Presence of TRPA1 Modifies CD4+/CD8+ T Lymphocyte Ratio and Activation

Abstract

Transient Receptor Potential Ankyrin 1 (TRPA1) has been reported to influence neuroinflammation and lymphocyte function. We analysed the immune phenotype and activation characteristics of TRPA1-deficient mice (knockout—KO) generated by targeted deletion of the pore-loop domain of the ion channel. We compared TRPA1 mRNA and protein expression in monocyte and lymphocyte subpopulations isolated from primary and secondary lymphatic organs of wild type (WT) and KO mice. qRT-PCR and flow cytometric studies indicated a higher level of TRPA1 in monocytes than in lymphocytes, but both were orders of magnitude lower than in sensory neurons. We found lower CD4+/CD8+ thymocyte ratios, diminished CD4/CD8 rates, and B cell numbers in the KO mice. Early activation marker CD69 was lower in CD4+ T cells of KO, while the level of CD8+/CD25+ cells was higher. In vitro TcR-mediated activation did not result in significant differences in CD69 level between WT and KO splenocytes, but lower cytokine (IL-1β, IL-6, TNF-α, IL-17A, IL-22, and RANTES) secretion was observed in KO splenocytes. Basal intracellular Ca2+ level and TcR-induced Ca2+ signal in T lymphocytes did not differ significantly, but interestingly, imiquimod-induced Ca2+ level in KO thymocytes was higher. Our results support the role of TRPA1 in the regulation of activation, cytokine production, and T and B lymphocytes composition in mice.

TRPA1 Covalent Ligand JT010 Modifies T Lymphocyte Activation

“…The present results are consistent with our earlier results by qRT-PCR analyses, where we also detected low levels of Trpa1 mRNA in monocytes and lymphocytes isolated from primary and secondary lymphoid organs of mice [39]. While the expression of Trpa1 mRNA was reported in mouse CD4 + splenocytes and thymocytes either by RT-PCR or qRT-PCR analyses [34,35,39], other studies did not support transcription of Trpa1 in lymphocytes or in peritoneal macrophages [83][84][85]. The differences in these findings may be explained by the methodical differences.…”

“…As the representative plots in Figure 5e demonstrate, 40 µM of JT010 treatment did not cause elevation of the Annexin V labeling or the 7-AAD fluorescence of PEC lymphocytes; only addition of ionomycin (2 µM) at the end of the experiment resulted in maximum PS exposure of cells. Taken together, JT010 administration, even at high concentrations, did not modify cell size and granulation, 7-AAD staining or Annexin V-binding, as had been detected and reported earlier by other agonists (AITC, cinnamaldehyde) and antagonists (HC-030031 and A967079) [39].…”

“…PEC lymphocytes; only addition of ionomycin (2 µM) at the end of the experiment resulted in maximum PS exposure of cells. Taken together, JT010 administration, even at high concentrations, did not modify cell size and granulation, 7-AAD staining or Annexin V-binding, as had been detected and reported earlier by other agonists (AITC, cinnamaldehyde) and antagonists (HC-030031 and A967079) [39]. As detailed in Figure 4a, we gated PEC lymphocytes based on FSC/SSC parameter distribution; in these cases, the T lymphocyte activation was distinguished based on the specific nature of the activation, since anti-CD3 cross-linking activates T cells selectively (see detailed method in Section 2.5).…”

Mechanosensitive Ion Channels: Their Physiological Importance and Potential Key Role in Cancer

“…In a melanoma orthotopic mouse model, Trpa1 −/− mice showed higher CD8+ T-cell activation and tumor infiltration as well as lower tumor progression than Trpa1 +/+ animals [ 152 ], indicating that TRPA1 diminishes CD8+ T-cell cytotoxicity. Likewise, a recent publication highlights that Trpa1 −/− mice show a lower CD4+/CD8+ T-cell ratio and higher CD8+ cell activation than WT mice, and this TRPA1 deficiency also reduced the percentage of peripheral blood CD19+ B cells [ 153 ]. On the other hand, the role of TRPA1 in CD4+ T-cell biology is controversial.…”

“…On the other hand, the role of TRPA1 in CD4+ T-cell biology is controversial. While some studies report that TRPA1 restrains CD4+ T-cell activation by a crosstalk mechanism that implies the inhibition of TRPV1 [ 154 ], others indicate that its inhibition is correlated with the reduction of CD4+ T-cell activation markers such as CD69 [ 137 , 153 ], thus requiring further research to fully understand the role of TRPA1 in CD4+ T-cell biology.…”