Evidence for Activation of AMP Kinase (AMPK) in Nephropathic Cystinosis

Abstract

The objective of this study was to investigate the involvement of AMPK in the altered signaling that results in nephropathic cystinosis. Cystinosis is characterized by cysteine accumulation, due to mutations in CTNS, encoding for cystinosin, a lysosomal cystine transporter. Patients develop the Fanconi Syndrome, due to defective reabsorption by apical Na+/solute cotransport systems in the renal proximal tubule (RPT). In this study, cystinosin was knocked down >80% in primary rabbit RPT cells using cystinosin siRNA. This reduction in cystinosin strongly inhibited Na+ dependent Phosphate (Pi) uptake (70%). Intracellular Na+and Na,K‐ATPase were unaffected. However, ATP dropped 52%, the ADP/ATP ratio increased 300%, and apical NaPi2a transporters were reduced. Because increases in ADP result in AMPK phosphorylation and activation, the phosphorylated AMPKα subunit (pAMPKα) was examined. In primary RPT cells with a cystinosin knockdown the level of pAMPKα was elevated (8.0 +/− 4.2 fold relative to the control), and was unaffected by AICAR (5‐Aminoimidazole‐4‐Carboxamide‐1‐β‐D‐ribofuranoside), an AMPK activator (unlike control cells). Further studies are in progress to determine whether AMPK activation is responsible for the reduced growth, increased apoptotis, as well as reduced transport in primary RPT cells with a cystinosin knockdown. The funding was from the Cystinosis Research Foundation.