Biomedical subjects

T F Rinke de Wit

Publications and source records attributed to T F Rinke de Wit.

At least 19 recordsLinked to original sources

PubMed2001

No difference in in vitro susceptibility to HIV type 1 between high-risk HIV-negative Ethiopian commercial sex workers and low-risk control subjects.

Host factors such as increased beta-chemokine production, HIV-1 coreceptor expression level, and HIV-1 coreceptor polymorphism have been thought to influence susceptibility to HIV-1 infection. To determine the protective role of these factors in Ethiopians who remained HIV-1 uninfected, despite multiple high-risk sexual exposures, we studied 21 Ethiopian women who had been employed as commercial sex workers (CSWs) for five or more years. The HIV-1-resistant CSWs were compared with low-risk age-matched female controls who had a comparable CD4+ cell percentage and mean fluorescence intensity (MFI). Genetic polymorphism in the CCR5, CCR2b, or SDF-1 genes appeared not to be associated with resistance in the Ethiopian CSWs. Expression levels of CCR5 and CXCR4 on naive, memory, and total CD4+ T cells tended to be higher in the resistant CSWs, while the production of beta-chemokines RANTES, MIP-1alpha, and MIP-1beta by phytohemagglutinin (PHA)-stimulated peripheral blood mononuclear cells (PBMCs) was lower compared with low-risk HIV-1 negative controls. In vitro susceptibility of PHA-stimulated PBMCs to primary, CCR5-restricted, Ethiopian HIV-1 isolates was comparable between resistant CSWs and low-risk controls. In vitro susceptibility was positively correlated to CD4+ cell mean fluorescence intensity and negatively correlated to CCR5 expression levels, suggesting that infection of PBMCs was primarily dependent on expression levels of CD4 and that CCR5 expression, above a certain threshold, did not further increase susceptibility. Our results show that coreceptor polymorphism, coreceptor expression levels, beta-chemokine production, and cellular resistance to in vitro HIV-1 infection are not associated with protection in high-risk HIV-1-negative Ethiopian CSWs.

Adult↗

PubMed2001

Plasma levels of viro-immunological markers in HIV-infected and non-infected Ethiopians: correlation with cell surface activation markers.

Cross-sectional studies were conducted to measure soluble viral and immunological markers in plasma in order to determine the prognostic value of these markers for HIV disease progression in Ethiopians and to see their association with cell surface markers in HIV-1-infected and noninfected Ethiopians. Whole blood samples were collected from 52 HIV-1-negative Ethiopians, 32 HIV-1-positive Ethiopians with absolute CD4(+) T-cell count >200/microl whole blood and no AIDS defining conditions, and 39 HIV-positive Ethiopians with CD4(+) T-cell count <200/microl and/or AIDS defining conditions. Plasma levels of b(2)-microglobulin (b(2)m), soluble CD27 (sCD27), soluble tumor necrosis factor alpha receptor type II (sTNFR-II), IgG, IgA, IGE, and IL12 were elevated in HIV-1-infected individuals. The plasma levels of sTNFR-II, sCD27, b(2)m, IL12, and IgG were inversely correlated with numbers of CD4(+) T-cells, the proportion of naïve (CD45RA(+)CD27(+)) CD8(+) T-cells, and the proportion of CD8(+) T-cells expressing CD28 (CD8(+)CD28(+)) were positively correlated with the proportions of activated (HLA-DR(+)CD38(+)) CD4(+) T-cells, as well as activated (HLA-DR(+)CD38(+)) CD8(+) T-cells. A strong positive correlation was also observed when soluble immune markers were compared to each other. Multivariate regression analyses of soluble markers with numbers of CD4(+) T-cells showed that sCD27 is the best independent marker for CD4(+) T-cell decline in the HIV-1-infected Ethiopians. Our results indicate that measurement of soluble immune markers, which is relatively easy to perform, could be a good alternative to the quantification of T-cell subsets for monitoring HIV-1 disease progression in places where there is no facility for flow cytometric measurements.

Acquired Immunodeficiency Syndrome↗

PubMed2000

Identification of a genetic subcluster of HIV type 1 subtype C (C') widespread in Ethiopia.

Others and we have previously shown that subtype C is the predominant HIV-1 subtype and the major cause of AIDS in Ethiopia. The present study shows that subtype C in Ethiopia has a genetic subcluster, designated C', has not increased in frequency, or spread geographically, over the period 1988 (%C' = 23/53) to 1996-1997 (%C' = 26/50). There is no association of the HIV-1 subtype C or subcluster C' with geographic location, time of sample collection, or risk group in Ethiopia. Of 105 randomly collected samples representing 7 different towns in Ethiopia, all but 2 (1 subtype A from Addis Ababa, 1997 and 1 subtype D from Dessie, 1996) belong to subtype C.

Amino Acid Sequence↗

PubMed2000

HIV-1 subtype C in commerical sex workers in Addis Ababa, Ethiopia.

In this study, we have investigated the diversity of the current HIV-1 strains circulating in Addis Ababa, Ethiopia; in addition, we have evaluated the applicability of peptide enzyme-linked immunosorbent assay (ELISA) and heteroduplex mobility assay (HMA) for HIV-1 subtyping. Previous studies have indicated that HIV-1 subtype C is the major subtype present in HIV-positive samples collected from various risk groups between 1988 and 1995 in Addis Ababa. To assess the possible influx of new HIV-1 subtypes, 150 commercial sex workers (CSW) reporting in 1997 to two Health Centers in Addis Ababa were enrolled in an unlinked anonymous cross-sectional study. Subtyping was performed according to the World Health Organization algorithm of peptide ELISA, followed by HMA and DNA sequencing. As a result, the HIV-1 prevalence among these CSWs was found to be 45% (67 of 150). Of the 67 samples, 66 contained HIV-1 of subtype C and only one was of subtype D. This confirms the persistent overall presence of HIV-1 subtype C in Addis Ababa and a low influx of other subtypes into this location.

Amino Acid Sequence↗

PubMed2000

Increased cell division but not thymic dysfunction rapidly affects the T-cell receptor excision circle content of the naive T cell population in HIV-1 infection.

Recent thymic emigrants can be identified by T cell receptor excision circles (TRECs) formed during T-cell receptor rearrangement. Decreasing numbers of TRECs have been observed with aging and in human immunodeficiency virus (HIV)-1 infected individuals, suggesting thymic impairment. Here, we show that in healthy individuals, declining thymic output will affect the TREC content only when accompanied by naive T-cell division. The rapid decline in TRECs observed during HIV-1 infection and the increase following HAART are better explained not by thymic impairment, but by changes in peripheral T-cell division rates. Our data indicate that TREC content in healthy individuals is only indirectly related to thymic output, and in HIV-1 infection is mainly affected by immune activation.

Anti-HIV Agents↗

PubMed2000

Nonradioactive techniques for measurement of in vitro T-cell proliferation: alternatives to the [(3)H]thymidine incorporation assay.

T-cell proliferation is an important in vitro parameter of in vivo immune function and has been used as a prognostic marker of human immunodeficiency virus type 1 (HIV-1) disease progression. The proliferative capacity of T cells in response to various stimuli is commonly determined by a radioactive assay based on incorporation of [(3)H]thymidine ([(3)H]TdR) into newly generated DNA. In order to assess techniques for application in laboratories where radioactive facilities are not present, two alternative methods were tested and compared to the [(3)H]TdR assay as a "gold standard." As an alternative, T-cell proliferation was measured by flow cytometric assessment of CD38 expression on T cells and by an enzyme-linked immunosorbent assay (ELISA) based on bromo-2'-deoxyuridine (BrdU) incorporation. Peripheral blood mononuclear cells (PBMCs), either in whole blood or Ficoll-Isopaque separated, from a total of 26 HIV-1-positive and 18 HIV-1-negative Dutch individuals were stimulated with CD3 monoclonal antibody (MAb) alone, a combination of CD3 and CD28 MAbs, or phytohemagglutinin. BrdU incorporation after 3 days of stimulation with a combination of CD3 and CD28 MAbs correlated excellently with the [(3)H]TdR incorporation in both study groups (HIV-1 positives, r = 0.96; HIV-1 negatives, r = 0.83). A significant correlation of absolute numbers of T cells expressing CD38 with [(3)H]TdR incorporation, both in HIV-1-positive (r = 0.96) and HIV-1-negative (r = 0.84) individuals, was also observed under these conditions. The results of this study indicate that determination of both the number of CD38-positive T cells and BrdU incorporation can be used as alternative techniques to measure the in vitro T-cell proliferative capacity. The measurement of CD38 expression on T cells provides the additional possibility to further characterize the proliferating T-cell subsets for expression of other surface markers.

ADP-ribosyl Cyclase↗

PubMed1999

HIV-1 subtype C syncytium- and non-syncytium-inducing phenotypes and coreceptor usage among Ethiopian patients with AIDS.

OBJECTIVE: To assess syncytium-inducing (SI) and non-syncytium-inducing (NSI) frequencies, coreceptor usage and gp120 V3 sequences of HIV-1 isolates from Ethiopian AIDS patients. PATIENTS: Cross-sectional study on 48 hospitalized AIDS patients (CD4 T cells < 200 x 10(6) cell/l) with stage III or IV of the WHO staging system for HIV-1 infection and disease. METHODS: Peripheral blood mononuclear cells (PBMC) from all 48 patients were tested by MT-2 assay to determine SI/NSI phenotypes. Lymphocyte subsets were enumerated using Coulter counting and FACScan analysis. Viral load determination used a nucleic acid sequence-based amplification assay (NASBA). Coreceptor usage of HIV-1 biological clones was measured using U87 CD4/chemokine receptor transfectants and phytohemagglutinin-stimulated PBMC of healthy donors with wild-type CCR5 and homozygous mutation CCR5delta32 (a 32 base-pair deletion in CCR5). Reverse transcriptase polymerase chain reaction sequencing was performed on the third variable region (V3) of the HIV-1 gene gp120. Sequence alignments were done manually; phylogenetic analyses used PHYLIP software packages. RESULTS: SI viruses were detected for 3/48 (6%) AIDS patients only. Lower mean absolute CD4 counts were determined in patients with SI virus compared with NSI (P = 0.04), but no differences in viral load were observed. All patients were found to be infected with HIV-1 subtype C, based on V3 sequencing. NSI biological clones used CCR5 as coreceptor; SI biological clones used CXCR4 and/or CCR5 and/or CCR3. CONCLUSIONS: Ethiopian patients with HIV-1 C-subtype AIDS harbour a remarkably low frequency of SI phenotype viruses. Coreceptor usage of these viruses correlates with their biological phenotypes.

Adult↗

PubMed1999

Evaluation of the World Health Organization staging system for HIV infection and disease in Ethiopia: association between clinical stages and laboratory markers.

OBJECTIVE: To study the association between the clinical axis of the World Health Organization (WHO) staging system of HIV infection and disease and laboratory markers in HIV-infected Ethiopians. DESIGN: Cross-sectional study. METHODS: Clinical manifestations and stage of HIV-positive individuals participating in a cohort study of HIV infection progression, and of HIV-positive patients hospitalized with suspicion of AIDS, were compared to CD4+ T-cell count and viral load. RESULTS: Of the 86 HIV-positive participants of the cohort study, 53 (62%), 16 (19%), 16 (19%), and one (1.2%) were in stage 1, 2, 3 and 4, respectively. Minor weight loss (n = 15) and pulmonary tuberculosis (n = 9) were the most commonly diagnosed conditions among the 38 (44%) symptomatic HIV-positive individuals. Although 23 (27%) HIV-positive participants had CD4+ T-cell counts less than 200 x 10(6)/l, only one was in clinical stage 4. Among 79 hospitalized HIV-positive patients, 15 (19%) and 64 (81%) were in stage 3 and 4, respectively. The majority (83.5%) had CD4+ T-cell counts < 200 x 10(6)/l. Individuals at stage 3 had lower CD4+ T-cell counts and higher viral loads when seen in hospital as compared to cohort participants (P = 0.06 and 0.008, respectively). When grouping the two study populations, the median CD4+ T-cell count decreased (337, 262, 225, 126, and 78 x 10(6)/l, P< 0.01), and the median viral load increased (4.08, 3.89, 4.47, 5.65, and 5.65 log10 copies/ml, P < 0.01), with increasing clinical stage of HIV infection (1, 2, 3 cohort, 3 hospital, and 4, respectively). Median CD4+ T-cell counts were remarkably low in HIV-negative participants (749 x 10(6)/l), and in HIV-positive participants at stage 1 and 2 (337 and 262 x 10(6)/l, respectively). CONCLUSIONS: There was a good correlation between WHO clinical stages and biological markers. CD4+ T-cell counts were low in Ethiopians, particularly during early stages of HIV-1 infection, and preliminary reference values at different stages of HIV-1 infection were determined. In HIV-infected Ethiopians, lymphocyte counts less than 1,000 x 10(6)/l in non-hospitalized individuals, and less than 2,000 x 10(6)/l in hospitalized patients, had high positive predictive value, but low sensitivity, in identifying subjects with low CD4+ T-cell counts (< 200 x 10(6)/l) who would benefit from chemoprophylaxis of opportunistic infections. The on-going longitudinal study will be useful to confirm the prognostic value of the WHO staging system.

Biomarkers↗

PubMed1999

Reduced naive and increased activated CD4 and CD8 cells in healthy adult Ethiopians compared with their Dutch counterparts.

To assess possible differences in immune status, proportions and absolute numbers of subsets of CD4+ and CD8+ T cells were compared between HIV- healthy Ethiopians (n = 52) and HIV- Dutch (n = 60). Both proportions and absolute numbers of naive CD4+ and CD8+ T cells were found to be significantly reduced in HIV Ethiopians compared with HIV- Dutch subjects. Also, both proportions and absolute numbers of the effector CD8+ T cell population as well as the CD4+CD45RA-CD27- and CD8+CD45RA-CD27- T cell populations were increased in Ethiopians. Finally, both proportions and absolute numbers of CD4+ and CD8+ T cells expressing CD28 were significantly reduced in Ethiopians versus Dutch. In addition, the possible association between the described subsets and HIV status was studied by comparing the above 52 HIV- individuals with 32 HIV+ Ethiopians with CD4 counts > 200/microliter and/or no AIDS-defining conditions and 39 HIV+ Ethiopians with CD4 counts < 200/microliter or with AIDS-defining conditions. There was a gradual increase of activated CD4+ and CD8+ T cells, a decrease of CD8+ T cells expressing CD28 and a decrease of effector CD8+ T cells when moving from HIV- to AIDS. Furthermore, a decrease of naive CD8+ T cells and an increase of memory CD8+ T cells in AIDS patients were observed. These results suggest a generally and persistently activated immune system in HIV- Ethiopians. The potential consequences of this are discussed, in relation to HIV infection.

Acquired Immunodeficiency Syndrome↗

PubMed1999

Immunohematological reference ranges for adult Ethiopians.

A cross-sectional survey was carried out with 485 healthy working adult Ethiopians who are participating in a cohort study on the progression of human immunodeficiency virus type 1 (HIV-1) infection to establish hematological reference ranges for adult HIV-negative Ethiopians. In addition, enumeration of absolute numbers and percentages of leukocyte subsets was performed for 142 randomly selected HIV-negative individuals. Immunological results were compared to those of 1,356 healthy HIV-negative Dutch blood donor controls. Immunohematological mean values, medians, and 95th percentile reference ranges were established. Mean values were as follows: leukocyte (WBC) counts, 6.1 x 10(9)/liter (both genders); erythrocyte counts, 5.1 x 10(12)/liter (males) and 4.5 x 10(12)/liter (females); hemoglobin, 16.1 (male) and 14.3 (female) g/dl; hematocrit, 48.3% (male) and 42.0% (female); platelets, 205 x 10(9)/liter (both genders); monocytes, 343/microl; granulocytes, 3, 057/microl; lymphocytes, 1,857/microl; CD4 T cells, 775/microl; CD8 T cells, 747/microl; CD4/CD8 T-cell ratio, 1.2; T cells, 1, 555/microl; B cells, 191/microl; and NK cells, 250/microl. The major conclusions follow. (i) The WBC and platelet values of healthy HIV-negative Ethiopians are lower than the adopted reference values of Ethiopia. (ii) The absolute CD4 T-cell counts of healthy HIV-negative Ethiopians are considerably lower than those of the Dutch controls, while the opposite is true for the absolute CD8 T-cell counts. This results in a significantly reduced CD4/CD8 T-cell ratio for healthy Ethiopians, compared to the ratio for Dutch controls.

Adult↗

PubMed1996

Expression of tyrosine kinase gene in mouse thymic stromal cells.

Amongst the most important signal transduction molecules involved in regulating growth and differentiation are the protein tyrosine kinases (PTK). Since T cell development is a consequence of interactions between thymic stromal cells (TSC) and thymocytes, identification of the PTK in both compartments is required to dissect the mechanisms that control this process. Here we report a search for PTK in mouse TSC, using RT-PCR to survey the repertoire of PTK mRNAs expressed in a freshly isolated TSC preparation. We identified 10 different PTK cDNAs among the 216 cDNAs sequenced, and demonstrate that transcripts of three of those (ufo, fyn and fer) are widely expressed among a large panel of immortalized thymic epithelial cell lines (TEC) and in primary cultures of TSC. Of the other seven, none were expressed in established TEC lines but, instead, displayed distinct expression patterns in cell types likely to have contaminated the fresh TSC preparation, i.e., macrophages, B cells, T cells and fibroblasts. Among the three PTK expressed in TEC lines, only one, ufo, exhibited expression exclusively in cells of non-hemopoietic origin. Although expression of ufo (also known as tyro 7, axl or ark) is not thymic-specific, in that it is also expressed in cell types of mesodermal origin in other tissues, its presence in TEC suggests a role for ufo in differentiation of the TSC compartment. Consistent with this notion, high-level expression of this receptor PTK at the protein level could be documented in every TEC line investigated, as well as in fresh thymus tissue sections. These data provide the first example of a receptor PTK in TSC and open new approaches to study the regulation of TSC differentiation.

Amino Acid Sequence↗

PubMed1995

Immunological and functional characterization of Mycobacterium leprae protein antigens: an overview.

A major focus of leprosy research in the last 10 years has been the identification and characterization of antigens of Mycobacterium leprae that interact with antibodies and T cells of the host's immune response. Through the combined efforts of many different laboratories, a substantial number of protein antigens have been identified and characterized. In this MicroReview we present an updated list of M. leprae protein antigens, and, with emphasis on recent developments, summarize what is known regarding their functional and immunological features.

Antigens, Bacterial↗

PubMed1994

Identification of an antigenic domain on Mycobacterium leprae protein antigen 85B, which is specifically recognized by antibodies from patients with leprosy.

Sixty-three overlapping 15-oligomer peptides covering the 30-kDa protein antigen 85B of Mycobacterium leprae were tested by ELISA to identify epitopes recognized by human antibodies. Serum samples from patients with lepromatous leprosy (LL) reacted mainly with peptides comprising amino acid regions (AA) 206-230, 251-280, and 291-325. Sera of patients with active tuberculosis who responded to the native 30-kDa antigen did not recognize these peptides. The antibody-binding specificity to the defined B cell regions was evaluated in a blind study with 71 serum samples from patients and household contacts living in Ethiopia where leprosy is endemic. The peptide of AA 256-280 was recognized by 88% of LL patients, 15% of patients with tuberculoid leprosy, and none of the contacts. These findings suggest that there are major linear B cell epitopes on the M. leprae 30-kDa protein that are recognized by lepromin-negative LL patients, whereas lepromin-positive patients respond preferentially to conformational epitopes.

Amino Acid Sequence↗

PubMed1993

A Mycobacterium leprae-specific gene encoding an immunologically recognized 45 kDa protein.

By screening a Mycobacterium leprae lambda gt11 expression library with a serum from an Ethiopian lepromatous leprosy (LL) patient a clone was isolated (LL4) belonging to hybridization group III of a panel of previously isolated M. leprae clones. Members of this hybridization group encode a serologically recognized 45 kDa protein. The complete DNA sequences of the partially overlapping clones LL4 and L1 (hybridization group III) are presented and these revealed the presence of an open reading frame (ORF) predicting a protein with a molecular size of 42,448 Da. Southern hybridizations on total genomic DNA of M. leprae, Mycobacterium tuberculosis and eight atypical mycobacteria showed that the LL4 DNA fragment is specific for M. leprae DNA even under low-stringency conditions. The M. leprae specificity of LL4 DNA was further confirmed by the polymerase chain reaction using four different sets of primers. Western blotting analyses showed that the M. leprae 45 kDa protein is frequently recognized by antibodies from leprosy patients and that this recognition is specific since no antibodies could be detected in sera of tuberculosis patients. T-cell proliferation assays also demonstrated T-cell recognition by leprosy patients and healthy contacts of the M. leprae 45 kDa protein. The specificity of the LL4 DNA region and the 45 kDa antigen that is encoded by hybridization group III could provide unique tools for the development of M. leprae-specific immunological and DNA reagents.

Amino Acid Sequence↗

PubMed1993

The Mycobacterium leprae antigen 85 complex gene family: identification of the genes for the 85A, 85C, and related MPT51 proteins.

The genes for two novel members (designated 85A and 85C) of the Mycobacterium leprae antigen 85 complex family of proteins and the gene for the closely related M. leprae MPT51 protein were isolated. The complete DNA sequence of the M. leprae 85C gene and partial sequences of the 85A and MPT51 genes are presented. As in M. tuberculosis, the M. leprae 85A, 85C, and previously identified 85B component genes are not closely linked on the genome. However, the MPT51 genes of both species localize close to the respective 85A component genes. Like the 85B component, the M. leprae 85A-MPT51 and 85C antigens are recognized by T cells from healthy contacts and leprosy patients.

Amino Acid Sequence↗

PubMed1992

Mycobacteria contain two groEL genes: the second Mycobacterium leprae groEL gene is arranged in an operon with groES.

In contrast to other bacterial species, mycobacteria were thus far considered to contain groEL and groES genes that are present on separate loci on their chromosomes, Here, by screening a Mycobacterium leprae lambda gt11 expression library with serum from an Ethiopian lepromatous leprosy patient, two DNA clones were isolated that contain a groEL gene arranged in an operon with a groES gene. The complete DNA sequence of this groESL operon was determined. The predicted amino acid sequences of the GroES and GroEL proteins encoded by this operon are 85-90% and 59-61% homologous to the sequences from previously characterized mycobacterial GroES and GroEL proteins. Southern blotting analyses with M. leprae groES- and groEL-specific probes demonstrate that similar groESL homologous DNA is present in the genomes of other mycobacteria, including Mycobacterium tuberculosis. This strongly suggests that mycobacteria contain a groESL operon in addition to a separately arranged second groEL gene. Using five T-cell clones from two leprosy patients as probes, expression of the M. leprae GroES protein in Escherichia coli after heat shock was demonstrated. Four of these clones recognized the same M. leprae-specific GroES-derived peptide in a DR2-restricted fashion. No expression of the groEL gene from this operon was detected in E. coli after heat shock, as tested with a panel of T-cell clones and monoclonal antibodies reactive to previously described GroEL proteins of mycobacteria.

Amino Acid Sequence↗

PubMed1991

Monoclonal antibodies to human embryonal carcinoma cells: antigenic relationships of germ cell tumors.

Fifteen monoclonal antibodies (mAb) that show specificity for human embryonal carcinoma cells are described. C57BL/6 mice were immunized with Tera-2 embryonal carcinoma cells, and hybridomas were isolated and tested versus a set of human developmental tumor cell lines. The antigens exhibit relatively restricted and unique distributions on normal tissues as shown by immunohistochemistry. The mAbs recognize a series of differentiation antigens since their expression changes when teratocarcinoma stem cells are induced to differentiate in vitro. The expression of these molecules in germ cell and related tumors is consistent with the data obtained from in vitro cell studies. Seven of these mAbs immunoprecipitate molecules from surface labeled Tera-2 cells that show distinct molecular weights on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The antigenic phenotypes defined by the set of mAbs can be used to investigate possible pathogenetic relationships of the various testicular tumors.

Animals↗

PubMed1991

The role of MHC class I expression in developmental tumours.

Expression of HLA class I (-like) genes by two developmental tumour cell lines representing embryonic (Tera-2) and extra-embryonic (Jeg-3) origins is reviewed. The Tera-2 embryonal carcinoma cells are HLA negative but can be induced by gamma interferon to express HLA-A, -B, -C, and apparently -E and G genes. Jeg-3 choriocarcinoma cells constitutively express HLA-G and low levels of a novel HLA-C gene product. The NK sensitivity of the developmental tumour cell types has been assessed and the biochemical characteristics of the surface molecules expressed by the trophoblast derived cells has been further documented. The biological role of these novel HLA molecules in the context of the maternal acceptance of the fetal-semi-allograft is discussed.

Antibodies, Monoclonal↗