Kidney Week

Abstract: TH-PO1068

Activation of the PKA Signaling Pathway Stimulates Oxalate Transport by Human Intestinal Caco2-BBE Cells

Session Information

• Mineral Disease: Nephrolithiasis
  November 02, 2017 | Location: Hall H, Morial Convention Center
  Abstract Time: 10:00 AM - 10:00 AM

Category: Mineral Disease

• 1204 Mineral Disease: Nephrolithiasis

Authors

• Hassan, Hatim A., University of Chicago, Chicago, Illinois, United States

Hatim A. Hassan,

• Arvans, Donna L., University of Chicago, Chicago, Illinois, United States

Donna L. Arvans,

• Alshaikh, Altayeb, University of Chicago, Chicago, Illinois, United States

Altayeb Alshaikh,

• Bashir, Mohamed, University of Chicago, Chicago, Illinois, United States

Mohamed Bashir,

Background

Most kidney stones are composed of calcium oxalate, and small increases in urine oxalate affect the stone risk. The mammalian intestine plays a crucial role in oxalate homeostasis and we had recently reported that oxalobacter-derived factors stimulate oxalate transport by human intestinal Caco2-BBE (C2) cells through mechanisms including PKA activation. We therefore evaluated whether intestinal oxalate transport is directly regulated by activation of the PKA signaling pathway. To this end, PKA is activated with forskolin and IBMX (F/I). F/I significantly stimulated (4-fold) ¹⁴C-oxalate transport by C2 cells (≥ 49% of which is mediated by the oxalate transporter SLC26A6 [A6]), an effect completely blocked by the PKA inhibitor H89, indicating that it is PKA-dependent. Utilizing selective pharmacological inhibitors in preliminary studies, we found that the PKC, ERK1/2, PI3K, P38, and Src kinases are not involved in the observed stimulation. Evaluating A6 total and surface (using brush border membrane vesicles) protein expression revealed that the observed stimulation is not due to changes in A6 total and surface protein expression. Assessing ¹⁴C-oxalate transport as a function of increasing ¹⁴C-oxalate concentration in the flux medium showed that the observed stimulation is due to F/I-induced increase (1.8-fold) in V_max (the maximal velocity) and reduction (2-fold) in K_m (the apparent affinity for oxalate) in preliminary studies. siRNA knockdown studies showed that significant components of the observed stimulation are mediated by the A6 and SLC26A2 (A2) oxalate transporters. Since F/I did not affect A6 total and surface protein expression, and in view of the reduced K_m (reflecting greater A6 affinity for oxalate upon PKA activation), it is likely possible that the observed stimulation is due to mechanisms including F/I-induced enhanced A6 transport activity resulting from an increase in the intrinsic activity of the preexisting A6 membrane transporters. We conclude that activation of the PKA signaling pathway significantly stimulates intestinal oxalate transport by C2 cells through mechanisms including increased intrinsic activity of preexisting A6 membrane transporters, as well as enhanced A2 transport activity (resulting from more A2 membrane transporters and/or increased intrinsic activity).

Funding

• NIDDK Support