Sensitivity and Specificity of Standard and Rapid HIV-Antibody Tests Evaluated by Seroconversion and Non-Seroconversion Low-Titre Panels

Background and objectives: The aim of this study is to compare the relative sensitivity and specificity of commercial HIV-antibody assays using seroconversion, non-seroconversion panels, and negative blood donor samples. Materials and methods: We evaluated the sensitivity of five standard ELISA HIV-antibody assays: Vironostika HIV Uni-Form II, Abbott recombinant HIV-1/HIV-2 third-generation EIA, Biotest Anti-HIV-1/-2 recombinant, Recombigen HIV-1/HIV-2 EIA and Wellcozyme HIV 1+2 (VK54/55), and three rapid screening tests, Capillus HIV-1/HIV-2, Abbott Test Pack HIV-1/HIV-2 third-generation EIA, and Sensy-Test HIV 1/2. All tests were assessed using four panels of plasma samples obtained from individuals who were seroconverting and a low-titre HIV-antibody panel of samples. Specificity of the standard screening tests was determined on 3,500 HIV-antibody-negative blood donor samples. Results: There was no statistically significant difference in sensitivity between the five standard ELISA tests. One of these tests was significantly less specific than the others. The standard ELISA tests detected all the low-titre HIV-antibody-positive samples. Two of the rapid screening tests were significantly less sensitive on the seroconversion panels and all three tests failed to detect at least one of the positive samples in the low-titre panel. Conclusions: The additional risk of using one or other of the standard ELISA tests under review of not detecting all HIV-positive units of blood is not statistically significant. Using some of the rapid screening tests will, however, add a significant additional risk. A rapid screening test should therefore be adopted only after careful consideration of the effect of a possible lack of sensitivity on the safety of the blood supply.     

Levels of soluble E-selectin in patients with active Behçet’s disease

Behçets disease is a systemic vasculitis of unknown aetiology. Endothelial cell injury plays an important role in the pathogenesis and immunopathology of Behçets disease. E-selectin is expressed by activated endothelial cells. Because the selectin adhesion molecules are shed from activated cells, soluble forms of these proteins can be used as activation markers of endothelium (E-selectin). The pathogenesis of Behçets disease (BD) is closely related to endothelial cells, leucocyte functions and immunity. The aim of this study was to investigate circulating E-selectin adhesion molecules, which are known to play a significant part in the immune response especially by regulating interaction of the leucocytes with endothelium in BD. Plasma E-selectin concentrations were evaluated in 23 patients with BD and 20 healthy control subjects. The disease activity was evaluated by clinical manifestations (oral aphthous ulcer, genital ulceration, positive pathergy test, skin lesions, eye involvement, thrombophlebitis and arthritis) and by laboratory investigations [erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP)]. The patients were newly or previously diagnosed cases not taking any drug for BD. Levels of E-selectin were measured with commercially available sandwich enzyme-linked immunosorbent assay (ELISA) kits using human sE-selectin (cat. no: BMS 205). Plasma E-selectin concentrations of patients and controls were compared with the Mann-Whitney U test. Statistical significance was assigned to p values lower than 0.05. Serum levels (mean±SD) of soluble E-selectin (sE-selectin) were significantly higher in 23 patients with BD than in 20 healthy controls (53.2±18.2 ng/ml vs 33.8±7.5 ng/ml, p<0.0001). A statistically significant positive correlation was observed between sE-selectin levels and CRP and ESR in patients with BD (r=0.78, p<0.001 and r=0.56, p<0.01, respectively). Increases in the E-selectin in BD may be a direct consequence of the leucocyte and endothelium activations observed during the disease process. The noninvasive investigations can be used as biochemical markers for inflammation. This may provide additional information regarding disease activity along with the traditional indices such as ESR and CRP.     

CD52 antibodies for prevention of graft-versus-host disease and graft rejection following transplantation of allogeneic peripheral blood stem cells

Graft-versus-host disease (GVHD) is a major cause of mortality and morbidity after allogeneic bone marrow transplantation, but can be avoided by removing T lymphocytes from the donor bone marrow. However, T cell depletion increases the risk of graft rejection. In this study, two strategies are used to overcome rejection: (1) use of high doses of stem cells obtained from peripheral blood (PBSC), (2) admixture with a CD52 monoclonal antibody in order to deplete both donor and residual recipient lymphocytes. Two antibodies are compared: CAMPATH-1G (rat IgG2b) and its humanized equivalent CAMPATH-1H (human IgG1). A total of 187 consecutive patients at six centers received PBSC transplants from HLA-matched siblings between 1997 and 1999. A wide spectrum of diseases, both malignant and non-malignant, was included. The recovery of CD34+ cells after antibody treatment was close to 100%. The risk of acute GVHD (grade 2 to 4) was 11% in the CAMPATH-1G group and 4% in the CAMPATH-1H group (P = NS). The risk of chronic GVHD (any grade) was 11% in the CAMPATH-1G group and 24% in the CAMPATH-1H group (P = 0.03) but the risk of extensive chronic GVHD was only 2%. The overall risk of graft failure/rejection was 2%, not significantly different between the two antibodies. Antibody treatment was equally effective at concentrations between 10 microg/ml and 120 microg/ml and it made no significant difference to the outcome whether the patients received post-transplant immunosuppression or not (87% did not). Transplant-related mortality in this heterogenous group of patients (including high-risk and advanced disease) was 22% at 12 months. It is proposed that treatment of peripheral blood stem cells with CAMPATH-1H is a simple and effective method for depleting T cells which may be applicable to both autologous and allogeneic transplants from related or unrelated donors. Special advantages of this approach are the simultaneous depletion of donor B cells (which reduces the risk of EBV-associated lymphoproliferative disease) and the concomitant infusion of CAMPATH-1H to deplete residual recipient T cells and thus prevent graft rejection.     

Regulation of the luminal Na+/H+ exchanger NHE3 by intracellular protein trafficking

The article summarizes some of the recent developments in the understanding of the mechanisms of regulation of the proximal tubule apical membrane Na+/H+ antiporter NHE3. NHE3 antiporter has a major role in HCO3- and NaCl reabsorption in the proximal tubule. NHE3 protein is associated with the regulatory factor NHERF which interacts with ezrin, an actin-binding protein. This multi-protein complex constitutes a link between a membrane protein, NHE3, and actin cytoskeleton. Cytoskeleton organization has a key role to control NHE3 activity under normal conditions. Pharmacological perturbations of actin polymerization interfere with NHE3 activity. Parathyroid hormone-induced NHE3 activity inhibition results first, from a protein kinase A-mediated phosphorylation without protein trafficking, and then from endocytosis involving dynamin. The stimulatory effect of systemic angiotensin II concentrations on NHE3 activity is protein kinase C-dependent and results, at least in part, from exocytic insertion of the protein in luminal membranes. It requires cytoskeleton integrity.     

Relationship of SARS-CoV-2–specific CD4 response to COVID-19 severity and impact of HIV-1 and tuberculosis coinfection

T cells are involved in control of coronavirus disease 2019 (COVID-19), but limited knowledge is available on the relationship between antigen-specific T cell response and disease severity. Here, we used flow cytometry to assess the magnitude, function, and phenotype of SARS coronavirus 2–specific (SARS-CoV-2–specific) CD4⁺ T cells in 95 hospitalized COVID-19 patients, 38 of them being HIV-1 and/or tuberculosis (TB) coinfected, and 38 non–COVID-19 patients. We showed that SARS-CoV-2–specific CD4⁺ T cell attributes, rather than magnitude, were associated with disease severity, with severe disease being characterized by poor polyfunctional potential, reduced proliferation capacity, and enhanced HLA-DR expression. Moreover, HIV-1 and TB coinfection skewed the SARS-CoV-2 T cell response. HIV-1–mediated CD4⁺ T cell depletion associated with suboptimal T cell and humoral immune responses to SARS-CoV-2, and a decrease in the polyfunctional capacity of SARS-CoV-2–specific CD4⁺ T cells was observed in COVID-19 patients with active TB. Our results also revealed that COVID-19 patients displayed reduced frequency of Mycobacterium tuberculosis–specific CD4⁺ T cells, with possible implications for TB disease progression. These results corroborate the important role of SARS-CoV-2–specific T cells in COVID-19 pathogenesis and support the concept of altered T cell functions in patients with severe disease.     

Alloimmunization to platelets in heavily transfused patients with sickle cell disease

Bone marrow transplantation (BMT) is now an option for some patients with sickle cell disease (SCD). Many SCD patients are multiply transfused with red blood cells (RBCs), and may be immunized to alloantigens other than erythrocyte antigens. Because platelet refractoriness is a significant complication during BMT, we wished to determine the prevalence of alloimmunization to platelets in transfused SCD patients. Sera collected from 47 transfused and 14 untransfused SCD patients were screened for HLA and platelet-specific antibodies. Transfusion and RBC antibody histories were reviewed. A subset of the patients were rescreened 1 year later. Eighty-five percent of patients with at least 50 RBC transfusions (22 of 26), 48% of patients with less than 50 transfusions (10 of 21), and none of 14 untransfused patients demonstrated platelet alloimmunization (P < .05). Platelet alloimmunization was more prevalent than RBC alloimmunization (20% to 30%). Half of the platelet reactivity was chloroquine-elutable. Eighteen of 22 patients (82%) on chronic RBC transfusion remained platelet-alloimmunized 11 to 22 months after initial testing. In summary, 85% of heavily transfused SCD patients are alloimmunized to HLA and/or platelet-specific antigens. These patients may be refractory to platelet transfusion, a condition that would increase their risk during BMT. Leukodepletion in the transfusion support of SCD patients should be considered to prevent platelet alloimmunization.     

Immunohistochemical localization of membrane and alpha-granule proteins in human megakaryocytes: application to plastic-embedded bone marrow biopsy specimens

Using a new technique for antigen localization, we have demonstrated platelet proteins in megakaryocytes in plastic-embedded biopsy specimens of normal human bone marrow. In a series of 25 specimens, megakaryocytes showed labeling with antibodies to the integral membrane glycoproteins IIIa, IIb, and the IIb-IIIa complex; granule membrane protein 140; and five alpha-granule matrix proteins: thrombospondin, factor VIII-related antigen, beta-thromboglobulin, platelet factor 4, and fibrinogen. The antibodies to the membrane glycoproteins IIIa, IIb, and IIb-IIIa produced diffuse cytoplasmic staining and heavier staining on the plasma membrane, whereas the antibodies to the alpha-granule matrix proteins produced a distinct granular staining within the cytoplasm. Staining for granule membrane protein 140 was also granular in distribution. Rare mononuclear cells consistent with megakaryocyte precursors were labeled with these markers. Other enzyme histochemical and lectin-binding studies showed that the enzyme alpha-naphthyl acetate esterase, the lectin Ulex europaeus I, and the periodic-acid Schiff reaction were consistent, but not specific, markers of megakaryocytes. This immunohistochemical technique should facilitate the examination of qualitative and quantitative changes in megakaryocytes in a variety of physiologic and pathologic processes.     

Posttraumatic incarceration of medial collateral ligament into knee joint with anterior cruciate ligament injury

Medial collateral ligament of the knee is an important coronal stabiliser and often injured in isolation or as combination of injuries. The article reports a case of incarcerated medial collateral ligament (MCL) injury in combination with anterior cruciate ligament (ACL) injury in 20 year old male who presented to us 4 weeks after injury. Clinical examination and MRI was correlated to complete ACL tear with torn distal MCL and incarceration into the joint. Patient was taken up for ACL hamstring graft reconstruction with mini-arthrotomy and repair of the torn MCL. Patient was followed up with dedicated rehabilitation protocol with good functional results. At one year follow-up, patient exhibited full range of motion with negative Lachman, Pivot shift and valgus stress tests. This article highlights the rare pattern of MCL tear and also reviews the literature on this pattern of injury.