Dopaminergic modulation of phenotypical and functional characteristics of human T lymphocytes: perspectives for nonconventional immunomodulation.

Dopamine (DA) besides its action in the nervous system, plays an important role in immune cells interactions. Emerging role of DA as a regulator of CD4+ T cells physiology is important since dysregulation of different T cell subsets, showing abnormal cell numbers, functions, expression of dopamine receptors (DR) and/or response to DA, could contribute to the onset and development of some immune-related disorders. Thus, directly and indirectly acting dopaminergic therapeutics, currently used in approved clinical indications, could represent an attractive source of non-conventional agents for the modulation of CD4+ T cell functions. The aim of the present work was to develop in vitro methods to investigate the effects of dopaminergic agents, currently used in the pharmacotherapy, on the functional responses of CD4+ T cells, namely: (i) CD4+ T naïve (Tn), T central memory (TCM) and T effector memory (TEM) cells, and their responses to recall antigen (Ag); (ii) CD4+ T regulatory cells (Treg), and their suppressive effects on T effector cells (Teff) and (iii) CD4+ T naïve cells, and their ability to differentiate towards different T helper (Th) lineages (Th1/Th2/Th17). In cultured CD4+ T cells, our results have shown higher expression of DR in apoptotic cells in comparison to viable cells and stimulation-induced DR upregulation of all DR on viable cells. Addition of high concentrations of DA and L-DOPA (100 μM) have shown profound effect on survival of CD4+ T cells. Interestingly, based on preliminary experiments, our ex vivo data have shown trend of proliferating cells expressing DR in higher percentages that still need to be validated in subsequent studies on more subjects. So far, in vitro tested concentrations of dopaminergic agonists have not shown any major effects on proliferation of CD4+ T cells. In addition, through the use of flow cytometric analysis, expression of DR was examined on human: CD4+ naïve T lymphocytes (CD3+CD4+CD45RA+CCR7+), TCM (CD3+CD4+CD45RA-CCR7+), TEM (CD3+CD4+CD45RA-CCR7-), Treg cells (CD4+CD25highCD127low), and also frequency of different Th subsets: Th1 (CD4+CXCR3+CCR4-CCR6-), Th2 (CD4+CXCR3-CCR4+CCR6-), Th17 (CD4+CXCR3-CCR4-CCR6+) and Th1/Th17 (CD4+CXCR3+CCR4-CCR6+) were analysed. DR expression of all five DR was confirmed on each subset, present in a different extension potentially represents an opportunity to develop targeted immunomodulating strategies. Validated and developed in vitro method to test functional response of memory CD4+ T cells towards recall Ag have potential relevance for a wide range of different fields of T cell biology research in health and disease. Additionally, obtained preliminary results have confirmed in vitro experimental conditions likely appropriate to study commitment of naïve CD4+ T cells and factors mimicking specific polarisation routes (Th1/Th2/Th17), which are T subsets important in onset and development of some dopamine-related disorders. Further, in vitro methods have shown CD4+CD25high T cell-dependent inhibition of CD4+ T effector lymphocyte proliferation. Treg cells also suppressed production of IFN-γ and TNF-α from Teff cells. In addition, effects of DA and L-DOPA treatments seems to suppress Treg suppressive capacity in healthy subjects and in the group of Parkinson’s disease (PD) patients who had never been treated (PD-dn), but not in PD patients that were on dopaminergic therapy (PD-dt). Available evidence supports the possibility to repurpose dopaminergic agents as modulators of dopaminergic pathways, shifting the balance towards beneficial outcomes in some pathological conditions, such as PD. Over the last decades, an impressive number of studies in the animal model of immune diseases and in the clinical setting supported this evidence, and strongly required futher testing. The development of therapeutic protocols needs to take into account that DR exists in multiple subtypes and their patterns of expression, and that fuctional relevance differs among immune cells - and may even depend on the functional status (e.g. resting/activated) of specific cells. Proposed in vitro methods examined and characterised the various CD4+ T cell lineages, providing both the conceptual as well as the experimental framework for more in-depth investigation of dopaminergic pathways modulating CD4+ T cell function.