Loss of apical monocilia on collecting duct principal cells impairs ATP secretion across the apical cell surface and ATP-dependent and flow-induced calcium signals

Purinergic Signalling

Michael B. Hovater¹  Brian Siroky²  Bradley K. Yoder²  Elizabeth L. Hanson¹  Nai-Lin Cheng¹  P. Darwin Bell²  Dragos Olteanu¹  Erik M. Schwiebert³  Peter Komlosi²  Wen Liu⁴  Lisa M. Satlin⁴  Attila Fintha¹ 

[1] University of Alabama at Birmingham$$;University of Alabama at Birmingham$$University of Alabama at Birmingham$$;University of Alabama at Birmingham$$University of Alabama at Birmingham$$University of Alabama at Birmingham$$University of Alabama at Birmingham$$;Mount Sinai School of Medicine$$

关键词: Cilia;   

DOI  :  10.1007/s11302-007-9072-0

学科分类：分子生物学，细胞生物学和基因

来源: Springer

【 摘 要 】

Renal epithelial cells release ATP constitutively under basal conditions and release higher quantities of purine nucleotide in response to stimuli. ATP filtered at the glomerulus, secreted by epithelial cells along the nephron, and released serosally by macula densa cells for feedback signaling to afferent arterioles within the glomerulus has important physiological signaling roles within kidneys. In autosomal recessive polycystic kidney disease (ARPKD) mice and humans, collecting duct epithelial cells lack an apical central cilium or express dysfunctional proteins within that monocilium. Collecting duct principal cells derived from an Oak Ridge polycystic kidney (orpk^Tg737) mouse model of ARPKD lack a well-formed apical central cilium, thought to be a sensory organelle. We compared these cells grown as polarized cell monolayers on permeable supports to the same cells where the apical monocilium was genetically rescued with the wild-type Tg737 gene that encodes Polaris, a protein essential to cilia formation. Constitutive ATP release under basal conditions was low and not different in mutant versus rescued monolayers. However, genetically rescued principal cell monolayers released ATP three- to fivefold more robustly in response to ionomycin. Principal cell monolayers with fully formed apical monocilia responded three- to fivefold greater to hypotonicity than mutant monolayers lacking monocilia. In support of the idea that monocilia are sensory organelles, intentionally harsh pipetting of medium directly onto the center of the monolayer induced ATP release in genetically rescued monolayers that possessed apical monocilia. Mechanical stimulation was much less effective, however, on mutant orpk collecting duct principal cell monolayers that lacked apical central monocilia. Our data also show that an increase in cytosolic free Ca²⁺ primes the ATP pool that is released in response to mechanical stimuli. It also appears that hypotonic cell swelling and mechanical pipetting stimuli trigger release of a common ATP pool. Cilium-competent monolayers responded to flow with an increase in cell Ca²⁺ derived from both extracellular and intracellular stores. This flow-induced Ca²⁺ signal was less robust in cilium-deficient monolayers. Flow-induced Ca²⁺ signals in both preparations were attenuated by extracellular gadolinium and by extracellular apyrase, an ATPase/ADPase. Taken together, these data suggest that apical monocilia are sensory organelles and that their presence in the apical membrane facilitates the formation of a mature ATP secretion apparatus responsive to chemical, osmotic, and mechanical stimuli. The cilium and autocrine ATP signaling appear to work in concert to control cell Ca²⁺. Loss of a cilium-dedicated autocrine purinergic signaling system may be a critical underlying etiology for ARPKD and may lead to disinhibition and/or upregulation of multiple sodium (Na⁺) absorptive mechanisms and a resultant severe hypertensive phenotype in ARPKD and, possibly, other diseases.

【 授权许可】

Unknown