Effects of cigarette smoke extract on primary activated T cells.

Tobacco smoking predisposes the development of diseases characterized by chronic inflammation and T cell dysfunction. In this study, we aimed to determine the direct effects of cigarette smoke on primary T cells and to identify the corresponding molecular mediators. Activated T cells cultured in the presence of cigarette smoke extract (CSE) displayed a dose-dependent decrease in cell proliferation, which associated with the induction of cellular apoptosis. T cell apoptosis by CSE was independent of caspases and mediated through reactive oxygen and nitrogen species endogenously contained within CSE. Additional results showed that exposure of T cells to CSE induced phosphorylation of the stress mediator eukaryotic-translation-initiation-factor 2 alpha (eIF2α). Inhibition of the phosphorylation of eIF2α in T cells prevented the cellular apoptosis induced by CSE. Altogether, the results show the direct effects of CSE on T cells, which advance in the understanding of how cigarette smoking promotes chronic inflammation and immune dysfunction.

Role of c-Myb in the survival of pre B-cell acute lymphoblastic leukemia and leukemogenesis.

Acute lymphoblastic leukemia (ALL) is the most common cancer in children. The current treatment protocol for ALL involves an intense chemotherapy regimen yielding cure rates of nearly 80%. However, new therapies need to be designed not only to increase the survival rate but also to combat the risk of severe therapy associated toxicities including secondary malignancies, growth problems, organ damage, and infertility. The c-Myb proto-oncogene is highly expressed in immature hematopoietic cells. In this study, we demonstrate that loss of c-Myb itself decreased the viability of these leukemic cells. Additionally, the inhibition of c-Myb caused a decrease in cell proliferation, significantly increased the number of cells in G(0) /G(1) phase of the cell cycle, increased the sensitivity of pre-B-ALL cells to cytotoxic agents in vitro, and significantly delayed disease onset in a mouse model of leukemia. Furthermore, we demonstrate that Bcl-2 is a target of c-Myb in pre-B-ALL cells. Our results identify c-Myb as a potential therapeutic target in pre-B-ALL and suggest that suppression of c-Myb levels or activity, in combination with currently used therapies and/or dose reduction, may lead to a decrease in toxicity and an increase in patient survival rates. Because c-Myb is aberrantly expressed in several other malignancies, targeting c-Myb will have broad clinical applications.

L-arginine deprivation regulates cyclin D3 mRNA stability in human T cells by controlling HuR expression.

Myeloid-derived suppressor cells are a major mechanism of tumor-induced immune suppression in cancer. Arginase I-producing myeloid-derived suppressor cells deplete l-arginine (L-Arg) from the microenvironment, which arrests T cells in the G(0)-G(1) phase of the cell cycle. This cell cycle arrest correlated with an inability to increase cyclin D3 expression resulting from a decreased mRNA stability and an impaired translation. We sought to determine the mechanisms leading to a decreased cyclin D3 mRNA stability in activated T cells cultured in medium deprived of L-Arg. Results show that cyclin D3 mRNA instability induced by L-Arg deprivation is dependent on response elements found in its 3'-untranslated region (UTR). RNA-binding protein HuR was found to be increased in T cells cultured in medium with L-Arg and bound to the 3'-untranslated region of cyclin D3 mRNA in vitro and endogenously in activated T cells. Silencing of HuR expression significantly impaired cyclin D3 mRNA stability. L-Arg deprivation inhibited the expression of HuR through a global arrest in de novo protein synthesis, but it did not affect its mRNA expression. This alteration is dependent on the expression of the amino acid starvation sensor general control nonderepressible 2 kinase. These data contribute to an understanding of a central mechanism by which diseases characterized by increased arginase I production may cause T cell dysfunction.

Pegylated arginase I: a potential therapeutic approach in T-ALL.

Adult patients with acute lymphoblastic T cell leukemia (T-ALL) have a very poor prognosis and few effective therapeutic options. Therefore, novel therapies that increase the efficacy of the treatments and that prolong T-ALL patient survival are needed. Malignant T cells require high concentrations of nutrients to sustain their increased rate of proliferation. In this study, we determined whether L-Arginine depletion by the pegylated form of the L-Arginine-metabolizing enzyme arginase I (peg-Arg I) impairs the proliferation of malignant T cells. Our results show that peg-Arg I depleted L-Arginine levels in vitro and in vivo. In addition, treatment of malignant T-cell lines with peg-Arg I significantly impaired their proliferation, which correlated with a decreased progression into the cell cycle, followed by the induction of apoptosis. Furthermore, peg-Arg I impaired the expression of cyclin D3, a fundamental protein in T-ALL proliferation, through a global arrest in protein synthesis. Injection of peg-Arg I plus chemotherapy agent Cytarabine prolonged survival in mice bearing T-ALL tumors. This antitumoral effect correlated with an inhibition of T-ALL proliferation in vivo, a decreased expression of cyclin D3, and T-ALL apoptosis. The results suggest the potential benefit of L-Arginine depletion by peg-Arg I in the treatment of T-cell malignancies.

Arginase I in myeloid suppressor cells is induced by COX-2 in lung carcinoma.

Myeloid suppressor cells (MSCs) producing high levels of arginase I block T cell function by depleting l-arginine in cancer, chronic infections, and trauma patients. In cancer, MSCs infiltrating tumors and in circulation are an important mechanism for tumor evasion and impair the therapeutic potential of cancer immunotherapies. However, the mechanisms that induce arginase I in MSCs in cancer are unknown. Using the 3LL mouse lung carcinoma, we aimed to characterize these mechanisms. Arginase I expression was independent of T cell-produced cytokines. Instead, tumor-derived soluble factors resistant to proteases induced and maintained arginase I expression in MSCs. 3LL tumor cells constitutively express cyclooxygenase (COX)-1 and COX-2 and produce high levels of PGE2. Genetic and pharmacological inhibition of COX-2, but not COX-1, blocked arginase I induction in vitro and in vivo. Signaling through the PGE2 receptor E-prostanoid 4 expressed in MSCs induced arginase I. Furthermore, blocking arginase I expression using COX-2 inhibitors elicited a lymphocyte-mediated antitumor response. These results demonstrate a new pathway of prostaglandin-induced immune dysfunction and provide a novel mechanism that can help explain the cancer prevention effects of COX-2 inhibitors. Furthermore, an addition of arginase I represents a clinical approach to enhance the therapeutic potential of cancer immunotherapies.

Helicobacter pylori arginase inhibits T cell proliferation and reduces the expression of the TCR zeta-chain (CD3zeta).

Helicobacter pylori infects approximately half the human population. The outcomes of the infection range from gastritis to gastric cancer and appear to be associated with the immunity to H. pylori. Patients developing nonatrophic gastritis present a Th1 response without developing protective immunity, suggesting that this bacterium may have mechanisms to evade the immune response of the host. Several H. pylori proteins can impair macrophage and T cell function in vitro through mechanisms that are poorly understood. We tested the effect of H. pylori extracts and live H. pylori on Jurkat cells and freshly isolated human normal T lymphocytes to identify possible mechanisms by which the bacteria might impair T cell function. Jurkat cells or activated T lymphocytes cultured with an H. pylori sonicate had a reduced proliferation that was not caused by T cell apoptosis or impairment in the early T cell signaling events. Instead, both the H. pylori sonicate and live H. pylori induced a decreased expression of the CD3zeta-chain of the TCR. Coculture of live H. pylori with T cells demonstrated that the wild-type strain, but not the arginase mutant rocF(-), depleted L-arginine and caused a decrease in CD3zeta expression. Furthermore, arginase inhibitors reversed these events. These results suggest that H. pylori arginase is not only important for urea production, but may also impair T cell function during infection.

L-Arginine modulates CD3zeta expression and T cell function in activated human T lymphocytes.

Engagement of the T cell receptor (TCR) by antigen or anti-CD3 antibody results in a cycle of internalization and re-expression of the CD3zeta. Following internalization, CD3zeta is degraded and replaced by newly synthesized CD3zeta on the cell surface. Here, we provide evidence that availability of the amino acid L-arginine modulates the cycle of internalization and re-expression of CD3zeta and cause T cell dysfunction. T cells stimulated and cultured in presence of L-arginine, undergo the normal cycle of internalization and re-expression of CD3zeta. In contrast, T cells stimulated and cultured in absence of L-arginine, present a sustained down-regulation of CD3zeta preventing the normal expression of the TCR, exhibit a decreased proliferation, and a significantly diminished production of IFNgamma, IL5, and IL10, but not IL2. The replenishment of L-arginine recovers the expression of CD3zeta. The decreased expression of CD3zeta is not caused by a decreased CD3zeta mRNA, an increased CD3zeta degradation or T cell apoptosis.