Tim-3 expression defines a novel population of dysfunctional T cells with highly elevated frequencies in progressive HIV-1 infection

Progressive loss of T cell functionality is a hallmark of chronic infection with human immunodeficiency virus 1 (HIV-1). We have identified a novel population of dysfunctional T cells marked by surface expression of the glycoprotein Tim-3. The frequency of this population was increased in HIV-1–infected individuals to a mean of 49.4 ± SD 12.9% of CD8+ T cells expressing Tim-3 in HIV-1–infected chronic progressors versus 28.5 ± 6.8% in HIV-1–uninfected individuals. Levels of Tim-3 expression on T cells from HIV-1–infected inviduals correlated positively with HIV-1 viral load and CD38 expression and inversely with CD4+ T cell count. In progressive HIV-1 infection, Tim-3 expression was up-regulated on HIV-1–specific CD8+ T cells. Tim-3–expressing T cells failed to produce cytokine or proliferate in response to antigen and exhibited impaired Stat5, Erk1/2, and p38 signaling. Blocking the Tim-3 signaling pathway restored proliferation and enhanced cytokine production in HIV-1–specific T cells. Thus, Tim-3 represents a novel target for the therapeutic reversal of HIV-1–associated T cell dysfunction

HIV-Specific T-Cells Accumulate in the Liver in HCV/HIV Co-Infection

BACKGROUND AND AIMS: Hepatitis C Virus (HCV)-related liver disease progresses more rapidly in individuals co-infected with Human Immunodeficiency Virus-1 (HIV), although the underlying immunologic mechanisms are unknown. We examined whether HIV-specific T-cells are identified in the liver of HCV/HIV co-infected individuals and promote liver inflammation through bystander immune responses. METHODS: Ex-vivo intra-hepatic lymphocytes from HCV mono-infected and HCV/HIV co-infected individuals were assessed for immune responses to HIV and HCV antigens by polychromatic flow cytometry. RESULTS: HCV/HIV liver biopsies had similar frequencies of lymphocytes but lower percentages of CD4+ T-cells compared to HCV biopsies. In co-infection, intra-hepatic HIV-specific CD8+ and CD4+ T-cells producing IFN-gamma and TNF-alpha were detected and were comparable in frequency to those that were HCV-specific. In co-infected individuals, viral-specific CD8+ T-cells produced more of the fibrogenic cytokine, TNF-alpha. In both mono- and co-infected individuals, intra-hepatic HCV-specific T-cells were poorly functional compared to HIV-specific T-cells. In co-infection, HAART was not associated with a reconstitution of intra-hepatic CD4+ T-cells and was associated with reduction in both HIV and HCV-specific intra-hepatic cytokine responses. CONCLUSION: The accumulation of functional HIV-specific T-cells in the liver during HCV/HIV co-infection may represent a bystander role for HIV in inducing faster progression of liver disease

Evaluating T-cell Immunity in HCV/HIV Co-infection

Due to shared routes of transmission, co-infection with Hepatitis C (HCV) and Human Immuno-deficiency Virus (HIV) has become prevalent worldwide, with approximately one-third of all HIV-infected individuals and about 10% of all HCV infected individuals in North America being co-infected with the other virus (1). Recent advances in HIV treatment have increased the life expectancy of HIV-infected patients, resulting in HCV-associated disease to develop into a major cause of morbidity and mortality among the co-infected population. HIV is consistently shown to have a negative effect on different aspects of HCV disease, from increased HCV RNA levels (2) to aggravated liver fibrosis and more rapid progression to cirrhosis and end-stage liver disease (3). The host immune responses play a major role in not only controlling HCV infection, but also in causing hepatic inflammation and damage. Despite major advances in the understanding of the pathogenesis of these two infections, the mechanisms underlying the role of immune responses in more rapid progression of HCV disease in HCV/HIV co-infection is not quite clear. This thesis is generated based on an investigation to understand why HIV infection worsens HCV pathogenesis. This question is addressed throughout this thesis from different immunological aspects, with a focus on the function of T-cells. I have demonstrated that in HCV/HIV co-infection, functional HIV-specific T-cells accumulate in the liver and produce an array of cytokines, including INF- and TNF-, which may represent a bystander role for HIV in the aggravation of HCV-induced liver damage. My data also demonstrate that co-expression of two defined T-cell exhaustion markers, Tim-3 and PD-1 may play a significant role in HCV-specific T-cell dysfunction, in the setting of HIV co-infection. Both total and HCV-specific peripheral T-cells co-express Tim-3 and PD-1 in significantly higher frequencies, compared to HCV mono-infection. In co-infection, HCV-specific CD8+ T-cells showed greater frequencies of Tim-3/PD-1 dual-expression than those being HIV-specific, indicating a greater degree of exhaustion in the former. Additionally, I demonstrated that some HIV mono-infected individuals may contain CD8+ T-cells that cross-recognize two defined HLA-A2-restricted epitopes within the HIV and HCV proteome, the HIV-Gag: SLYNTVATL and HCV-NS5b: ALYDVVSKL. This T-cell cross-reactivity was further elaborated in the context of HCV/HIV co-infection, demonstrating that degeneracy of HIV-specific T-cells may play a role in the immuno-pathology of co-infection. Altogether, these data could be integrated into the foundation of potential mechanisms involved in the immunopathogenesis of HCV/HIV co-infection, and be applied to further investigation in basic science and clinical studies in this field.Ph

The Effect of Leverage and Liquidity on the Management of Earnings and Capital of Iranian Commercial Banks

Financial theorists have always focused on the ability of companies to manage earnings through the use of accounting methods for accruals. Evidence from some empirical research shows that earnings management is possible through the use of financial leverage and capital management and generally managing capital structure. The earnings management by banks is also implemented through monitoring loan-loss provision and net charge-off loans. This study investigates the relationship between "financial leverage and liquidity" and "earnings and capital management" of commercial banks listed in Tehran Stock Exchange (TSE) during the years 1385 to 1393. Data have been extracted from financial statements and their accompanying notes of some selected banks and Excel has been used to classify the data. Data analysis is also done using EViews software. The results from hypothesis testing show that there is a significant relationship between the liquidity ratios and financial leverage, earnings management and capital management in commercial banks

Tim-3 Negatively Regulates Cytotoxicity in Exhausted CD8⁺ T Cells in HIV Infection

<div><p>Cytotoxic CD8⁺ T cells (CTLs) contain virus infections through the release of granules containing both perforin and granzymes. T cell ‘exhaustion’ is a hallmark of chronic persistent viral infections including HIV. The inhibitory regulatory molecule, T cell Immunoglobulin and Mucin domain containing 3 (Tim-3) is induced on HIV-specific T cells in chronic progressive infection. These Tim-3 expressing T cells are dysfunctional in terms of their capacities to proliferate or to produce cytokines. In this study, we evaluated the effect of Tim-3 expression on the cytotoxic capabilities of CD8⁺ T cells in the context of HIV infection. We investigated the cytotoxic capacity of Tim-3 expressing T cells by examining 1) the ability of Tim-3⁺ CD8⁺ T cells to make perforin and 2) the direct ability of Tim-3⁺ CD8⁺ T cells to kill autologous HIV infected CD4⁺ target cells. Surprisingly, Tim-3⁺ CD8⁺ T cells maintain higher levels of perforin, which was mainly in a granule-associated (stored) conformation, as well as express high levels of T-bet. However, these cells were also defective in their ability to degranulate. Blocking the Tim-3 signalling pathway enhanced the cytotoxic capabilities of HIV specific CD8⁺ T cells from chronic progressors by increasing; a) their degranulation capacity, b) their ability to release perforin, c) their ability to target activated granzyme B to HIV antigen expressing CD4⁺ T cells and d) their ability to suppress HIV infection of CD4⁺ T cells. In this latter effect, blocking the Tim-3 pathway enhances the cytotoxcity of CD8⁺ T cells from chronic progressors to the level very close to that of T cells from viral controllers. Thus, the Tim-3 receptor, in addition to acting as a terminator for cytokine producing and proliferative functions of CTLs, can also down-regulate the CD8⁺ T cell cytotoxic function through inhibition of degranulation and perforin and granzyme secretion.</p> </div

Polychromatic FACS analysis of viral-specific T cells in HCV/HIV co-infection.

<p>Shown are representative FACS data of the HIV and HCV specific multi-parameter CD8⁺ T-cell responses from (a) liver and (b) blood of subject OM 405, a therapy-naïve HCV/HIV co-infected individual, after in vitro stimulation using pool of HIV and HCV peptides. Initial gating on forward scatter area (FSC-A) versus height (FSC-H) was used to remove doublets. The events were further gated on forward scatter (FSC) versus the dead cell marker to remove dead cells. Lymphocytes were gated on the remaining live cells on a FSC versus SSC plot. Gates on CD3⁺/CD8⁺ cells were then generated. All responses are gated on a CD3⁺/CD8⁺ population and presented against TNF-α on the x-axis. Figure (c) shows a comparison of the frequency of HIV and HCV-specific CD8⁺ T-cells in the liver and blood of therapy-naïve, co-infected individuals. All intra-hepatic HCV-specific responses are significantly stronger than peripheral HCV-specific responses.</p

Characterization of the functional hierarchy of viral specific CD8⁺ T-cells in HCV/HIV co-infection.

<p>Representative functional profiles of virus specific CD8⁺ T-cells in blood and liver are depicted from an individual with (a) HCV/HIV co-infection (OM 405) and from an individual with (b) HCV mono-infection (OM 428). For multi-parameter analysis, the Boolean gating platform was used to create all of the possible combinations of functions, generating 32 response patterns for 5 of the different functions analyzed. All data are reported after background correction. Nonspecific background is shown to become extremely low when examining combinations of functions, nearly reaching 0 events for multiple functions simultaneously. This permits a very low threshold for detection of positive responses from multiple combinations. Consequently, for multi-parameter analysis, the results were thresholded based on a minimum criterion of positivity, as calculated by SPICE software and presented as the 90^th percentile of negative values for each analysis. Each pie chart generated by SPICE software, represents the hierarchy of responses to either HCV or HIV antigen stimulation. For simplicity, responses are grouped by number of functions and matched to the colored bars, with black bars representing the percentage of responding cells to HIV peptides and gray bars representing the percentage of responses to HCV peptide stimulation. In all pie charts, color red represents the 5+ responding population and the colors blue, green, turquoise, and yellow representing the 4+, 3+, 2+, and 1+ populations respectively. Color-coded arcs represent the dominant marker within each pie slice, with color blue representing TNF-α, red for CD107-a, green for IFN-γ and black for MIP-1β. Although IL-2 is included in the presentation and demonstrated by bar graphs, the software would not allow for arc colors for more than 4 responses. As a result there is no arc representative for IL-2. Figure (c) represents the average frequency of intra-hepatic viral specific responses within the pool of CD8⁺ T-cell populations simultaneously expressing 2 functions or less, compared with those within the pool of populations expressing more than 2 functions simultaneously; as analyzed in 3 subjects within each cohort of HCV mono and HCV/HIV co-infected individuals. The cutoff point of simultaneous expression of more than 2 measured markers is considered to show CD8⁺ T-cell poly-functionality. Figure (d) represents PD-1 levels on HIV-1 and HCV-specific T-cells from HCV/HIV Co-infected liver (OM 385). Liver cells were stained with two pools of HLA-A*0201-restricted pentamers (Proimmune): HCV pentamers: NS3-CINGVCWTV, NS3-KLVALGINAV and HIV pentamers: Pol-ILKEPVHGV, Gag p24-TLNAWVKVV. PD-1 gating was based on FMO (fluorescence minus one) of the control sample.</p

Characteristics of intra-hepatic lymphocytes from HCV and HCV/HIV co-infected individuals.

<p>Frequencies of lymphocytes obtained from liver biopsies normalized to total live mononuclear cell count from FACS analysis are illustrated in (a) with percent composition of intra-hepatic CD4 and CD8 expressing T-cells shown in (b).</p

Characteristics of HCV mono-infected and HCV/HIV co-infected individuals, untreated for HCV.

<p>1. Normalized to 100,000 events count.</p><p>na = not applicabale.</p><p>no = number of participants.</p><p>HAART individuals were treated>1 year.</p