Collecting duct cells that lack normal cilia have mislocalized vasopressin-2 receptors

Dept. of Medicine, Division of Nephrology, Medical University of South Carolina, 173 Ashley Ave., Charleston, SC 29425, USA.
AJP Renal Physiology (Impact Factor: 3.25). 12/2011; 302(7):F801-8. DOI: 10.1152/ajprenal.00253.2011
Source: PubMed

ABSTRACT Polycystic kidney disease (PKD) is a ciliopathy characterized by renal cysts and hypertension. These changes are presumably due to altered fluid and electrolyte transport in the collecting duct (CD). This is the site where vasopressin (AVP) stimulates vasopressin-2 receptor (V2R)-mediated aquaporin-2 (AQP2) insertion into the apical membrane. Since cysts frequently occur in the CD, we studied V2R and AQP2 trafficking and function in CD cell lines with stunted and normal cilia [cilia (-), cilia (+)] derived from the orpk mouse (hypomorph of the Tg737/Ift88 gene). Interestingly, only cilia (-) cells grown on culture dishes formed domes after apical AVP treatment. This observation led to our hypothesis that V2R mislocalizes to the apical membrane in the absence of a full-length cilium. Immunofluorescence indicated that AQP2 localizes to cilia and in a subapical compartment in cilia (+) cells, but AQP2 levels were elevated in both apical and basolateral membranes in cilia (-) cells after apical AVP treatment. Western blot analysis revealed V2R and glycosylated AQP2 in biotinylated apical membranes of cilia (-) but not in cilia (+) cells. In addition, apical V2R was functional upon apical desmopressin (DDAVP) treatment by demonstrating increased cAMP, water transport, and benzamil-sensitive equivalent short-circuit current (I(sc)) in cilia (-) cells but not in cilia (+) cells. Moreover, pretreatment with a PKA inhibitor abolished DDAVP stimulation of I(sc) in cilia (-) cells. Thus we propose that structural or functional loss of cilia leads to abnormal trafficking of AQP2/V2R leading to enhanced salt and water absorption. Whether such apical localization contributes to enhanced fluid retention and hypertension in PKD remains to be determined.

6 Reads
  • Source
    • "Collecting duct cells derived from the orpk mouse model expressing a hypomorphic allele of the Tg737 gene (pCDNA cells, cilia ( À)) and genetically rescued cells with the wild-type orpkTg737 gene (BAP2 cells, cilia (þ)) were provided by P. Darwin Bell., Ph.D. (Department of Nephrology of the Medical University of South Carolina) [19]. Cells were grown to confluence in Dulbecco's modified Eagle's medium/F12 medium (Mediatech, Manassas, VA, USA) with 0.2 mg/ml dexamethasone, 10 nM triiodothreonine, 1 Â insulin–transferrin–sodium selenite, 12 U/ml interferon-γ (IFN-γ), 268 mg/ml G418, 1% penicillin–streptomycin, and 5% fetal bovine serum (FBS) at 33 1C, 5% CO 2 . "
    [Show abstract] [Hide abstract]
    ABSTRACT: Traumatic brain injury (TBI) patients would benefit from the identification of reliable biomarkers to predict outcomes and treatment strategies. In our study, cerebrospinal fluid (CSF) from patients with severe TBI was evaluated for oxidant stress-mediated damage progression after hospital admission and subsequent ventriculostomy placement. Interestingly, substantial levels of peroxiredoxin VI (Prdx6), a major antioxidant enzyme normally found in astrocytes, were detected in CSF from control and TBI patients, and were not associated with blood contamination. Functionally, Prdx6 and its associated binding partner glutathione S-transferase pi (GSTP1-1, also detected in CSF) act in tandem to detoxify lipid peroxidation damage to membranes. We found Prdx6 was fully active in CSF of control patients but becomes significantly inactivated (oxidized) under TBI. Furthermore, significant and progressive oxidation of "buried" protein thiol in CSF of TBI patients (as compared to that of non-trauma control) were detected over a 24h period following hospital admission, with increased oxidation correlating with severity of trauma. Conversely, recovery of Prdx6 activity after 24h indicated more favorable patient outcome. Not only is this the first report of an extracellular form of Prdx6 but also the first report of its detection at a substantial level in CSF. Taken together, our data suggest a meaningful correlation between TBI-initiated oxidation of Prdx6, its specific phospholipid hydroperoxide peroxidase activity, and severity of trauma outcome. Consequently, we propose that Prdx6 redox status detection has the potential to be a biomarker for TBI outcome and a future indicator of therapeutic efficacy.
    Free Radical Biology and Medicine 04/2014; 72. DOI:10.1016/j.freeradbiomed.2014.04.002 · 5.74 Impact Factor
  • Source
    • "In the rat and mouse efferent ductules, AQP-1 was present not only on microvilli of non-ciliated cells but also on cilia of ciliated cells, whereas it was absent from the epididymal epithelium (Badran & Hermo, 2002; Ruz et al., 2006), similar to that seen in the hamster. Its presence on ductal cilia is not understood, but recent studies have linked the sensory function of the cilium with cell signaling pathways and the expression of AQPs in kidney tubules (Marion et al., 2011; Saigusa et al., 2012). In nonciliated cells, AQP-1 was also found along the basolateral plasmalemma (Badran & Hermo, 2002; Hermo & Smith, 2011), where Na+/K+-ATPase (Ilio & Hess, 1992) would couple its presence to allow for rapid movement of water into expanded intercellular channels along basolateral regions of the epithelium (Jones & "
    [Show abstract] [Hide abstract]
    ABSTRACT: Efferent ductules are responsible for the transportation of spermatozoa from the testis to the epididymis and their epithelium is responsible for the reabsorption of over 90% of the luminal fluid. The purpose of this research was to characterize the gross morphology and histology of efferent ductules in the male Golden Syrian hamster. The efferent ductules emerge from rete testis with a unique polarity at the apex or cephalic pole of the testis. The number of efferent ductules varied from 3 to 10 with an average of 6.0 and blind ending ducts were observed in approximately 56% of the males. The ductules merged into a single common duct prior to entering the caput epididymidis. The proximal efferent ductule lumen was wider than the distal (conus and common ducts), consistent with reabsorption of most of the luminal fluid, as was morphology of the ductal epithelium. Non-ciliated cells in the proximal region had prominent endocytic apparatuses, showing both coated pits and apical tubules in the apical cytoplasm. Large basolateral, intercellular spaces were also present in the epithelium of the proximal region. Distal non-ciliated cells had an abundance of large endosomes and lysosomal granules. Localisation of sodium/hydrogen exchanger-3 (NHE3; SLC9A3) and aquaporins 1 and 9 (AQP1, AQP9) along the microvillus border was also consistent with ion transport and fluid reabsorption by this epithelium. In comparison, the caput epididymidis epithelium expressed only AQP9 immunostaining. Another unusual feature of the hamster efferent ductules was the presence of glycogen aggregates in the basal cytoplasm of small groups of epithelial cells, but only in the proximal ducts near the rete testis. Androgen (AR), estrogen (ESR1 and ESR2) and vitamin D receptors (VDR) were also abundant in epithelial nuclei of proximal and distal efferent ductules. In comparison, caput epididymidis showed very little immunostaining for ESR1.
    Andrology 03/2014; 2(4):510-520. DOI:10.1111/j.2047-2927.2014.00194.x · 2.30 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: not applicable to EF.
    AJP Cell Physiology 03/2012; 302(10):C1434-5. DOI:10.1152/ajpcell.00070.2012 · 3.78 Impact Factor
Show more