Biomedical subjects
Publications and source records attributed to Esper Georges Kallas.
Three HIV-1-seronegative patients with disseminated tuberculosis presented significant depletion of T-cell counts, in CD4+ and/or CD8+ cells, associated with increased expression of activation marker CD38 on CD8+ T-lymphocytes. This finding raises the question of potential mechanisms involved in the activation or loss of T-cells in disseminated tuberculosis.
BACKGROUND: Monitoring recipient's alloreactivity has shown to be critical for limiting overimmunosuppression besides allowing preemptive treatment of acute rejection (AR). METHODS: Flow cytometry and real time RT-PCR were performed in urine of kidney transplant recipients with AR (n = 13) and compared with pyelonephritis (n = 10), chronic allograft nephropathy (n = 13), acute tubular necrosis (n = 13) and stable graft function (n = 11). Expression of CD3, CD4, CD8, HLA-DR, Fas-L, ICAM-1 and CD25 were assessed using flow cytometry. mRNA of perforin, granzyme B and Fas-L were quantified by real time RT-PCR. RESULTS: Frequencies of CD3+, HLA-DR+, Fas-L+, ICAM-1+ and CD25+ cells were significantly higher in AR group (p < 0.05). ROC curves showed sensitivity from 70% to 91% and specificity from 30% to 100%, whereas the highest sensitivity and specificity was 91% and 100% respectively, for Fas-L+ cells. Levels of mRNA of perforin, granzyme B and Fas-L were significantly augmented in AR, while the sensitivity and specificity ranged from 85% to 88% and from 55% to 100%, respectively. CONCLUSIONS: Analyses of immune activation markers by flow cytometry and real time RT-PCR are equally useful for noninvasive monitoring kidney allografts.
BACKGROUND: Previous studies have reported that incubation of polymorphonuclear cells (PMN) in uraemic plasma or with different haemodialysis membranes and peritoneal dialysis solutions increases apoptosis in this cell type. In addition, PMN harvested from uraemic patients show a reduced ability to generate superoxide in response to stimuli as well as impaired phagocytosis, chemotaxis and degranulation. The aim of the current study was to investigate the effect of uraemia and dialysis modality on apoptosis and function in freshly harvested non-incubated PMN. METHODS: Polymorphonuclear cells were harvested from 14 chronic haemodialysis (HD) patients, from 14 continuous peritoneal dialysis patients (CAPD), 28 chronic kidney disease (CKD), pre-dialysis patients and from 14 healthy subjects (Controls). In these in vivo experiments, PMN apoptosis was studied by means of flow cytometric analysis of annexin V binding to freshly isolated cells. Polymorphonuclear cell phagocytosis and production of reactive oxygen species by unstimulated or stimulated (S.aureus, fMLP, PMA) cells were also studied by flow cytometry using whole blood. RESULTS: We observed increased PMN apoptosis in CKD patients. CAPD and HD patients displayed PMN apoptosis rates similar to controls. In the HD group, PMN exhibited decreased phagocytosis rates. In contrast, phagocytosis rates in PMN from CAPD were not significantly different from controls. In the CKD and HD groups, apoptosis was inversely correlated with respiratory burst activity and phagocytosis. CONCLUSION: Our results suggest that both uraemia and treatment modality may interfere with PMN apoptosis and function. Dialysis appears to normalize the increased PMN apoptosis rates observed in CKD patients.
An interesting interaction pattern has been found between HIV-1 and GBV-C/HGV, resulting in protection against progression to AIDS. The mechanisms involved in this interaction remain to be clarified. We examined the current knowledge concerning this coinfection and developed hypotheses to explain its effects. A better understanding of this interaction could result in new concepts, which may lead to new strategies to control HIV-1 replication and progression to AIDS.
Several strategies aim at characterizing the AIDS epidemic in different parts of the world. Among these, the identification of recent HIV-1 infections using the recently described serologic testing algorithm for recent human immunodeficiency virus (HIV) seroconversion (STARHS) strategy was employed in four testing sites of the City of Sao Paulo Public Health Department (CSPPHD). Those identified as recently infected were invited to participate in a prospective clinical and laboratory evaluation study. We describe the establishment of the patient identification network and the success in enrolling the participants, as well as their clinical and laboratory characteristics. From May to December 2002, 6,443 persons were tested for HIV in the four participating sites, of whom 384 (5.96%) tested HIV-1 positive; 43 (11.2%) of them were identified as recently infected. Twenty-two were successfully enrolled in the follow-up study, but three of them did not meet clinical and/or laboratory criteria for recent HIV-1 infection. After these exclusions, the laboratory findings revealed a median CD4+ T lymphocyte count of 585 cells/microL (inter-quartile range 25-75% [IQR], 372-754), a CD8+ T lymphocyte count of 886 cells/microL (IQR, 553-1098), a viral load of 11,000 HIV-RNA copies/mL (IQR, 3,650-78,150), log10 of 4.04 (IQR 3.56-4.88). The identification of recent HIV infections is an extremely valuable way to evaluate the spread of the virus in a given population, especially when cohort studies, considered the gold standard method to evaluate incidence, are not available. This work demonstrated that establishing a network to identify such patients is a feasible task, even considering the difficulties in a large, resource-limited country or city.
After the identification of HIV-2 in 1986, most of the cases reported have been concentrated in West Africa. We identified a case of HIV-2 infection in São Paulo, Brazil of a 45-year-old female who presented with Pneumocystis carinii pneumonia, with a CD4 count of 22 cells/ml. DNA samples from this patient were subjected to end-point PCR amplification of the LTR region. Clones were sequenced and subjected to phylogenetic analyses. All clones were subtype A related, and four presented an insertion, corresponding to an extra NF-kappaB site. This is the first confirmed case report of an HIV-2-infected subject identified in Brazil whose transmission occurred within the country. Furthermore, the NF-kappaB duplication would potentially be associated with an increase in viral cytopathogenicity. This raises concern for the need for permanent monitoring of the spread of HIV-2 in different areas of the world, even considering its lower rate of transmission and pathogenicity when compared to HIV-1.
BACKGROUND: Apoptosis is one of the possible explanations for the progressive loss of CD4(+) T lymphocytes in infection with the human immunodeficiency virus (HIV), which may interfere with cell cycle distribution. This study evaluated the cell cycle of CD4(+) and CD8(+) lymphocytes in HIV-infected subjects and controls. METHODS: Two methods to identify lymphocytes for cell cycle analysis were evaluated, magnetic beads and concurrent staining, and both were followed by propidium iodide DNA labeling. The chosen method was used to evaluate the cell cycle of lymphocytes in HIV-1-infected subjects and controls. RESULTS: There was no significant difference between the two methods, although a higher variability was observed with the magnetic bead cell separation method. A higher proportion of cells in the S phase was observed in HIV-1 patients (2.69% vs. 1.19%, P = 0.016), coupled with a decrease in G(1) (96.11% vs. 98.10%, P = 0.005) in CD4(+) lymphocytes, a phenomenon not observed in CD8(+) lymphocytes. No correlation was detected between the different cell cycle phases and T-lymphocyte counts or viral load. CONCLUSIONS: The present work developed a new approach to evaluate lymphocyte cell cycle distribution, applied in the setting of HIV-1 infection. It may contribute to the understanding of the CD4(+) T-lymphocytes depletion seen in these patients.
Human immunodeficiency virus type 1 (HIV-1) and type 2 (HIV-2) are the causative agents of AIDS. HIV-2 is prevalent at moderate to high rates in West African countries, such as Senegal, Guinea, Gambia, and Cape Verde. Diagnosis of HIV-2 is made with a positive HIV-1/HIV-2 ELISA or simple/rapid assay, followed by one or two confirmatory tests specific for HIV-2. Following CD(4)(+)T cell counts, HIV-2 viral burden and clinical signs and symptoms of immunodeficiency are beneficial in monitoring HIV-2 disease progression. Although non-nucleoside reverse transcriptase inhibitors are ineffective in treating HIV-2, nucleoside reverse transcriptase inhibitors and protease inhibitors can be effective in dual and triple antiretroviral regimens. Their use can decrease HIV-2 viral load, increase CD(4)(+)T cell counts and improve AIDS-related symptoms. HIV-2 resistance to various nucleoside reverse transcriptase inhibitors and protease inhibitors, including zidovudine, lamivudine, ritonivir and indinavir, has been identified in some HIV-2 infected patients on antiretroviral therapy. The knowledge of HIV-2 peculiarities, when compared to HIV-1, is crucial to helping diagnose and guide the clinician in the choice of the initial antiretroviral regimen and for monitoring therapy success.
We evaluated neutrophil activation by measuring its phagocytic ability and oxidative burst activity in 16 patients with sepsis and 16 healthy volunteers. We also focused on neutrophil apoptosis as a regulatory mechanism of the inflammatory response. Neutrophil phagocytosis was evaluated by the detection of propidium iodide (PI)-labeled Staphylococcus aureus added to whole blood. Reactive oxygen species (ROS) formation was quantified by measuring the oxidation of 2',7' dichlorofluorescein diacetate (DCFH-DA) at baseline and after cell stimulation with phorbol myristate acetate (PMA), and bacterial cells (killed S. aureus) or products (lipopolysaccharide [LPS] and N-formyl-methionyl-leucyl-phenylalanine [FMLP]). Apoptosis was assessed in neutrophils stained with annexin V and PI. Neutrophil phagocytic ability was increased in patients with sepsis compared with healthy controls (median geometric mean fluorescence intensity [GMFI] was 101.9 and 54.7, respectively; P = 0.05). ROS formation was enhanced in patients with sepsis compared with healthy volunteers at baseline (median GMFI 275.6 and 52.1, respectively; P < 0.001), and after stimulation with S. aureus (median GMFI 2395.8 and 454.9, respectively; P < 0.001), PMA (median GMFI 1120.6 and 307.5, respectively; P = 0.003), FMLP (median GMFI 792.4 and 123.2, respectively; P < 0.001), and LPS (median GMFI 624.8 and 144.8, respectively; P < 0.001). Early neutrophil apoptosis was increased in patients with sepsis compared with healthy volunteers (median 11.3% and 9.1%, respectively; P = 0.03). These data demonstrate that neutrophil function is enhanced in patients with sepsis. Additionally, circulating neutrophils from patients with sepsis presented with increased early apoptosis, which may be consequence of a regulatory mechanism of the inflammatory response.
CD(4)(+) and CD(8)(+) T lymphocyte counts, naive and memory/effector CD(4)(+) T subpopulations, and the expression of CD(38) on CD(8)(+) T lymphocytes were evaluated in four groups: AIDS patients with tuberculosis (HIV/TB, n=14), HIV-1 infected patients (HIV, n=10), HIV-1 negative patients with tuberculosis (TB, n=20) and healthy controls (CTL, n=17). TB and HIV had fewer +CD(4)(+) T cells than CTL, with the lowest values observed in TB/HIV (p<0.001). No difference between groups was observed in the percentage of naive and memory/effector subpopulations in +CD(4)(+) T lymphocytes. TB (355 cells/ micro L) and HIV (517 cells/ micro L) had diverging effects on +CD(8)(+) T cell counts, with a marked depletion observed in HIV/TB (196 cells/ micro L). TB and HIV up-regulated CD(38) expression on CD(8)(+) T cells, a finding also present in TB/HIV. While the decrease of +CD(4)(+) T cell counts in HIV/TB may be attributed to HIV and tuberculosis, the decrease of +CD(8)(+) T cell counts is likely to be due to tuberculosis.
Leptospira interrogans glycolipoprotein (GLP) has been implicated in pathological and functional derangement seen in leptospirosis. The goal of this study was to evaluate GLP's ability to induce cellular activation, as assessed by cytokine production and expression of surface activation markers. GLP extracted from either pathogenic L. interrogans serovar Copenhageni or nonpathogenic Leptospira biflexa serovar Patoc (GLPp) was used to stimulate peripheral blood mononuclear cell cultures from healthy donors. Supernatant cytokine levels were measured by enzyme-linked immunosorbent assay. Expression of CD69 and HLA-DR on lymphocytes and monocytes, as well as lipopolysaccharide (LPS) binding, were measured by flow cytometry. At 6 h of incubation, GLP induced a significant rise in tumor necrosis factor alpha levels, which dropped progressively until 72 h of incubation. Interleukin-10 peak levels were obtained at between 24 and 48 h, with sustained levels until 72 h of incubation. The response magnitude was proportional to the GLP dose. CD69 expression on T lymphocytes and monocytes increased significantly, as did HLA-DR expression on monocytes. GLPp induced no CD69 or HLA-DR expression. GLP did not block biotinylated LPS binding to monocytes, suggesting that different pathways are used to induce cell activation. In conclusion, GLP induces cellular activation and may play a major role in the pathogenesis of leptospirosis.