Ossama Allam

Immunosuppressive tryptophan catabolism and gut mucosal dysfunction following early HIV infection

BACKGROUND Tryptophan (Trp) catabolism into kynurenine (Kyn) contributes to immune dysfunction in chronic human immunodeficiency virus (HIV) infection. To better define the relationship between Trp catabolism, inflammation, gut mucosal dysfunction, and the role of early antiretroviral therapy (ART), we prospectively assessed patients early after they acquired HIV. METHODS Forty patients in the early phase of infection were longitudinally followed for 12 months after receiving a diagnosis of HIV infection; 24 were untreated, and 16 were receiving ART. Kyn/Trp ratio, regulatory T-cells (Tregs) frequency, T-cell activation, dendritic cell counts, and plasma levels of gut mucosal dysfunction markers intestinal-type fatty acid-binding protein, soluble suppression of tumorigenicity 2, and lipopolysaccharide were assessed. RESULTS Compared with healthy subjects, patients in the early phase of infection presented with elevated Kyn/Trp ratios, which further increased in untreated patients but normalized in ART recipients. Accordingly, in untreated subjects, the elevated Treg frequency observed at baseline continued to increase over time. The highest CD8(+) T-cell activation was observed during the early phase of infection and decreased in untreated patients, whereas activation normalized in ART recipients. The Kyn/Trp ratio was positively associated with CD8(+) T-cell activation and levels of inflammatory cytokines (interleukin 6, interferon γ-inducible protein 10, interleukin 18, and tumor necrosis factor α) and negatively associated with dendritic cell frequencies at baseline and in untreated patients. However, ART did not normalize plasma levels of gut mucosal dysfunction markers. CONCLUSIONS Early initiation of ART normalized enhanced Trp catabolism and immune activation but did not improve plasma levels of gut mucosal dysfunction markers.

Imbalanced production of IL-18 and its antagonist in human diseases, and its implications for HIV-1 infection.

IL-18 is a pleiotropic and multifunctional cytokine that belongs to the IL-1 family. It is produced as a biologically inactive precursor, which is cleaved into its active mature form mainly by caspase-1. The caspase becomes active from its inactive precursor (procaspase-1) upon assembly of an inflammasome. Because of IL-18s potential pro-inflammatory and tissue destructive effects, its biological activities are tightly controlled in the body by its naturally occurring antagonist called IL-18BP. The antagonist is produced in the body both constitutively and in response to an increased production of IL-18 as a negative feedback mechanism. Under physiological conditions, most of IL-18 in the circulation is bound with IL-18BP and is inactive. However, an imbalance in the production of IL-18 and its antagonist (an increase in the production of IL-18 with a decrease, no increase or an insufficient increase in the production of IL-18BP) has been described in many chronic inflammatory diseases in humans. The imbalance results in an increase in the concentrations of free IL-18 (unbound with its antagonist) resulting in increased biological activities of the cytokine that contribute towards pathogenesis of the disease. In this article, we provide an overview of the current biology of IL-18 and its antagonist, discuss how the imbalance occurs in HIV infections and how it contributes towards development of AIDS and other non-AIDS-associated clinical conditions occurring in HIV-infected individuals undergoing combination anti-retroviral therapy (cART). Finally, we discuss challenges facing immunotherapeutic strategies aimed at restoring balance between IL-18 and its antagonist in these patients.

Hammering out HIV-1 incidence with Hamming distance.

In epidemiology, the rate at which new infections occur with HIV-1 (or any other pathogen) in a population over a certain period of time is called incidence. Determining incidence is a very important measure for monitoring epidemics, future projections of the infected individuals, and evaluating success of vaccinations and preventive measures. It is also important to calculate appropriate sample size in any prospective vaccination trial. For measuring incidence, it is important to identify incident (recent) infections and distinguish them from pre-existing (chronic) ones. This distinction is of particular importance in the case of HIV-1, as the existing infections with the virus cannot and do not prevent subsequent infections with the same or different viral strains [1]. In line with the importance of detecting incident HIV-1 infections, several clinical tests have been developed [2]. The tests are based upon sequential expression of several HIV markers including viral RNA, viral antigens and various characteristics (e.g. antigen specificity, isotype, avidity, etc.) of virus-specific antibodies at precise predetermined times after the infection. The tests, however, are very cumbersome, time-consuming, intrusive, resourceintensive and difficult to perform on a large number of samples. Furthermore, they lack standardization, reproducibility and broader applicability to all infecting viral clades. Therefore, developing novel HIV-1 incidence assays remains a public health priority. In the original research paper presented in this issue, Park et al. [3] have devised a novel sequence-based assay to estimate HIV-1 incidence with high sensitivity and specificity. In order to

Mesenchymal stem cell therapy in HIV-infected HAART-treated nonimmune responders restores immune competence.

In this issue of AIDS, Zhang et al. [1] report about safety and immunological responses of therapy with mesenchymal stem cells (MSCs), derived from human umbilical cord or more specifically Wharton’s jelly, in HIV-infected nonimmune responder (NIR) individuals. The NIRs respond to highly active antiretroviral therapy (HAART) and effectively suppress HIV replication, but do not show any improvement in their immune status, as measured by increase in CD4þ T-cell counts. The authors conducted a prospective and open-label controlled pilot clinical trial. They made 3-monthly intravenous MSC transfusions in seven NIRs, whereas six NIRs received saline as controls. All the participants remained on HAART during the study and a 12-month follow-up period. The MSC recipients tolerated the therapy well and showed neither any adverse clinical effect nor increase in viral burden. More importantly, the MSC recipients showed a significant increase in their naive and central memory CD4þ T-cell counts compared with the control group. The therapy also restored their ability to produce interleukin (IL)-2 and IFN-g in response to HIV antigens, and down-regulated signs of immune activation, immune exhaustion, and inflammation. This study has provided a new glimpse of hope for the HIV-infected NIRs. It is noteworthy that more than 20% HAARTtreated HIV-infected persons exhibit the NIR phenotype, and are at increased risk for opportunistic infections, cancer and reduced life expectancy [2].

Differential synthesis and release of IL-18 and IL-18 Binding Protein from human platelets and their implications for HIV infection.

HighlightsPlatelets contain IL‐18 transcripts but no pre‐formed protein.They synthesize and release it de novo upon activation.They contain preformed IL‐18BP but not its transcripts.Fresh human platelet poor plasma (PRP) contains little IL‐18 in healthy individuals.Fresh PRP from HIV‐infected individual contain readily detectable IL‐18. Abstract IL‐18 is a pro‐inflammatory cytokine belonging to the IL‐1 family and is produced in the body from macrophages, epithelial and dendritic cells, keratinocytes, adrenal cortex etc. The cytokine is produced as an inactive precursor that is cleaved inside cells into its mature form by activated caspase 1, which exists as an inactive precursor in human cells and requires assembly of an inflammasomes for its activation. We show here for the first time that human platelets contain transcripts for the IL‐18 gene. They synthesize the cytokine de novo, process and release it upon activation. The activation also results in the assembly of an inflammasome and activation of caspase‐1. Platelets also contain the IL‐18 antagonist, the IL‐18‐Binding Protein (IL‐18BP); however, it is not synthesized in them de novo, is present in pre‐made form and is released irrespective of platelet activation. IL‐18 and IL‐18BP co‐localize to &agr; granules inside platelets and are secreted out with different kinetics. Platelet activation contributes to plasma concentrations in healthy individuals, as their plasma samples contain abundant IL‐18, while their platelet‐poor plasma samples contain very little amounts of the cytokine. The plasma and PPP samples from these donors, however, contain comparable amounts of IL‐18BP. Unlike healthy individuals, the platelet‐poor plasma from HIV‐infected individuals contains significant amounts of IL‐18. Our findings have important implications for viral infections and other human diseases that are accompanied by platelet activation.

A potentially protective role of IL-18 Binding Protein in HIV-infected Long-Term Non-Progressors

HighlightsIL‐18BP, an antagonist of IL‐18, production decreases in HIV infected individuals.No increase in production of IFN‐&ggr; in HIV infection may underlie this decrease.Its production is maintained at normal physiological levels in LTNP.The antagonist levels correlate negatively with viral loads.The levels tend to correlate positively with CD4+ T cell counts. Abstract An imbalance between IL‐18 and its antagonist, IL‐18 Binding Protein, occurs in the circulation of HIV‐infected individuals. We show here for the first time that HIV‐infected Long Term Non‐Progressors (LTNPs) do not develop this imbalance, and maintain normal levels of IL‐18BP in the circulation. Their circulating levels of the antagonist correlate negatively with viral loads and show a positive trend with CD4+ T cells counts. The maintenance of normal production of IL‐18BP may contribute, at least in part, to the ability of LTNPs to delay AIDS progression.

Microbial translocation and AIDS-associated lymphomas: crux of the problem.

In this issue of AIDS, Marks et al. [1] have reported significant associations between markers of microbial translocation and increased risk for non-Hodgkin’s lymphoma (NHL) in HIV-infected individuals. The authors examined prediagnostic blood samples from NHL and non-NHL HIV-infected individuals for different markers of microbial translocation. The results show that elevated plasma levels of sCD14 associated independently with 2.5-fold increased risk for NHL. The authors used Limulus Amebocyte Lysate Assay for measuring lipopolysaccharide (LPS). As plasma/serum contains substances that interfere with the assay, the authors measured percentage recovery of LPS from each sample after spiking it with LPS. Considering the samples from which they recovered the recommended (50–200) percentage of the spiked LPS, they found significant associations of the above-median values with increased (3.0-fold) risk for NHL. These results suggest, for the first time, an etiologic role of microbial translocation across gut mucosa in lymphomagenesis in HIV-infected individuals.

HIV induces production of IL-18 from intestinal epithelial cells that increases intestinal permeability and microbial translocation

Interleukin-18 (IL-18) is a pleiotropic cytokine of the IL-1 family with multiple context dependent functions. We and others have shown that HIV infection is accompanied by increased circulating levels of IL-18 along with decreased levels of its antagonist, Interleukin-18 Binding Protein (IL-18BP). The infection is also accompanied by intestinal inflammation and decreased intestinal integrity as measured by intestinal permeability, regeneration and repair. However, little is known concerning the relation between high level of IL-18 associated with the viral infection and intestinal permeability. Here we demonstrate that HIV treatment increases production of IL-18 and decreases that of IL-18BP production in human intestinal epithelial cell (IEC) lines. IL-18 causes apoptosis of the IEC by activating caspase-1 and caspase-3. It induces epithelial barrier hyperpermeability by decreasing and disrupting both tight and adherens junction proteins, occludin, claudin 2 and beta-catenin. Disorganization of F-actin was also observed in the IEC that were exposed to the cytokine. Moreover IL-18 decreases transepithelial electrical resistance (TEER) in Caco-2 and increases permeability in HT29 monolayers. The cells’ treatment with IL-18 causes an increase in the expression of phosphorylated myosin II regulatory light-chain (p-MLC) and myosin light-chain kinase (MLCK), and a decrease in phosphorylated Signal Transducer and Activator of Transcription (p-STAT)-5. This increase in p-MLC is suppressed by a Rho-kinase (ROCK)-specific inhibitor. Interestingly, the levels of the cytokine correlate with those of LPS in the circulation in three different categories of HIV infected patients (HAART-naïve and HAART-treated HIV-infected individuals, and Elite controls) as well as in healthy controls. Collectively, these results suggest that the HIV-induced IL-18 plays a role in increased intestinal permeability and microbial translocation observed in HIV-infected individuals.

Abstract 115: KIR haplotype polymorphism determines innate susceptibility/resistance to childhood leukemia

Background: The Killer-cell Immunoglobulin-like Receptor (KIR) genes encode receptors that are expressed on the surface of Natural Killer (NK) cells and a subset of T cells. The receptors regulate functional activities of these immune cells in the body. There exist seventeen distinct KIR genes in humans of which nine encode inhibitory, and six encode activating receptors, while two are pseudo genes. KIR haplotypes show extensive variability with respect to their repertoire of activating KIR genes. Humans may have none or up to six different activating KIR genes. The acquisition of these genes is believed to decrease activation threshold of NK cells in humans, and provide resistance to malignancy. Hypothesis: We hypothesized that the acquisition of activating KIR genes by an individual may protect him/her from childhood leukemia. Methods: We used the candidate gene approach. In a case-control study, we determined the presence or absence of different activating KIR genes using gene-specific primer pairs. The frequencies of each gene were compared between cases and controls using Chi2 test and multivariate logistic regression. Results: Our results show for the first time that the inheritance of activating KIR genes is associated with increased resistance to childhood leukemia (both B-ALL and T-ALL; in adjusted analysis p-values being as low as 1.14x10-7). Interestingly, this resistance increased as the number of activating KIR genes increased in the genome of the individual. Furthermore, we found that that this resistance/susceptibility is differentially modulated by co-inheritance of different inhibitory KIR-MHC class I gene pairs. Conclusions: The inheritance of activating KIR gene-containing haplotypes confers innate resistance to childhood leukemia. These receptors may serve as novel target molecules for therapeutic interventions in childhood leukemia. The study was supported by a research grant (≠2010-700554) from the Canadian Cancer Society Research Institute (CCSRI). Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 115. doi:1538-7445.AM2012-115