Limits...
Foxp3+ Treg expanded from patients with established diabetes reduce Helios expression while retaining normal function compared to healthy individuals.

Du W, Shen YW, Lee WH, Wang D, Paz S, Kandeel F, Liu CP - PLoS ONE (2013)

Bottom Line: After isolation, Treg from both T1D patients and healthy subjects were successfully expanded with high purity.Importantly, expanded Treg from both subject groups were able to suppress autologous or allogeneic CD8(+) effector T cells equally well.Thus, based on the positive outcomes, these potent expanded Treg from diabetic human patients may be useful in treating T1D or preventing islet graft rejection.

View Article: PubMed Central - PubMed

Affiliation: Department of Diabetes and Metabolic Diseases Research, Beckman Research Institute, City of Hope, Duarte, California, United States of America.

ABSTRACT
Foxp3(+) regulatory T cells (Treg) play a crucial role in regulating immune tolerance. The use of Treg to restore immune tolerance is considered an attractive novel approach to inhibit autoimmune disease, including type 1 diabetes (T1D), and to prevent rejection of organ transplants. In view of the goal of developing autologous Treg-based cell therapy for patients with long-term (>15 years) T1D, it will be necessary to expand a sufficient amount of functional Treg in vitro in order to study and compare Treg from T1D patients and healthy subjects. Our results have demonstrated that there is a comparable frequency of Treg in the peripheral blood lymphocytes (PBLs) of patients with long-term T1D relative to those in healthy subjects; however, Th1 cells, but not Th17 cells, were increased in the T1D patients. Further, more Treg in PBLs from T1D patients than from healthy subjects expressed the CD45RO(+) memory cell phenotype, suggesting they were antigen-experienced cells. After isolation, Treg from both T1D patients and healthy subjects were successfully expanded with high purity. Although there was no difference in Helios expression on Treg in PBLs, in vitro expansion led to fewer Helios-expressing Treg from T1D patients than healthy subjects. While more Th1-like Treg expressing IFN-γ or TNF-α were found in the PBLs of T1D patients than healthy controls, there was no such difference in the expanded Treg. Importantly, expanded Treg from both subject groups were able to suppress autologous or allogeneic CD8(+) effector T cells equally well. Our findings demonstrate that a large number of ex vivo expanded functional Treg can be obtained from long-term T1D patients, although fewer expanded Treg expressed a high level of Helios. Thus, based on the positive outcomes, these potent expanded Treg from diabetic human patients may be useful in treating T1D or preventing islet graft rejection.

Show MeSH

Related in: MedlinePlus

In vitro expansion leads to decreased expression of Helios on T1D patient but not healthy subject Treg.PBMCs and expanded Treg were stained with indicated antibodies and analyzed by FACS. (A) Percentage of Helios+ cells in total CD4+ T cells (CD4+Helios+) or in Foxp3+CD4+ T cells (CD4+Foxp3+Helios+) in PBLs of T1D patients or healthy controls. (B) FACS analyses of Helios and Foxp3 expression in PBLs and expanded Treg on D14 from one representative individual of each subject group. The cells were electronically gated on CD4+ T cell population. (C) Collective analyses of the percentage of Helios+ cell frequency in PBLs or expanded Foxp3+ Treg on D14 in each subject group. (D) Paired comparison analyses of Helios expression on Foxp3+ Treg in PBLs vs. its expression on expanded Foxp3+ Treg in each subject group.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3569420&req=5

pone-0056209-g004: In vitro expansion leads to decreased expression of Helios on T1D patient but not healthy subject Treg.PBMCs and expanded Treg were stained with indicated antibodies and analyzed by FACS. (A) Percentage of Helios+ cells in total CD4+ T cells (CD4+Helios+) or in Foxp3+CD4+ T cells (CD4+Foxp3+Helios+) in PBLs of T1D patients or healthy controls. (B) FACS analyses of Helios and Foxp3 expression in PBLs and expanded Treg on D14 from one representative individual of each subject group. The cells were electronically gated on CD4+ T cell population. (C) Collective analyses of the percentage of Helios+ cell frequency in PBLs or expanded Foxp3+ Treg on D14 in each subject group. (D) Paired comparison analyses of Helios expression on Foxp3+ Treg in PBLs vs. its expression on expanded Foxp3+ Treg in each subject group.

Mentions: Helios, an Ikaros transcription factor family member, has been reported as a marker that distinguishes thymically-derived nTreg from peripherally-induced Treg (iTreg) and/or as a marker for T-cell activation and proliferation [32]–[34]. We first compared the expression of Helios on non-cultured CD4+ T cells and CD4+Foxp3+ Treg in PBLs of T1D and healthy subjects and in the expanded Foxp3+ Treg from their PBLs. Our analyses on non-cultured CD4+ T cells showed comparable percentages of Helios+ or Helios+Foxp3+ CD4+ T cells detected in T1D subjects with those detected in healthy controls (Fig. 4A). Next, we assessed Helios expression within the CD4+Foxp3+ Treg population in PBLs, and found no difference of Helios expression in these Treg between the two subject groups (Fig. 4B–C). In addition, there was also no difference in the Helios expression on Foxp3+ Treg in CD4+CD25+CD127−/lo PBLs from T1D patients or healthy subjects (data not shown).


Foxp3+ Treg expanded from patients with established diabetes reduce Helios expression while retaining normal function compared to healthy individuals.

Du W, Shen YW, Lee WH, Wang D, Paz S, Kandeel F, Liu CP - PLoS ONE (2013)

In vitro expansion leads to decreased expression of Helios on T1D patient but not healthy subject Treg.PBMCs and expanded Treg were stained with indicated antibodies and analyzed by FACS. (A) Percentage of Helios+ cells in total CD4+ T cells (CD4+Helios+) or in Foxp3+CD4+ T cells (CD4+Foxp3+Helios+) in PBLs of T1D patients or healthy controls. (B) FACS analyses of Helios and Foxp3 expression in PBLs and expanded Treg on D14 from one representative individual of each subject group. The cells were electronically gated on CD4+ T cell population. (C) Collective analyses of the percentage of Helios+ cell frequency in PBLs or expanded Foxp3+ Treg on D14 in each subject group. (D) Paired comparison analyses of Helios expression on Foxp3+ Treg in PBLs vs. its expression on expanded Foxp3+ Treg in each subject group.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3569420&req=5

pone-0056209-g004: In vitro expansion leads to decreased expression of Helios on T1D patient but not healthy subject Treg.PBMCs and expanded Treg were stained with indicated antibodies and analyzed by FACS. (A) Percentage of Helios+ cells in total CD4+ T cells (CD4+Helios+) or in Foxp3+CD4+ T cells (CD4+Foxp3+Helios+) in PBLs of T1D patients or healthy controls. (B) FACS analyses of Helios and Foxp3 expression in PBLs and expanded Treg on D14 from one representative individual of each subject group. The cells were electronically gated on CD4+ T cell population. (C) Collective analyses of the percentage of Helios+ cell frequency in PBLs or expanded Foxp3+ Treg on D14 in each subject group. (D) Paired comparison analyses of Helios expression on Foxp3+ Treg in PBLs vs. its expression on expanded Foxp3+ Treg in each subject group.
Mentions: Helios, an Ikaros transcription factor family member, has been reported as a marker that distinguishes thymically-derived nTreg from peripherally-induced Treg (iTreg) and/or as a marker for T-cell activation and proliferation [32]–[34]. We first compared the expression of Helios on non-cultured CD4+ T cells and CD4+Foxp3+ Treg in PBLs of T1D and healthy subjects and in the expanded Foxp3+ Treg from their PBLs. Our analyses on non-cultured CD4+ T cells showed comparable percentages of Helios+ or Helios+Foxp3+ CD4+ T cells detected in T1D subjects with those detected in healthy controls (Fig. 4A). Next, we assessed Helios expression within the CD4+Foxp3+ Treg population in PBLs, and found no difference of Helios expression in these Treg between the two subject groups (Fig. 4B–C). In addition, there was also no difference in the Helios expression on Foxp3+ Treg in CD4+CD25+CD127−/lo PBLs from T1D patients or healthy subjects (data not shown).

Bottom Line: After isolation, Treg from both T1D patients and healthy subjects were successfully expanded with high purity.Importantly, expanded Treg from both subject groups were able to suppress autologous or allogeneic CD8(+) effector T cells equally well.Thus, based on the positive outcomes, these potent expanded Treg from diabetic human patients may be useful in treating T1D or preventing islet graft rejection.

View Article: PubMed Central - PubMed

Affiliation: Department of Diabetes and Metabolic Diseases Research, Beckman Research Institute, City of Hope, Duarte, California, United States of America.

ABSTRACT
Foxp3(+) regulatory T cells (Treg) play a crucial role in regulating immune tolerance. The use of Treg to restore immune tolerance is considered an attractive novel approach to inhibit autoimmune disease, including type 1 diabetes (T1D), and to prevent rejection of organ transplants. In view of the goal of developing autologous Treg-based cell therapy for patients with long-term (>15 years) T1D, it will be necessary to expand a sufficient amount of functional Treg in vitro in order to study and compare Treg from T1D patients and healthy subjects. Our results have demonstrated that there is a comparable frequency of Treg in the peripheral blood lymphocytes (PBLs) of patients with long-term T1D relative to those in healthy subjects; however, Th1 cells, but not Th17 cells, were increased in the T1D patients. Further, more Treg in PBLs from T1D patients than from healthy subjects expressed the CD45RO(+) memory cell phenotype, suggesting they were antigen-experienced cells. After isolation, Treg from both T1D patients and healthy subjects were successfully expanded with high purity. Although there was no difference in Helios expression on Treg in PBLs, in vitro expansion led to fewer Helios-expressing Treg from T1D patients than healthy subjects. While more Th1-like Treg expressing IFN-γ or TNF-α were found in the PBLs of T1D patients than healthy controls, there was no such difference in the expanded Treg. Importantly, expanded Treg from both subject groups were able to suppress autologous or allogeneic CD8(+) effector T cells equally well. Our findings demonstrate that a large number of ex vivo expanded functional Treg can be obtained from long-term T1D patients, although fewer expanded Treg expressed a high level of Helios. Thus, based on the positive outcomes, these potent expanded Treg from diabetic human patients may be useful in treating T1D or preventing islet graft rejection.

Show MeSH
Related in: MedlinePlus