Activation of Calcium-Sensing Receptor increases intracellular calcium and decreases cAMP and mTOR in PKD1 deficient cells

Abstract

Clinical and fundamental research suggest that altered calcium and cAMP signaling might be the most proximal events in ADPKD pathogenesis. Cells from ADPKD cysts have a reduced resting cytosolic calcium [Ca²⁺]_i and increased cAMP levels. CaSR plays an essential role in regulating calcium homeostasis. Its activation is associated with [Ca²⁺]_i increase and cAMP decrease, making CaSR a possible therapeutic target. Human conditionally immortalized Proximal Tubular Epithelial cells (ciPTEC) with stable knockdown of PKD1 (ciPTEC-PC1KD) and ciPTEC generated from an ADPKD1 patient (ciPTEC-PC1Pt) were used as experimental tools. CaSR functional expression was confirmed by studies showing that the calcimimetic NPS-R568 induced a significant increase in [Ca²⁺]_i in ciPTEC-PC1KD and ciPTEC-PC1Pt. Resting [Ca²⁺]_i were significantly lower in ciPTEC-PC1KD with respect to ciPTECwt, confirming calcium dysregulation. As in native cyst cells, significantly higher cAMP levels and mTOR activity were found in ciPTEC-PC1KD compared to ciPTECwt. Of note, NPS-R568 treatment significantly reduced intracellular cAMP and mTOR activity in ciPTEC-PC1KD and ciPTEC-PC1Pt. To conclude, we demonstrated that selective CaSR activation in human ciPTEC carrying PKD1 mutation increases [Ca²⁺]_i, reduces intracellular cAMP and mTOR activity, reversing the principal dysregulations considered the most proximal events in ADPKD pathogenesis, making CaSR a possible candidate as therapeutic target.