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The present study is designed to characterize the role of dopamine in neuroimmunomodulation of mice. The neurotoxin-induced neurodegeneration was used to investigate the mechanism in lymphoid tissue atrophy. In addition, effects of the dopamine and its agonists on lymphocyte proliferation were assessed. The presence of tyrosine hydroxylase in immune cells was identified. Correlation of tyrosine hydroxylase expression with cell growth was also elucidated. 6-Hydroxydopamine (6-OHDA) was reported to induce the degeneration of noradrenergic nerves and alter the immune responses. In this study, intraperitoneal administration of 6-OHDA induces thymus atrophy in mice. The lowest levels of thymus weight and cell number were reached between day 3 to 5 in mice receiving 6-OHDA treatment; they gradually recovered thereafter. On flow cytometry analysis, the most substantial reductions were recorded for CD4+CD8+ thymocytes, although the numbers of other subpopulations such as CD4+CD8-, CD4- CD8+, CD4-CD8- cells were also reduced. DNA fragmentation, a characteristic of apoptosis, was detected in the thymocytes following 6- OHDA injection. Pretreatment with desipramine, a catecholamine uptake inhibitor, greatly blocked the effects of 6-OHDA on reduction in thymus size and thymocyte number, the changes in thymocyte subpopulations, the percentage of apoptotic cells and the appearance of DNA fragmented bands. These results indicate that 6-OHDA induces mouse thymocytes to undergo apoptosis in vivo. The mechanism of 6-OHDA-induced thymocyte apoptosis were further investigated in vitro. Results showed that 6-OHDA-induced thymocyte apoptosis could be detected in vitro and blocked by desipramine treatment. Flow cytometry analysis indicated that CD4+CD8+ cells were most susceptible to 6-OHDA, and Bcl-2 expression was reduced after 6-OHDA treatment. Both VAD-FMK, a broad spectrum interleukin-1b-converting enzyme (ICE)-family protease inhibitor, and DEVD-CHO, a potent inhibitor of CPP32 (caspase-3), blocked 6-OHDA-induced thymocyte apoptosis. However, the ICE (caspase-1) inhibitor YVAD-CMK failed to cause such effect. This cell death process was prevented by the treatment of protein synthesis inhibitor, cycloheximide, and by the antioxidants such as N- acetylcysteine or butylated hydroxyanisole. These results suggest that down-regulation of Bcl-2, activation of ICE-like proteases such as CPP32 but not ICE, generation of reactive oxygen species, and synthesis of new proteins are involved in the apoptotic mechanism of 6-OHDA in thymocytes. Dopamine is known as the precursor of catecholamine and one of the neurotransmitters in brain and peripheral tissues. Recent studies implicated the important role of dopamine in immune responses. Intraperitoneal administration of 1-methyl-4-phenyl-1,2,3,6- tetrahydropyridine (MPTP) at dose sufficient to lower endogenous dopamine suppressed splenocyte proliferation in response to mitogens such as lipopolysaccharide (LPS) or concanavalin A (Con A). Moreover, intravenous injection of the specific agonists for dopamine DA-1 receptor (SKF38393) or DA-2 receptor (LY171555) into mice enhanced the splenocyte proliferation stimulated by LPS or Con A. In the in vitro cultures, dopamine, SKF38393 and LY171555 all promoted cell proliferation in the presence of LPS or Con A. These results indicate that dopamine has an ability to regulate B- and T-cell proliferation both in vivo and in vitro. The role of endogenous dopamine in immune cell growth was further investigated. Haloperidol, a general antagonist of dopamine receptor, reduced cell growth rate of T cell hybridoma (10I) and rat nervous pheochromocytoma cells (PC12). Tyrosine hydroxylase (TH) is the initial rate-limiting enzyme of catecholamine biosynthesis in nervous system. Flow cytometric analysis indicated TH also expresses in various immune cells including T-cell lineage (murine thymocytes, spleen T cells, T cell hybridoma 10I, human Jurkat-T and MOLT-4 cells), B-cell lineage (A20 cells), and monocyte/macrophage lineage (P388D1 and U937 cells). The presence of TH in PC12 cells was used as control. TH expression was down-regulated in low serum condition in which cell growth rate was reduced. Temporal studies indicated that the expression of TH increased during 10I cell growth. Both a-methyl-p-tyrosine and MPTP reduced TH expression and cell growth in a dose-dependent manner. These results suggest that immune T cells expresses TH which is correlated to cell growth, and that dopamine released from immune cells may bind to the receptors to act in an autocrine or paracrine way.
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