Emerging and Underrecognized Complications of Illicit Drug Use.

Illicit drug use can result in a wide range of medical complications. As the availability, synthesis, and popularity of illicit drugs evolve over time, new syndromes associated with their use may mimic infections. Some of these symptoms are anticipated drug effects, and others are complications of adulterants mixed with drugs or complications from the method of using drugs. Some illicit drugs are associated with rare infections, which are difficult to diagnosis with standard microbiological techniques. The goal of this review is to orient a wide range of clinicians-including general practitioners, emergency medicine providers, and infectious diseases specialists-to complications of illicit drug use that may be underrecognized. Improving awareness of infectious and noninfectious complications of illicit drug can expedite diagnosis and medical treatment of persons who use drugs and facilitate targeted harm reduction counseling to prevent future complications.

Fully differentiated HIV-1 specific CD8+ T effector cells are more frequently detectable in controlled than in progressive HIV-1 infection.

BACKGROUND: CD8+ T cells impact control of viral infections by direct elimination of infected cells and secretion of a number of soluble factors. In HIV-1 infection, persistent HIV-1 specific IFN-gamma+ CD8+ T cell responses are detected in the setting of disease progression, consistent with functional impairment in vivo. Recent data suggest that impaired maturation, as defined by the lineage markers CD45RA and CCR7, may contribute to a lack of immune control by these responses. METHODOLOGY/PRINCIPAL FINDINGS: We investigated the maturation phenotype of epitope-specific CD8+ T cell responses directed against HIV-1 in 42 chronically infected, untreated individuals, 22 of whom were "Controllers" (median 1140 RNA copies/ml plasma, range<50 to 2520), and 20 "progressors" of whom had advanced disease and high viral loads (median 135,500 RNA copies/ml plasma, range 12100 to >750000). Evaluation of a mean of 5 epitopes per person revealed that terminally differentiated CD8+ T cells directed against HIV-1 are more often seen in HIV-1 Controllers (16/22; 73%) compared to HIV-1 progressors (7/20; 35%)(p = 0.015), but the maturation state of epitope-specific responses within a given individual was quite variable. Maturation phenotype was independent of the HLA restriction or the specificity of a given CD8+ T cell response and individual epitopes associated with slow disease progression were not more likely to be terminally differentiated. CONCLUSIONS/SIGNIFICANCE: These data indicate that although full maturation of epitope-specific CD8+ T cell responses is associated with viral control, the maturation status of HIV-1 specific CD8+ T cell responses within a given individual are quite heterogeneous, suggesting epitope-specific influences on CD8+ T cell function.

Expansion of HIV-specific CD4+ and CD8+ T cells by dendritic cells transfected with mRNA encoding cytoplasm- or lysosome-targeted Nef.

Transfection with synthetic mRNA is a safe and efficient method of delivering antigens to dendritic cells for immunotherapy. Targeting antigens to the lysosome can sometimes enhance the CD4+ T-cell response. We transfected antigen-presenting cells (APCs) with mRNA encoding Gag-p24 and cytoplasmic, lysosomal, and secreted forms of Nef. Antigen-specific cytotoxic T cells were able to lyse the majority of transfected targets, indicating that transfection was efficient. Transfection of APCs with a Nef construct bearing lysosomal targeting signals produced rapid and prolonged antigen presentation to CD4+ and CD8+ T cells. Polyclonal CD4+ and CD8+ T-cell lines recognizing multiple distinct epitopes were expanded by coculture of transfected dendritic cells with peripheral blood mononuclear cells from viremic and aviremic HIV-infected subjects. Importantly, lysosome-targeted antigen drove a significantly greater expansion of Nef-specific CD4+ T cells than cytoplasmic antigen. The frequency of recognition of CD8 but not CD4 epitopes by mRNA-expanded T cells was inversely proportional to sequence entropy and was similar to ex vivo responses from a large chronic cohort. Thus human dendritic cells transfected with mRNA encoding lysosome-targeted HIV antigen can expand a broad, polyclonal repertoire of antiviral T cells, offering a promising approach to HIV immunotherapy.

Differences in the expressed HLA class I alleles effect the differential clustering of HIV type 1-specific T cell responses in infected Chinese and caucasians.

China is a region of the world with a rapidly spreading HIV-1 epidemic. Studies providing insights into HIV-1 pathogenesis in infected Chinese are urgently needed to support the design and testing of an effective HIV-1 vaccine for this population. HIV-1-specific T cell responses were characterized in 32 HIV-1-infected individuals of Chinese origin and compared to 34 infected caucasians using 410 overlapping peptides spanning the entire HIV-1 clade B consensus sequence in an IFN-gamma ELISpot assay. All HIV-1 proteins were targeted with similar frequency in both populations and all study subjects recognized at least one overlapping peptide. HIV-1-specific T cell responses clustered in seven different regions of the HIV-1 genome in the Chinese cohort and in nine different regions in the caucasian cohort. The dominant HLA class I alleles expressed in the two populations differed significantly, and differences in epitope clustering pattern were shown to be influenced by differences in class I alleles that restrict immunodominant epitopes. These studies demonstrate that the clustering of HIV-1-specific T cell responses is influenced by the genetic HLA class I background in the study populations. The design and testing of candidate vaccines to fight the rapidly growing HIV-1 epidemic must therefore take the HLA genetics of the population into account as specific regions of the virus can be expected to be differentially targeted in ethnically diverse populations.

Comparison of overlapping peptide sets for detection of antiviral CD8 and CD4 T cell responses.

Increasing efforts are directed towards the development of effective vaccines through induction of virus-specific T cell responses. Although emerging data indicate a significant role of these cells in determining viral set point in infections such as HIV, there is as yet no consensus as to the best methods for assaying the breadth of these responses. In this study, we used sensitive interferon gamma-based intracellular cytokine staining (ICS) and Elispot assays to determine the optimal overlapping peptide set to screen for these responses. Twenty persons with established HIV infection were studied, focusing on responses to the highly immunogenic Nef protein. Six different HIV-1 Nef peptide sets were used, ranging in length from 15 to 20 amino acids (aa), in overlap from 10 to 11 amino acids, and derived from two different B clade sequences. A total of 54 CD8 T cell responses to Nef peptides were found in this cohort, of which only 12 were detected using previously defined Nef optimal epitopes. No single peptide set detected all responses. Though there was a trend of the shorter peptides detecting more CD8 T cell responses than the 20 amino acid long peptides and longer peptides detecting more CD4 T cell responses, neither was statistically significant. There was no difference between an overlap of 10 or 11 amino acids. All responses detected with the six different sets of overlapping peptides were towards the more highly conserved regions of Nef. We conclude that peptides ranging from 15 to 20 amino acids yield similar results in IFN-gamma-based Elispot and ICS assays, and that all are likely to underestimate the true breadth of responses to a given reference strain of virus.

Comprehensive epitope analysis of human immunodeficiency virus type 1 (HIV-1)-specific T-cell responses directed against the entire expressed HIV-1 genome demonstrate broadly directed responses, but no correlation to viral load.

Cellular immune responses play a critical role in the control of human immunodeficiency virus type 1 (HIV-1); however, the breadth of these responses at the single-epitope level has not been comprehensively assessed. We therefore screened peripheral blood mononuclear cells (PBMC) from 57 individuals at different stages of HIV-1 infection for virus-specific T-cell responses using a matrix of 504 overlapping peptides spanning all expressed HIV-1 proteins in a gamma interferon-enzyme-linked immunospot (Elispot) assay. HIV-1-specific T-cell responses were detectable in all study subjects, with a median of 14 individual epitopic regions targeted per person (range, 2 to 42), and all 14 HIV-1 protein subunits were recognized. HIV-1 p24-Gag and Nef contained the highest epitope density and were also the most frequently recognized HIV-1 proteins. The total magnitude of the HIV-1-specific response ranged from 280 to 25,860 spot-forming cells (SFC)/10(6) PBMC (median, 4,245) among all study participants. However, the number of epitopic regions targeted, the protein subunits recognized, and the total magnitude of HIV-1-specific responses varied significantly among the tested individuals, with the strongest and broadest responses detectable in individuals with untreated chronic HIV-1 infection. Neither the breadth nor the magnitude of the total HIV-1-specific CD8+-T-cell responses correlated with plasma viral load. We conclude that a peptide matrix-based Elispot assay allows for rapid, sensitive, specific, and efficient assessment of cellular immune responses directed against the entire expressed HIV-1 genome. These data also suggest that the impact of T-cell responses on control of viral replication cannot be explained by the mere quantification of the magnitude and breadth of the CD8+-T-cell response, even if a comprehensive pan-genome screening approach is applied.

Antiretroviral drug resistance mutations in antiretroviral-naive prisoners.

We assessed the incidence of antiretroviral drug resistance in a cohort of 25 antiretroviral-naive, human immunodeficiency virus-positive inmates in Massachusetts. Silent mutations, unexpected mutations at resistant sites, and resistance mutations were recorded. Among these inmates, we found a prevalence of drug resistance mutations that was equivalent to the prevalence previously found in nonprison populations in the same state.