Human CCR6+ Th17 Lymphocytes Are Highly Sensitive to Radiation-Induced Senescence and Are a Potential Target for Prevention of Radiation-Induced Toxicity

Abstract

Purpose: This study addresses the sensitivity of different peripheral CD4⁺ T-lymphocyte subsets to irradiation (IR) and identifies potential targets for the prevention or treatment of radiation-induced toxicity.

Methods: This study was performed on peripheral blood mononuclear cells or sorted peripheral memory lymphocytes of CCR6⁺ mucosa-homing Th17/CCR6^negTh and regulatory T subtypes of healthy volunteers. Cells were irradiated with a 2 Gy with or without pharmacologic inhibitors of different signaling pathways. Senescence of irradiated cells was assessed by resistance to apoptosis and determination of various senescence-associated biomarkers (senescence associated b-galactosidase activity, p16Ink4a-, p21Cdkn1a-, gH2A.X-, H2A.J expression). Cytokine production was measured in supernatants of irradiated cells by Luminex technology.

Results: Not all CD4⁺ memory T lymphocyte subsets were equally radiosensitive. High sensitivity of CCR6⁺Th17 lymphocytes to IR-induced senescence was shown by expression of the histone variant H2A.J, higher SA-b-Gal activity, and upregulation of p16^Ink4a and p21^Cdkn1a expression. Lower Annexin V staining and cleaved caspase-3, and higher expression of antiapoptotic genes Bcl-2 and Bcl-xL LF, showed that CCR6⁺Th17 lymphocytes were more resistant to IR-induced apoptosis than CCR6^neg memory Th and regulatory T lymphocytes. After a 2 Gy IR, both CCR6⁺Th17 and CCR6^neg cells acquired a moderate senescence-associated secretory phenotype, but only CCR6⁺Th17 cells secreted interleukin 8 (IL-8) and vascular endothelial growth factor-A (VEGF-A). Pharmacologic targeting of reactive oxygen species (ROS), mitogen-activated protein kinases (MAPKs), and mammalian target of rapamycin (mTOR) signaling pathways prevented the expression of senescent markers and IL-8 and VEGF-A expression by CCR6⁺Th17 cells after IR.

Conclusions: This study suggests that IR induces senescence of CCR6+Th17 lymphocytes associated with secretion of IL-8 and VEGF-A that may be detrimental to the irradiated tissue. ROS-MAPKs signaling pathways are candidate targets to prevent this CCR6⁺Th17-dependent radiation-induced potential toxicity. Finally, the ratio of circulating H2A.J⁺ senescent CCR6⁺ Th17/CD4⁺ T lymphocytes may be a candidate marker of individual intrinsic radiosensitivity.