AMPKα1-Sensitivity of Orai1 and Ca²⁺ Entry in T - Lymphocytes

Abstract

Background/Aims: T-lymphocyte activation and function critically depends on Ca²⁺ signaling, which is regulated by store operated Ca²⁺ entry (SOCE). Human and mouse T lymphocytes express AMP activated kinase AMPKα1, which is rapidly activated following elevation of cytosolic Ca²⁺ concentration ([Ca²⁺]_i) by treatment of the cells with Ca²⁺ ionophore or following inhibition of endosomal Ca²⁺ ATPase with thapsigargin. AMPK is further activated by triggering of the T cell antigen receptor (TCR). The present study explored whether AMPK influences Ca²⁺ entry and Ca²⁺-sensitive regulation of T-lymphocyte function. Methods: T-lymphocytes were isolated and cultured from AMPKα1-deficient (ampk^-/-) mice and from their wildtype (ampk^+/+) littermates. The phenotype of the cells was analysed by flow cytometry, [Ca²⁺]_i estimated from Fura-2 fluorescence, SOCE from increase of [Ca²⁺]_i following thapsigargin treatment (1 µM), and cell function analysed by measuring cytokine secretion and western blotting. Results: Expression of surface markers in CD4⁺ and CD8⁺ T-cells were similar in ampk^-/- and ampk^+/+ T-lymphocyte blasts. Moreover, total STIM1 protein abundance was similar in ampk^-/- and ampk^+/+ T-lymphocyte blasts. However, Orai1 cell membrane protein abundance was significantly higher in ampk^-/- than in ampk^+/+ T-lymphocyte blasts. SOCE and increase of [Ca²⁺]_i following TCR activation by triggering TCR with anti-CD3 and cross-linking secondary antibody were both significantly more pronounced in ampk^-/- than in ampk^+/+ T-lymphocyte blasts. The difference of Ca²⁺ entry between ampk^-/- and ampk^+/+ T-lymphocytes was abrogated by Orai1 inhibitor 2-aminoethoxydiphenyl borate (2-APB, 50 µM). Proliferation of unstimulated ampk^-/- lymphocytes was higher than proliferation of ampk^+/+ T-lymphocytes, a difference reversed by Orai1 silencing. Conclusions: AMPK downregulates Orai1 and thus SOCE in T-lymphocytes and thus participates in negative feed-back regulation of cytosolic Ca²⁺ activity.

Exendin-4 Alleviates High Glucose-Induced Rat Mesangial Cell Dysfunction through the AMPK Pathway

“…Correlative studies have shown that exendin-4 can enhance the phosphorylation of AMPK in the liver of DIO mice and in hepatocytes [17,21], which was down-regulated in the kidneys of db/db mice [18]. Initially, AMPK was solely recognized as a highly conserved sensor of cellular energy that regulated Ca 2+ and K + channels in different cell types [22,23]. However, further studies have demonstrated that AMPK also plays a role in the amelioration of epithelial cell damage caused by renal ischemia [24] and cell proliferation through the attenuation of MEK-ERK signaling [25] as well as mTOR activity [26,27].…”

AMP-Activated Protein Kinase α1 Regulates Cardiac Gap Junction Protein Connexin 43 and Electrical Remodeling Following Pressure Overload

“…Ampk is a powerful regulator of channels, carriers and Na + /K + ATPase [10,11,12,13,14,15,16,17,18,19]. The kinase is involved in the regulation of cardiac sodium and potassium channels [11,17].…”

Lithium- Sensitive Store-Operated Ca²⁺ Entry in the Regulation of FGF23 Release

“…Orai1 is upregulated by the serum & glucocorticoid inducible kinase SGK1 [61][62][63]75], which is in turn markedly upregulated by cell shrinkage [76]. Orai1 is further regulated by the AMPK activated kinase [77]. Future studies will be required to explore whether this kinase participates in the regulation of FGF23 release.…”