Role of endothelial Nox2 NADPH oxidase in angiotensin II-induced hypertension and vasomotor dysfunction

NADPH oxidase (Nox)-derived reactive oxygen species (ROS) are known to be involved in angiotensin II-induced hypertension and endothelial dysfunction. Several Nox isoforms are expressed in the vessel wall, among which Nox2 is especially abundant in the endothelium. Endothelial Nox2 levels rise during hypertension but little is known about the cell-specific role of endothelial Nox2 in vivo. To address this question, we generated transgenic mice with endothelial-specific overexpression of Nox2 (Tg) and studied the effects on endothelial function and blood pressure. Tg had an about twofold increase in endothelial Nox2 levels which was accompanied by an increase in p22phox levels but no change in levels of other Nox isoforms or endothelial nitric oxide synthase (eNOS). Basal NADPH oxidase activity, endothelial function and blood pressure were unaltered in Tg compared to wild-type littermates. Angiotensin II caused a greater increase in ROS production in Tg compared to wild-type aorta and attenuated acetylcholine-induced vasorelaxation. Both low and high dose chronic angiotensin II infusion increased telemetric ambulatory blood pressure more in Tg compared to wild-type, but with different patterns of BP change and aortic remodeling depending upon the dose of angiotensin II dose. These results indicate that an increase in endothelial Nox2 levels contributes to angiotensin II-induced endothelial dysfunction, vascular remodeling and hypertension

Mechanism Underlying Defective Interferon Gamma-Induced IDO Expression in Non-obese Diabetic Mouse Fibroblasts

Indoleamine 2,3-dioxygenase (IDO) can locally suppress T cell-mediated immune responses. It has been shown that defective self-tolerance in early prediabetic female non-obese diabetic (NOD) mice can be attributed to the impaired interferon-gamma (IFN-γ)- induced IDO expression in dendritic cells of these animals. As IFN-γ can induce IDO in both dendritic cells and fibroblasts, we asked the question of whether there exists a similar defect in IFN-γ-induced IDO expression in NOD mice dermal fibroblasts. To this end, we examined the effect of IFN-γ on expression of IDO and its enzymatic activity in NOD dermal fibroblasts. The results showed that fibroblasts from either prediabetic (8 wks of age) female or male, and diabetic female or male (12 and 24 wks of age respectively) NOD mice failed to express IDO in response to IFN-γ treatment. To find underlying mechanisms, we scrutinized the IFN- γ signaling pathway and investigated expression of other IFN-γ-modulated factors including major histocompatibility complex class I (MHC-I) and type I collagen (COL-I). The findings revealed a defect of signal transducer and activator of transcription 1 (STAT1) phosphorylation in NOD cells relative to that of controls. Furthermore, we found an increase in MHC-I and suppression of COL-I expression in fibroblasts from both NOD and control mice following IFN-γ treatment; indicating that the impaired response to IFN-γ in NOD fibroblasts is specific to IDO gene. Finally, we showed that an IFN-γ-independent IDO expression pathway i.e. lipopolysaccharide (LPS)-mediated-c-Jun kinase is operative in NOD mice fibroblast. In conclusion, the findings of this study for the first time indicate that IFN-γ fails to induce IDO expression in NOD dermal fibroblasts; this may partially be due to defective STAT1 phosphorylation in IFN-γ-induced-IDO signaling pathway

Interleukin-13 Promotes Susceptibility to Chlamydial Infection of the Respiratory and Genital Tracts

Chlamydiae are intracellular bacteria that commonly cause infections of the respiratory and genital tracts, which are major clinical problems. Infections are also linked to the aetiology of diseases such as asthma, emphysema and heart disease. The clinical management of infection is problematic and antibiotic resistance is emerging. Increased understanding of immune processes that are involved in both clearance and immunopathology of chlamydial infection is critical for the development of improved treatment strategies. Here, we show that IL-13 was produced in the lungs of mice rapidly after Chlamydia muridarum (Cmu) infection and promoted susceptibility to infection. Wild-type (WT) mice had increased disease severity, bacterial load and associated inflammation compared to IL-13 deficient (−/−) mice as early as 3 days post infection (p.i.). Intratracheal instillation of IL-13 enhanced bacterial load in IL-13−/− mice. There were no differences in early IFN-g and IL-10 expression between WT and IL-13−/− mice and depletion of CD4+ T cells did not affect infection in IL-13−/− mice. Collectively, these data demonstrate a lack of CD4+ T cell involvement and a novel role for IL-13 in innate responses to infection. We also showed that IL-13 deficiency increased macrophage uptake of Cmu in vitro and in vivo. Moreover, the depletion of IL-13 during infection of lung epithelial cells in vitro decreased the percentage of infected cells and reduced bacterial growth. Our results suggest that enhanced IL-13 responses in the airways, such as that found in asthmatics, may promote susceptibility to chlamydial lung infection. Importantly the role of IL-13 in regulating infection was not limited to the lung as we showed that IL-13 also promoted susceptibility to Cmu genital tract infection. Collectively our findings demonstrate that innate IL-13 release promotes infection that results in enhanced inflammation and have broad implications for the treatment of chlamydial infections and IL-13-associated diseases

Regulatory T Cells in the Pathogenesis and Healing of Chronic Human Dermal Leishmaniasis Caused by Leishmania (Viannia) Species

The immune inflammatory response is a double edged sword. During infectious diseases, regulatory T cells can prevent eradication of the pathogen but can also limit inflammation and tissue damage. We investigated the role of regulatory T cells in chronic dermal leishmaniasis caused by species of the parasite Leishmania that are endemic in South and Central America. We found that although individuals with chronic lesions have increased regulatory T cells in their blood and at skin sites where immune responses to Leishmania were taking place compared to infected individuals who do not develop disease, their capacity to control the inflammatory response to Leishmania was inferior. However, healing of chronic lesions at the end of treatment was accompanied by an increase in the number and capacity of regulatory T cells to inhibit the function of effector T cells that mediate the inflammatory response. Different subsets of regulatory T cells, defined by the expression of molecular markers, were identified during chronic disease and healing, supporting the participation of distinct regulatory T cells in the development of disease and the control of inflammation during the healing response. Immunotherapeutic strategies may allow these regulatory T cell subsets to be mobilized or mitigated to achieve healing

GTPase-Rac enhances depolarization-induced superoxide production by the macula densa during tubuloglomerular feedback

Superoxide (O2− ) enhances tubuloglomerular feedback (TGF) by scavenging nitric oxide at the macula densa (MD). The primary source of O2− in the MD during TGF is NADPH oxidase, which is activated by membrane depolarization. While Rac, a small GTP-binding protein, has been shown to enhance NADPH oxidase activity, its role in O2− generation by the MD is unknown. We hypothesized that depolarization of the MD leads to translocation of Rac to the apical membrane, and its activation, in turn, augments O2− generation during TGF. We tested this by measuring membrane potential and increased O2− levels during TGF responses in isolated, perfused tubules containing the intact MD plaque. Switching tubular NaCl from 10 to 80 mM, which induces TGF, depolarized membrane potential by 28.4 ± 4.5% from control (P < 0.05) and O2− levels from 124 ± 19 to 361 ± 27 U/min. This NaCl-induced depolarization and O2− generation were blocked by a Cl− channel blocker, 5-nitro-2(3-phenylpropylamino) benzoic acid (NPPB; 10−6 M). Inhibition of Rac blunted NaCl-induced O2− generation by 47%. When the NaCl content of the MD perfusate was increased from 10 to 80 mM, immunointensity of Rac on the apical side increased from 32 ± 3.1 to 46 ± 2.5% of the total immunofluorescence in the MD, indicating that high NaCl induces the translocation of Rac to the apical membrane. This NaCl-induced Rac translocation was blocked by a Cl− channel blocker, NPPB, indicating that depolarization of the MD induced Rac translocation. In conclusion, we found that depolarization of the MD during TGF leads to translocation of Rac to the apical membrane, which enhances O2− generation by the MD

HeLa cells cocultured with peripheral blood lymphocytes acquire an immuno-inhibitory phenotype through up-regulation of indoleamine 2,3-dioxygenase activity

The mechanisms by which tumour cells escape recognition by the immune system or subvert antitumour effector responses remain poorly understood. In the course of investigating the potential of costimulatory signals in anticancer immunotherapy strategies, we have observed that HeLa cells (a human cervical carcinoma cell line) cocultured with peripheral blood lymphocytes (PBL) acquire the capacity to inhibit PBL proliferation in response to interleukin-2 (IL-2). This immuno-inhibitory phenotype was further shown to result from induction of the tryptophan-catabolizing enzyme, indoleamine 2,3-dioxygenase (IDO), by interferon-γ (IFN-γ) secreted from cocultured allo-reactive PBL. This enzyme has recently been shown to be a critically important modulator of immunological responses, most notably through the capacity to protect allogeneic concepti from alloreactive maternal lymphocytes. While the cytostatic consequences of IDO activity in tumour cells has received attention, the data presented in this report support the hypothesis that IDO activity may also act to impair antitumour immune responses

Transcriptional regulation of indoleamine 2,3-dioxygenase (IDO) by tryptophan and its analogue: Down-regulation of the indoleamine 2,3-dioxygenase (IDO) transcription by tryptophan and its analogue

Indoleamine 2,3-dioxygenase (IDO; EC 1.13.11.42) is a rate-limiting enzyme involved in the catabolism of tryptophan, which is an essential amino acid. It is induced under pathological conditions, such as the presence of viral infections or tumour cells. This enzyme is induced by IFN-γ in the mouse rectal carcinoma cell line CMT-93. It is known that both 1-methyl-l-tryptophan (1-MT) and methylthiohydantoin-dl-tryptophan (MTH-trp) are tryptophan analogues, and are authentic inhibitors of the enzymatic activity of IDO. In this study, we examined the effects of both 1-MT and MTH-trp on the IFN-γ inducible IDO expression of CMT-93. As a result, the IFN-γ inducible IDO mRNA and the protein levels in CMT-93 were suppressed by 1-MT and MTH-trp, independently. Moreover, tryptophan (Trp), as a substrate of IDO, also suppressed IDO induction by IFN-γ at the transcriptional level. These results suggest that 1-MT and MTH-trp are as inhibitors of IDO enzymatic activity, and Trp suppresses IDO induction by IFN-γ at the transcriptional level