting them for cohort studies.
Blotting, Western↗
Evi-1 ZF1 DNA binding activity and a second distinct transcriptional repressor region are both required for optimal transformation of Rat1 fibroblasts.
The Evi-1 gene encodes a zinc finger transcriptional repressor protein that normally plays a role in development and is frequently activated in myeloid leukaemias. Evi-1 has two distinct DNA binding domains, ZF1 and ZF2, and a defined repressor domain but the function of the remainder of the molecule is unknown. The ZF2 and repressor domains have been shown to be required for transformation and we show here that ZF1 is also required. An alternative splice variant of Evi-1, designated delta324, encodes a protein which lacks a portion of the ZF1 DNA binding domain and the intervening amino acids 239-514 (designated IR) located between ZF1 and the repressor domain. We show that delta324 can neither bind ZF1, repress transcription through this site nor transform Rat1 fibroblasts. Reconstitution studies demonstrate that the defect in delta324 is partially complemented by recreating the ZF1 DNA binding activity. However, full function also requires the IR region which has transcriptional repressor activity. This study shows therefore, that ZF1, ZF2 and repressor domains and the IR region all contribute to the transformation efficiency of the Evi-1 protein.
Alternative Splicing↗
Both stroma and stem cell factor maintain long-term growth of ELM erythroleukemia cells, but only stroma prevents erythroid differentiation in response to erythropoietin and interleukin-3.
Defining how the stromal requirements of hematopoietic progenitors change during leukemia progression is an important topic that is not well understood at present. The murine ELM erythroleukemia is an interesting model because the erythroid progenitors retain dependence on bone marrow-derived stromal cells for long-term growth in vitro, and they also undergo erythroid differentiation in the presence of erythropoietin (EPO) and interleukin-3 (IL-3). In this report, we have shown using neutralizing antibodies that stem cell factor (SCF), insulin-like growth factor (IGF)-1, and integrin signaling pathways are all involved. We then determined whether ELM cells can be maintained long-term without stroma in various combinations of growth factors produced by stroma cells or growth factors for which ELM cells have receptors. This showed that ELM cells could be maintained with high efficiency in SCF alone; furthermore, the cells remained absolutely SCF-dependent and did not become more tumorigenic than cells maintained on stroma. In contrast, ELM cells underwent clonal extinction when serially cloned in IGF1; any cells that survived long-term growth in IGF-1 were found to be IGF1-independent. One important difference between maintaining ELM cells on stroma and growth in SCF is that stroma reversibly inhibits their differentiation in response to EPO and IL-3, whereas SCF does not.
Animals↗
Incidence of HTLV-I-associated myelopathy/tropical spastic paraparesis (HAM/TSP) in Jamaica and Trinidad.
HTLV-I is sexually transmitted more efficiently from men to women than vice versa, and the majority of HTLV-I endemic areas report a female preponderance of HTLV-I-associated myelopathy/tropical spastic paraparesis (HAM/TSP) cases. The objective of this study was to estimate the gender- and age-specific incidence rates of HAM/TSP in the general population as well as in the HTLV-I-infected population in Jamaica and in Trinidad and Tobago. Incidence rates for HAM/TSP were computed based on all reported incident cases in both countries between 1990 and 1994. Population census reports for 1990 were used to calculate the population at risk. The age-standardized HAM/TSP incidence rate (mean +/- standard error of the mean) in Jamaica was 1.8 +/- 0.2/100,000 person years (PY). Among individuals of African descent in Trinidad and Tobago, the rate was 1.7 +/- 0.4/100,000 PY. As in HTLV-I seroprevalence, the incidence rate of HAM/TSP increased with age through the fifth decade of life and was three times as high in women than in men. The HAM/TSP incidence rate, calculated as a function of the number of HTLV-I-infected persons in each age stratum, is higher in women (24.7/100,000 PY) than in men (17.3/100,000 PY). With HTLV-I infection, the lifetime risk of developing HAM/TSP was estimated to be 1.9% overall and is slightly higher in women (1.8%) than in men (1.3%). Thus, the higher prevalence of HTLV-I in women in endemic areas does not fully explain the preponderance of female HAM/TSP, suggesting that other cofactors must be present. The higher incidence rate in women between the ages of 40 and 59 years, as well as the increase in HAM/TSP incidence rates with age, are indicative of the importance of adult-acquired HTLV-I infection, presumably through sexual transmission.
Adult↗
The Evi-1 proto-oncogene encodes a transcriptional repressor activity associated with transformation.
The myeloid transforming gene Evi-1 encodes a protein with two zinc finger domains, designated ZF1 and ZF2, with distinct DNA binding specificities. For the first time we demonstrate that Evi-1 has transcriptional repressor activity which is directly proportional to the amount of Evi-1 protein in cells. Repression has been observed with two distinct promoters: the minimal HSV-1 tk promoter and a VP16 inducible adenovirus E1b minimal promoter. Optimal repression is DNA binding dependent and is mediated by either ZF1 or a heterologous GAL4 DNA binding domain (GAL4DBD) but is significantly less efficient through the ZF2 binding site. Both GAL4DBD/Evi-1 fusion and non-fusion proteins have been used to map the repressor activity to a proline-rich region located within amino acids 514-724 between the ZF1 and ZF2 domains. Constitutive expression of mutant proteins lacking the repressor domain are defective for transformation of Rat1 fibroblasts demonstrating that this region is required for the oncogenic activity of the Evi-1 protein. These studies show that the Evi-1 gene encodes a transcriptional repressor and has important implications for the mechanism of action of the Evi-1 protein both in development and in the progression of some myeloid leukaemias.
3T3 Cells↗
Extensive latent retroviral infection in bone marrow of patients with HTLV-I-associated neurologic disease.
Explore the source record for details and available documents.
Bone Marrow↗
Human T-cell lymphotropic virus type I-associated facial nerve palsy in Trinidad and Tobago.
To assess the association of the human T-cell lymphotropic virus type I (HTLV-I) and idiopathic facial nerve palsy of the lower motor neuron type, we studied 78 patients consecutively admitted to the Port of Spain General Hospital in Trinidad, the West Indies, with a confirmed diagnosis of idiopathic facial nerve palsy. Patients were compared with two control groups: a population-based group of persons 20 years and older and a hospital-based group of patients 15 to 84 years old admitted to the medical wards. Sixty-two patients were Trinidadians of African origin and 16 were Trinidadians of East Indian origin. None of the East Indian patients was HTLV-I antibody positive. Three Afro-Trinidadians were infected with human immunodeficiency virus type 1 and 1 was coinfected with this virus and HTLV-I. Of the remaining 58 Afro-Trinidadians, 12 (20.7%) were HTLV-I positive only. This rate was statistically higher than the HTLV-I seroprevalence in the Afro-Trinidadian general population (3.5%) and the hospital control group (5.6%). After age standardization, the HTLV-I prevalence for patients with facial nerve palsy remained significantly elevated. HTLV-I antibody assays should be performed on all patients with idiopathic facial nerve palsy of the lower motor neuron type who live in HTLV-I endemic areas or are migrants from these areas.
Adolescent↗
The Evi1 proto-oncogene is required at midgestation for neural, heart, and paraxial mesenchyme development.
The ecotropic viral integration site-1 (Evi1) locus was initially identified as a common site of retroviral integration in myeloid tumors of the AKXD-23 recombinant inbred mouse strain. The full-length Evi1 transcript encodes a putative transcription factor, containing ten zinc finger motifs found within two domains of the protein. To determine the biological function of the Evi1 proto-oncogene, the full-length, but not an alternately spliced, transcript was disrupted using targeted mutagenesis in embryonic stem cells. Evi1 homozygous mutant embryos die at approximately 10.5 days post coitum. Mutants were distinguished at 10.5 days post coitum by widespread hypocellularity, hemorrhaging, and disruption in the development of paraxial mesenchyme. In addition, defects in the heart, somites, and cranial ganglia were detected and the peripheral nervous system failed to develop. These results correlated with whole-mount in situ hybridization analyses of embryos which showed expression of the Evi1 proto-oncogene in embryonic mesoderm and neural crest-derived cells associated with the peripheral nervous system. These data suggest that Evi1 has important roles in general cell proliferation, vascularization, and cell-specific developmental signaling, at midgestation.
Animals↗
The risks and benefits of childhood bacille Calmette-Guérin immunization among adults with AIDS. International MAC study groups.
OBJECTIVE: To define the risks of disseminated bacille Calmette-Guérin (BCG) or disseminated Mycobacterium tuberculosis in adults with AIDS who were immunized with BCG in childhood. DESIGN: HIV-infected patients with CD4 < 200 x 10(6)/l were enrolled from five study sites (New Hampshire, Boston, Finland, Trinidad and Kenya). Prior BCG immunization was determined and blood cultures for mycobacteria were obtained at study entry and at 6 months. Acid-fast bacilli were identified as Mycobacterium tuberculosis complex (MTBC) using DNA probes. MTBC isolates were then typed by both IS6110 restriction fragment length polymorphism and polymerase chain reaction/restriction enzyme analysis. SETTING: Most patients in New Hampshire and Finland were outpatients; most patients in Trinidad were inpatients with terminal illness; and most patients in Kenya were outpatients, although 44 were inpatients with terminal illness. PARTICIPANTS: A total of 566 patients were enrolled, including 155 with childhood BCG immunization; 318 patients had a single study visit and culture, and 248 patients had two study visits and cultures. MAIN OUTCOME MEASURES: Isolation and identification of mycobacteria from blood cultures. RESULTS: Blood cultures were positive for MTBC in 21 patients; none were positive for M. bovis BCG, and 21 were M. tuberculosis-positive. In Trinidad, seven (87%) out of eight isolates of M. tuberculosis were indistinguishable by IS6110 typing; BCG immunization was associated with a decreased risk of bacteremic infection with M. tuberculosis (P = 0.05). CONCLUSIONS: The risk of disseminated BCG among adult AIDS patients with childhood BCG immunization is very low. Childhood BCG immunization is associated with protection against bacteremia with M. tuberculosis among adults with advanced AIDS in Trinidad.
AIDS-Related Opportunistic Infections↗