[Generation of T regulatory cells in children with type 1 diabetes mellitus]. / Generacja komórek T regulatorowych u dzieci z cukrzyca typu 1.

INTRODUCTION: In spite of intensive research the pathogenesis of type 1 diabetes mellitus is not thoroughly understood. One of the ideas which currently has a great number of supporters is the theory of the participation of T regulatory cells in the mechanism of insufficient suppression of the immune response against pancreatic self -antigens. According to some authors, the infusion of T regulatory cells in autoimmune diseases could lead to long -term remission or even a complete cure. AIM OF THE STUDY: The aim of our present study was to achieve T regulatory cells from conventional T lymphocytes isolated from a small amount of peripheral blood in children with type 1 diabetes mellitus and their comparison to the Tregs generated from the blood of control children. Additionally, we assessed the changes in the expression of selected genes essential for the function of these cells during Tregs generation. MATERIAL AND METHODS: The examined group consisted of 20 children with type 1 diabetes mellitus, the control group consisted of 20 non -diabetic children. From the peripheral blood CD4+CD25 - cells were separated and cultured with T -reg expander and interleukin (IL) 2. Before and after the culture the cells were analysed according to the expression of transcription factor FoxP3 and other molecules/cytokines: OX40, 4 -1BB, GITR, ICOS -1, CTLA -4 and IFN -γ, IL -10 and TGF -ß. RESULTS: We observed a significantly higher percentage of T regulatory cells after the culture (with no difference between diabetic and control children). Moreover, we observed a lower expression of mRNA for GITR molecule and a higher IL -10 expression in the cultures of diabetic children compared to the control ones. The cells cultured from the blood of control children were characterised by a higher increase in the expression of mRNA for ICOS -1 and a lower expression of mRNA for TGF -ß in comparison to the cultures from diabetic children. CONCLUSIONS: The results of our investigations confirm the possibility of generating T regulatory cells from conventional T lymphocytes from peripheral blood of children with newly recognised type 1 diabetes mellitus.

Transformation of conventional T cells into regulatory T cells in children with metabolic syndrome.

BACKGROUND: Much research has been done in the recent years to establish an association between obesity, metabolic syndrome and the immune system. Numerous data suggest that the decreased number and/or function of regulatory T cells (Treg cells) can lead to chronic minimal inflammation present in patients with obesity and trigger formation of atherosclerotic plaque. AIM: To generate Treg cells from the peripheral blood in children meeting the diagnostic criteria of metabolic syndrome. METHODS: A total of 25 children with metabolic syndrome and 25 controls were enrolled in the study. Peripheral blood was collected, CD4(+)/CD25(-) cells were separated and cultured for 4 weeks in the presence of a Treg expander (CD3/CD28) and interleukin-2. The expression of the transcription factor FoxP3 as a Treg marker was assessed before and after culture using reverse transcriptase polymerase chain reaction (RT-PCR) and flow cytometry. RESULTS: Before the culture we observed a slightly lower percentage of Treg cells in children with metabolic syndrome vs controls. After the culture we noted a significant increase in mRNA expression and in the percentage of FoxP3-positive cells. We observed no differences in the results between the children with metabolic syndrome and the controls. CONCLUSIONS: Our study shows that it is possible to generate Treg cells from peripheral blood of children with metabolic syndrome. In future, these findings could be used to develop a model of immunotherapeutic intervention for patients at risk of cardiovascular disease.

Diminished expression of ICOS, GITR and CTLA-4 at the mRNA level in T regulatory cells of children with newly diagnosed type 1 diabetes.

Diabetes mellitus is one of the most common chronic diseases in children. T regulatory cells (Tregs) modulate response to autoantigens and probably play a role in pathogenesis of type 1 diabetes (T1DM). The aim of the present study was the assessment of T regulatory cells including their percentages and expression of critical genes in these cells in children with newly diagnosed type 1 diabetes. The examined group consisted of 50 children with T1DM. A flow cytometric analysis of T-cell subpopulations was performed using the following markers: anti-CD4, anti-CD25 and anti-CD127 (=IL-7R). Additionally, T regulatory cells were isolated for assessment of mRNA levels for chosen genes with the real-time RT-PCR technique. The percentages of CD4(+)CD25(high)CD127(dim/-) were very low and did not differ between T1DM and control children. We did not observe any statistically significant differences between healthy and diabetic children in mRNA expression for FoxP3, IL-7R (CD127), IL-8RA, IL-10RA, IL-12A, IL-2RA (CD25), IL-21, STAT1, STAT3, SOCS2, SOCS3, TGF-beta1-R1, TGF-beta-R2 and TBX-21 genes. Interestingly the mRNA level for CTLA-4, ICOS1, IL-23, IL-27, SMAD3 and GITR were lower in Treg cells of children with diabetes compared to the control patients. No disturbances in the percentages of T regulatory cells in patients with diabetes but diminished expression of some elements important in Treg function could be the result of an immunologic imbalance accompanying the onset of the diabetes. The results of our study should be used in future research in the field of immunotherapy in pediatric diabetes.

Lower percentages of T regulatory cells in children with type 1 diabetes - preliminary report.

INTRODUCTION: Type 1 diabetes (T1D) isa well-known autoimmune disease, however there are still some processes in its pathogenesis to be elucidated. In the last few years the role of T regulatory cells in the pathogenesis of T1D has been investigated. The aim of study was to determine the percentages and numbers of T regulatory cells in the peripheral blood of children with type 1 diabetes. MATERIAL AND METHODS: A total of 25 children with newly diagnosed type 1 diabetes were studied, and compared to the control group consisted of 30 healthy children with no signs of autoimmune, chronic, inflammatory, neoplastic disease, and no evidence of T1D in their families. Flow cytometric analysis ofT-cell subpopulations was performed using the following markers: anti-CD3, anti-CD4, anti-CD25, and anti-CD127 (IL-7R). RESULTS: In the group of children with type 1 diabetes we noted statistically significant lower percentages of T regulatory cells, i.e., CD4+CD25 high and CD4+CD127 low comparing to the control children. There were no differences in other assessed parameters: white blood cell count, lymphocytes, CD4+ and CD4+CD25 high CD127 low cells (percentage and count). We did not find the correlation between patients' age and any of analysed parameters. CONCLUSIONS: Our results suggest the lower percentages of Tregs in children with T1D, however those data need to be confirmed in a larger cohort of patients and complemented with functional studies, e.g. at mRNA level. In our opinion T regulatory cells could be excellent candidates for cell therapy in newly diagnosed type 1 diabetic children.