Evolution of HIV-1 in an HLA-B*57-positive patient during virologic escape

Elite suppressors maintain normal CD4(+) T cell counts and viral loads of <50 copies of human immunodeficiency virus type 1 (HIV-1) RNA per milliliter of plasma without antiviral therapy. We report here a case of virologic escape in a human leukocyte antigen (HLA)-B*57-positive patient shortly after seroconversion. This escape was associated with the development of mutations in 2 HLA-B*57-restricted CD8(+) T cell Gag epitopes, reversion of the drug-resistance mutation M184V, and reversion of a novel polymorphism in Vpu. The present study suggests that control of viral replication in elite suppressors may be due to HIV-1-specific CD8(+) T cells and, in some cases, mutations that have subtle effects on viral fitness.     

Cationic Lipid-Mediated Transfection of Differentiated Caco-2 Cells: A Filter Culture Model of Gene Delivery to a Polarized Epithelium

Purpose. The use of rapidly dividing in vitro cell culture systems to assess the efficiency of gene delivery is now recognised as a poor indicator of in vivo success. We investigated whether differentiated Caco-2 cell filter-cultures would make a more suitable model for studying gene transfer to an epithelium. Methods. Caco-2 cells were cultured on semi-permeable membrane filters into differentiated polarised monolayers. Monolayer differentiation was assessed by monitoring the transport of taurocholic acid. Cells at different stages of differentiation were transfected with DNA/DOTAP lipoplexes and later analysed for reporter gene activity. The uptake of radiolabled DNA was also evaluated at various stages of differentiation. Results. Caco-2 cultures developed a resistance to lipoplex-mediated transfection as early as three days, when some cells were still dividing and undifferentiated. As cultures matured, expression of reporter gene progressively decreased partly due to reduced internalisation of DNA. The resistance to transfection could be overcome in part by pre-treatment of monolayers with calcium chelating agents or surfactants. However, transgene expression in treated monolayers was still significantly lower than that in dividing cultures. Conclusions. Differentiated Caco-2 cells are a more appropriate model for gene-transfer studies to the intestinal epithelium because they demonstrate a resistance to transfection similar to that observed in vivo.     

Venous thromboembolism in patients with HIV/AIDS: a case-control study

BACKGROUND: Retrospective cohort studies of HIV-infected patients suggest an incidence of venous thromboembolism (VTE) of 1% to 2%, which is 10 times that expected among people without HIV. We investigated the prevalence of established risk factors for VTE in this population and explored novel risk factors. METHODS: We conducted a case-control study using patients in the Johns Hopkins University AIDS Service cohort. We used conditional logistic regression and paired t tests to test for covariates significantly associated with VTE. RESULTS: We identified 160 patients with VTE diagnosed radiologically or with a clinical course consistent with VTE; 23% of the cases of VTE were diagnosed in hospitalized patients. The incidence of VTE was approximately 0.5% per patient-year. Patients with VTE and control patients did not differ by gender, but black patients were overrepresented among those with VTE (odds ratio [OR]=1.9, 95% confidence interval [CI]: 1.11 to 3.08) and patients with VTE were older than controls (mean: 39 vs. 37 years; P=0.001). Patients with VTE had lower CD4 counts (229 vs. 362 cells/mm; P<0.0001), higher HIV RNA titers (120,254 vs. 71,262 copies/mL; P=0.013), and lower hemoglobin concentrations (11.4 vs. 12.7 g/dL; P<0.0001) preceding the event than those without VTE. The use of highly active antiretroviral therapy was not associated with VTE. In multivariate analyses, independent risks for VTE were age, hospitalization in the past 3 months (OR=13, 95% CI: 6.4 to 27), central venous catheter use in the past 3 months (OR=6.0, 95% CI: 2.3 to 16), and a CD4 count<500 cells/mm (OR=3.0, 95% CI: 1.2 to 7.8). CONCLUSIONS: The incidence of VTE in our cohort is similar to that reported in other cohorts of patients with HIV. Recent hospitalization was the risk factor most strongly associated with VTE.     

Passenger lymphocyte syndrome in liver transplant recipients: a description of 12 cases

BackgroundPassenger lymphocyte syndrome is an important cause of immune haemolysis after solid organ transplantation. It mainly occurs in minor ABO and Rh mismatched transplants. The haemolysis is usually mild and self-limited. We present our experience in passenger lymphocyte syndrome and liver transplantation and review the literature.ResultsA total of 1,217 liver transplants were performed and 12 passenger lymphocyte syndromes were detected: of the 56 cases with minor ABO incompatibility, ten patients developed passenger lymphocyte syndrome (17.9%) and of 147 cases with minor Rh incompatibility, two patients developed the syndrome (1.40%). All patients with passenger lymphocyte syndrome had haemolysis, a decrease of haemoglobin (median 6.8 g/dL) and an increase of bilirubin (median 5.15 mg/dL). The treatment of passenger lymphocyte syndrome consisted of increasing the dose of corticosteroids that the patients were receiving as post-transplantation immunosuppressive therapy and, in the majority of cases, transfusion of donor compatible red blood cells.DiscussionPassenger lymphocyte syndrome in liver transplantation has significant clinical consequences. It is, therefore, important to make the diagnosis rapidly, performing pre-transfusion direct antiglobulin tests, and manage the problem correctly with donor compatible red blood cell transfusions and/or immunosuppressive treatment.     

Slow human immunodeficiency virus type 1 evolution in viral reservoirs in infants treated with effective antiretroviral therapy

A longitudinal study of viral reservoirs in children initiating highly active antiretroviral therapy (HAART) in early infancy was undertaken to test the hypothesis that early effective treatment affects the persistence of replication-competent viral latency and the evolution of HIV-1 in resting CD4(+) T cells. An end point dilution culture assay was used to measure the frequencies of latently-infected resting CD4(+) T cells harboring replication-competent virus in early and late treated children. Gag, pol, and env also were sequenced and compared to pretreatment sequences. HIV-1-specific humoral and cellular immune responses were also assessed. Blood samples were obtained from 12 HIV-1-infected children who started HAART at a median of 1.9 months of age and who maintained suppression of HIV-1 replication for up to 5.5 years. Replication-competent HIV-1 was recovered from 10/12 (84%) subjects. Evolution in gag, pol, and env was restricted for years in early-treated children. HAART initiated from early infancy does not prevent the establishment of a reservoir of latent provirus, but does significantly limit the evolution of HIV-1 in viral reservoirs. The effect of early therapy on HIV-1 evolution may have implications for long-term pharmacologic control of HIV-1.     

The role of bone marrow biopsy and FDG‐PET/CT in identifying bone marrow infiltration in the initial diagnosis of high grade non‐Hodgkin B‐cell lymphoma and Hodgkin lymphoma. accuracy in a multicenter series of 372 patients

Bone marrow infiltration (BMI), categorized as an extra‐nodal site, affects stage and is associated with poor prognosis in newly diagnosed lymphoma patients. We have evaluated the accuracy of PET/CT and bone marrow biopsy (BMB) to assess BMI in 372 lymphoma patients [140 Hodgkin Lymphoma (HL) and 232 High Grade B‐cell non‐Hodgkin Lymphoma (HG B‐NHL), among them 155 Diffuse Large B‐Cell Lymphoma (DLCL)]. For HL cases, and taking into account PET/CT, sensitivity, negative predictive value (NPV) and accuracy were 96.7, 99.3, and 99.3% while those of BMB were 32.3, 83.8, and 85%, respectively. For HG B‐NHL and considering PET/CT, sensitivity, NPV, and accuracy were 52.7, 81.7, and 84.1%, while those of BMB were 77.6, 90.2, and 90.7%, respectively. In the HG B‐NHL group, 25 patients would have been under‐staged without BMB. These results lead us to recommend PET/CT and the avoidance of BMB to assess BMI in HL. In the case of HG B‐NHL, bone marrow status should be assessed firstly by means of PET/CT; only in either focal or diffuse PET/CT with low borderline SUV max values or in negative cases, should BMB be carried out afterwards. In the HG B‐NHL setting and at the present moment, both techniques are complementary. Am. J. Hematol. 90:686–690, 2015. © 2015 Wiley Periodicals, Inc.     

Efficiency of non-viral gene delivery systems to rat lungs.

OBJECTIVE: Transient expression of therapeutic genes within lung allografts may modulate the pathological processes following allotransplantation. Whilst efficient gene transfer to lungs has been reported with viral vectors, their usefulness is limited on the grounds of safety. Since non-viral systems overcome many of these safety issues, our studies were designed to evaluate the efficiency of several non-viral gene delivery vectors for in vivo transfer of plasmid DNA to rat lungs via the airways. METHODS: Fischer rats (230--260 g) underwent a thoracotomy, right main bronchus occlusion and instillation of 300 microg naked or complexed DNA (pCIluci, luciferase gene/CMV promoter) to the left lung followed by ventilation for 10 min. Rats were divided into five treatment groups (n=5): (1) Glucose, (2) Naked DNA, (3) Linear polyethylenimine (PEI), (4) Branched PEI, (5) Lipid GL-67/DOPE and (6) DOTAP/cholesterol. Animals were sacrificed 24 h after gene delivery for measurement of reporter gene activity and gas exchange of the left lung. RESULTS: Linear PEI was the most efficient gene delivery vector and was significantly better than DOTAP/cholesterol (P=0.00002) and naked DNA (P=0.004). All gene delivery vectors impaired function of the transfected left lung compared with DNA alone. Of all the gene delivery vectors tested, lipid GL-67/DOPE exerted the least effect on lung function whilst DOTAP/cholesterol mediated the most adverse effect. CONCLUSION: Linear PEI was the most efficient vector for gene delivery to rat lungs in our experimental setting although it mediated a moderate impairment in lung function. Further studies are needed to evaluate whether this effect is transient.