1α,25-Dihydroxyvitamin D₃ restores thymocyte apoptosis sensitivity in non-obese diabetic (NOD) mice through dendritic cells

Abstract

Aims/hypothesis

Resistance of NOD thymocytes to apoptosis-inducing signals is restored by 1α,25-dihydroxyvitamin D₃ (1α,25(OH)₂D₃), a therapy preventing diabetes in NOD mice. We studied whether modulation of thymocyte apoptosis is due to direct effects on thymic T lymphocytes or indirect effects via thymic dendritic cells, since both cell types constitute known targets for 1α,25(OH)₂D₃.

Methods and results

Female NOD mice were treated with 1α,25(OH)₂D₃ (5 μg/kg/2d) from 21 to 70 days. Vehicle-treated NOD and NOR mice served as controls. Analysis of thymic T lymphocytes from 1α,25(OH)₂D₃-treated mice revealed a decrease in number of apoptosis-resistant CD4⁺CD8⁺ and CD4⁺CD8⁻HSA^high T lymphocyte subsets, higher pro-apoptotic IL-2 and FasL, and lower anti-apoptotic Bclx-L mRNA expression levels. Thymic dendritic cells from 1α,25(OH)₂D₃-treated NOD mice had increased CD8α⁺FasL⁺ and CD80⁺/86⁺ expression compared to control NOD mice. In a syngeneic co-culture system of thymocytes and thymic dendritic cells, apoptosis levels were 20% higher only in co-cultures where both T cell- and dendritic cell-compartments originated from 1α,25(OH)₂D₃-treated mice. Activation-induced cell death-sensitivity in peripheral T lymphocytes was comparable to levels present in NOR mice, confirming better thymic selection in 1α,25(OH)₂D₃-treated mice.

Conclusion/interpretation

We conclude that 1α,25(OH)₂D₃ needs both thymic T cell- and dendritic cell-compartments to exert its apoptosis-restorative effects in NOD thymocytes.

Introduction

Type 1 diabetes is an organ-specific T lymphocyte-mediated autoimmune disease, characterised by destruction of insulin producing pancreatic β cells with subsequent development of hyperglycaemia and insulin dependence. Since exogenous insulin therapy is not able to completely avoid long-term complications and subsequent enormous health care costs, an etiologic more than a purely symptomatic treatment is urgently needed, aiming at prevention of diabetes. Prevention, however, supposes insight in the early pathogenic mechanisms, before disease becomes overt. Previous studies in pre-diabetic NOD mice, a mouse model for human type 1 diabetes, demonstrated several defects in the immune system, both in the T cell and antigen-presenting cell (APC) compartments [1]. Immunomodulation at young age, aiming at re-setting one or several immune dysregulations, thus seems a logical approach in the prevention of autoimmune diabetes. Life long treatment with high doses of 1α,25-dihydroxyvitamin D₃ (1α,25(OH)₂D₃), the activated form of vitamin D, prevents diabetes in NOD mice [2]. Further research on its mechanism of action in primary prevention revealed that 1α,25(OH)₂D₃ induced a restoration of T lymphocyte sensitivity to in vivo apoptosis-inducing signals (dexamethasone, cyclophosphamide), especially in the thymus [3], [4]. No general immune suppression was observed.

Abnormal T cell selection, which has been reported in NOD mice [5], [6], can be due to aberrations in the T lymphocytes themselves or in the thymic APC compartment, responsible for the presentation of (auto)antigen to thymic T lymphocytes. Dendritic cells (DCs) represent crucial APCs and several DC abnormalities have been described in NOD mice [7], [8], [9], [10], [11], [12]. Moreover, DCs constitute important target cells for the action of 1α,25(OH)₂D₃ [13]. The aim of this work was to study whether modulation of thymocyte apoptosis sensitivity by 1α,25(OH)₂D₃ is due to direct effects on T lymphocytes or indirect effects on thymic DCs. The data presented here demonstrate that direct effects of 1α,25(OH)₂D₃ on T lymphocytes combined with indirect effects on DCs are indispensable for final modulation of central T cell apoptosis sensitivity.