Elsevier

Clinical Immunology

Volume 181, August 2017, Pages 67-74
Clinical Immunology

Attenuated IL-2R signaling in CD4 memory T cells of T1D subjects is intrinsic and dependent on activation state

https://doi.org/10.1016/j.clim.2017.06.004Get rights and content

Highlights

  • Low response to IL-2 is most prominent in memory CD4 Teff of T1D subjects.

  • Multiple factors influence T1D-associated low IL-2 response in Teff.

  • Low, but sustained TCR activation correlates with low basal IL-2R signaling.

  • Activation enhances low IL-2 responsiveness suggesting a regulated response.

Abstract

The IL-2/IL-2R pathway is implicated in type 1 diabetes (T1D). While its role in regulatory T cell (Treg) biology is well characterized, mechanisms that influence IL-2 responses in effector T cells (Teff) are less well understood. We compared IL-2 responses in 95 healthy control and 98 T1D subjects. In T1D, low IL-2 responsiveness was most pronounced in memory Teff. Unlike Treg, CD25 expression did not influence the Teff responses. Reduced IL-2 responses in memory Teff were not rescued by resting, remained lower after activation and proliferation, and were absent in type 2 diabetes. Comparing basal IL-2 responses in resting versus activated cells, memory Teff displayed lower, but more sustained, responses to IL-2 overtime. These results suggest that T1D-associated defects in the Teff compartment are due to intrinsic factors related to activation. Evaluation of both Teff and Treg IL-2R signaling defects in T1D subjects may inform selection of therapies.

Introduction

Type 1 diabetes (T1D) is a complex autoimmune disease caused by the selective destruction of insulin producing beta cells in the pancreatic islet. This immune-mediated destruction of beta cells ultimately leads to a life-long dependence on exogenous insulin treatment. While the etiology of T1D is unknown, it is clear that both genetic and environmental factors contribute to disease susceptibility and progression. Beta-cell specific T cells are known to play a key role in T1D, yet there is also strong evidence that systemic immunological defects impact T1D pathogenesis. Gaining a better understanding of the character of the global immune response may lead to discovery of new areas for disease prevention and intervention.

Low dose IL-2 therapy augments the frequency of regulatory CD4 T cells (Treg), an IL-2 dependent cell-type that constitutively expresses high levels of CD25. There is great interest in using low dose IL-2 therapy to boost Tregs in T1D given that genetic variants in the IL-2/IL-2R pathway are associated with T1D, and defects in both IL-2R signaling and Treg stability have been observed in T1D in humans and mouse models [1], [2], [3], [4], [5]. Beneficial clinical responses have been observed in low dose IL-2 trials in graft versus host disease, vasculitis, lupus and alopecia patients [6]. In these clinical settings, IL-2 therapy clearly augments Treg, but may also impact other cell types [2]. In a recent dose-escalation Phase I adaptive trial in T1D, a range of IL-2 doses that increase Treg frequency in peripheral blood was identified [7]. However, in the same trial, other cell types also responded to IL-2 including Teff at all doses tested and there was significant heterogeneity across subjects. This finding confirms initial trials with IL-2 therapy in T1D that suggest there is a narrow dosing window that varies across subjects to achieve Treg-specific tolerogenic effects [8], [9]. To this point, it is imperative to better understand patient heterogeneity and underlying mechanisms of disease by clearly defining the degree, cell source, and cause of variability in IL-2/IL-2R signaling.

It remains to be determined to what degree IL-2/IL-2R signaling variability is due to differences in the intrinsic or extrinsic regulation of this pathway. Intrinsically in humans, genome wide association studies have identified multiple T1D-associated single nucleotide polymorphisms (SNPs) that are located in genes (IL-2, IL2RA, and PTPN2) that encode proteins in the IL-2/IL-2R signaling pathway [3], [4]. Yet, this does not explain low IL-2 response in all subjects. IL-2R signaling is tuned at multiple levels with expression level of CD25 being key. Other intrinsic factors including signaling molecules and miRNA have been shown to control signaling in model systems [10], [11]. Extrinsically, metabolic factors and secreted receptors [12], [13], [14] regulate IL-2R signaling in other disease settings. In this study, we examine IL-2 responsiveness in 98 T1D subjects and 95 healthy controls and explore underlying mechanisms in Teff. Our results demonstrate that disease impacts IL-2/IL-2R signaling most prominently in the CD4 Teff compartment. Furthermore, these effects are most likely acquired and intrinsic given that they are present in memory (but not naive) cells, are not present in type 2 diabetic (T2D) subjects, persist following activation and proliferation, and are altered by activation state.

Section snippets

Human subjects and sample handling

Peripheral blood mononuclear cells (PBMC) were collected from subjects enrolled in the Benaroya Research Institute (BRI) Immune Mediated Disease Registry and Repository. Written informed consent was obtained from all subjects according to IRB approved protocols at BRI, Seattle Washington, USA. Age and gender matched healthy controls were selected for each experiment based on lack of personal or family history of autoimmunity or asthma. Control and disease subjects were included in daily

Attenuated IL-2 responses in T1D are prominent in memory CD4 Teff

Heterogeneity of subjects and cell types can confound some immunological findings in smaller studies. To better understand this heterogeneity, we compared IL-2 responses in T cell subsets in a large number (98 T1D and 95 healthy control) of subjects matched for age, gender and ethnicity (Table 1). We chose a dose of IL-2 known to elicit differences in all T cell subsets, and that is not driven exclusively by IL-2 availability and CD25 expression level. Importantly, this enabled measurement of

Conclusion

This comprehensive study expands our understanding of underlying mechanisms of reduced IL-2R signaling in a cell-type specific manner, underscores the heterogeneity across subjects and cell types, and highlights the predominance of acquired, disease-associated IL-2R signaling defects in the Teff compartment in T1D that may be linked to chronic activation or differentiation state. Together, these Treg and Teff IL-2 signaling defects could inform selection of therapies for an individual. For

Grant support

This work was supported by a JDRF Career Development Award 3-2012-205 to SAL, and NIH grants AI101990, AI083455, and DK097672 to JHB.

Acknowledgements

We are very grateful to the investigators and staff of the BRI Diabetes Clinical Research Program for subject recruitment, as well as the BRI Translational Research Clinical Core for sample processing and handling. We thank all members of the Human Immunophenotyping Core who helped with standardization and acquisition of flow cytometry data in the large cohorts of controls and T1D subjects. We thank Anne Hocking for review of the article and assistance with submission.

References (44)

  • D. Klatzmann et al.

    The promise of low-dose interleukin-2 therapy for autoimmune and inflammatory diseases

    Nat. Rev. Immunol.

    (2015)
  • J.A. Todd et al.

    Regulatory T Cell responses in participants with type 1 diabetes after a single dose of interleukin-2: a non-randomised, open label, adaptive dose-finding trial

    PLoS Med.

    (2016)
  • L.A. Truman et al.

    Protocol of the adaptive study of IL-2 dose frequency on regulatory T cells in type 1 diabetes (DILfrequency): a mechanistic, non-randomised, repeat dose, open-label, response-adaptive study

    BMJ Open

    (2015)
  • A. Yu et al.

    Selective IL-2 responsiveness of regulatory T cells through multiple intrinsic mechanisms support the use of low-dose IL-2 therapy in Type-1 diabetes

    Diabetes

    (2015)
  • Y. Rochman et al.

    New insights into the regulation of T cells by gamma(c) family cytokines

    Nat. Rev. Immunol.

    (2009)
  • V. De Rosa et al.

    Glycolysis controls the induction of human regulatory T cells by modulating the expression of FOXP3 exon 2 splicing variants

    Nat. Immunol.

    (2015)
  • S.A. Long et al.

    Defects in IL-2R signaling contribute to diminished maintenance of FOXP3 expression in CD4 + CD25 + regulatory T cells of type 1 diabetic subjects

    Diabetes

    (2010)
  • K. Cerosaletti et al.

    Multiple autoimmune-associated variants confer decreased il-2R signaling in CD4(+)CD25(hi) T Cells of type 1 diabetic and multiple sclerosis patients

    PLoS One

    (2013)
  • S.A. Long et al.

    Functional islet-specific Treg can be generated from CD4(+)CD25(−) T cells of healthy and type 1 diabetic subjects

    Eur. J. Immunol.

    (2009)
  • A. Schneider et al.

    The effector T cells of diabetic subjects are resistant to regulation via CD4 + FOXP3 + regulatory T cells

    J. Immunol.

    (2008)
  • J.H. Yang et al.

    Natural variation in interleukin-2 sensitivity influences regulatory T-cell frequency and function in individuals with long-standing type 1 diabetes

    Diabetes

    (2015)
  • A.K. Marwaha et al.

    Pre-diagnostic genotyping identifies T1D subjects with impaired Treg IL-2 signaling and an elevated proportion of FOXP3 + IL-17 + cells

    Genes Immun.

    (2017)
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