Bend or break: The primary cilium as a potential regulator of electrolyte reabsorption in the kidney

Tubular cell dedifferentiation and peritubular inflammation are coupled by the transcription regulator Id1 in renal fibrogenesis

Super-Resolution Microscopy and FIB-SEM Imaging Reveal Parental Centriole-Derived, Hybrid Cilium in Mammalian Multiciliated Cells.

The lymphotoxin β receptor is a potential therapeutic target in renal inflammation.

a human kidney contains approximately one million functional units (the nephrons) that consist of a filter (the glomerulus) and a processing portion (the proximal, intermediate, and distal tubule). The glomeruli produce ∼180 liters of primary filtrate every day of which only 1 to 2 liters are

Loss of primary cilia is frequently observed in tumor cells, including pancreatic ductal adenocarcinoma (PDAC) cells, suggesting that the absence of this organelle may promote tumorigenesis through aberrant signal transduction and the inability to exit the cell cycle. However, the molecular mechanisms that explain how PDAC cells lose primary cilia are still ambiguous. In this study, we found that inhibition or silencing of histone deacetylase 2 (HDAC2) restores primary cilia formation in PDAC cells. Inactivation of HDAC2 results in decreased Aurora A expression, which promotes disassembly of primary cilia. We further showed that HDAC2 controls ciliogenesis independently of Kras, which facilitates Aurora A expression. These studies suggest that HDAC2 is a novel regulator of primary cilium formation in PDAC cells.

The Emerging Complexity of the Vertebrate Cilium: New Functional Roles for an Ancient Organelle

The concept of fluid flow-induced nucleotide release was first suggested from research in vascular biology. Fluid flow elicits regulatory input signals in vascular endothelial cells. In response to shear stress, these cells release vasoactive autacoids (e.g. nitric oxide) (Moncada and Higgs, 2006). Mechanical perturbations are known to release nucleotides such as ATP and UTP from most cells, including endothelial and epithelial cells (Bodin et al. 1991; Lazarowski et al. 2003). In endothelia, extracellular ATP triggers nitric oxide release. More recently, the concept of endothelial shear stress-induced vasodilatation has been suggested to involve ATP release and endothelial purinergic receptors (P2X4 isoform) (Yamamoto et al. 2006). Endothelial and epithelial cells abundantly express purinergic receptors (P2). These receptors are localized to both the apical (lumen-facing) and basolateral sides of the epithelial barrier where they modulate transepithelial salt and water transport (Leipziger, 2003). The rate of fluid flow over the apical surface of epithelia varies widely under different physiological conditions (e.g. diuresis and antidiuresis in the renal tubule). The question is how epithelial cells coordinate ion transport and fluid flow. Many researchers have become fascinated by the possible role of the primary cilium as a cellular sensing organelle in epithelia (Praetorius & Spring, 2005). In the absence of fluid flow, the primary cilium protrudes into the epithelial lumen perpendicular to the cell membrane. Fluid flow triggers cilium bending, eliciting an increase of cytosolic Ca2+ ([Ca2+]i) (Praetorius & Spring, 2001, 2003). Thus, this organelle appears to have the capacity to be a sensory antenna to detect flow changes. The flow-induced [Ca2+]i increase, a hallmark of this novel epithelial function, is found in many epithelial tissues. The nature of this flow-stimulated [Ca2+]i increase is not currently fully understood. It was hypothesized to relate to influx of Ca2+ via the polycystin1/2 complex localized on the primary cilium (Nauli et al. 2003; Xu et al. 2007). Recently it was found that the flow-induced increase of [Ca2+]i in isolated perfused renal tubules is caused or augmented by a flow or stretch-induced release of nucleotides from the epithelial cells and paracrine activation of epithelial P2 receptors (Jensen et al. 2007). It is very likely that this process requires mechano-sensing via the primary cilium, but this remains to be proven. In bile duct epithelia, extracellular nucleotides provide autocrine/paracrine stimuli for anion secretion (Feranchak & Fitz, 2002). Like in renal epithelia, native perfused bile ducts were recently shown to respond with an increase of [Ca2+]i after increasing ductal flow in a cilium-dependent manner (Masyuk et al. 2006). In this issue of The Journal of Physiology, Woo et al. (2008) present direct evidence that fluid flow over the apical surface of cultured rat and human bile duct epithelial cells triggers shear-stress-dependent ATP release to the apical side of the tissue. At the same time, fluid flow also triggers an increase of [Ca2+]i and hence, the activation of Cl− secretion. These data highlight that ATP is released from the epithelial cells, themselves, and that fluid flow, itself, stimulates epithelial ATP secretion. Based on these data, Woo et al. (2008) propose an integrative cellular model of bile secretion (see Fig. 10 of their paper). In this model, augmented hepatic bile secretion carries a signal via luminal bile flow to the target site of the bile duct cell. This cell uses its property to sense flow and triggers the release of nucleotides, which activate luminal (and probably also basolateral) P2 receptors in the duct cell leading to an increase of [Ca2+]i. Elevated [Ca2+]i in turn activates ductal anion secretion, which is a major component of bile formation. As Woo et al. (2008) mostly use non-ciliated human bile duct cells their results argue that this mechanism does not require the primary cilium. Interestingly, in ciliated rat bile duct cells the flow-induced ATP secretion response curve is shifted to the left (Woo et al. 2008), suggesting that primary cilia accentuate the detection of flow changes by epithelia. Finally, the data of Woo et al. (2008) suggest that the activation of the atypical PKCζ is involved in flow-stimulated epithelial ATP secretion. Epithelial P2 receptors modulate ion transport in a dual fashion. In secretory epithelia (e.g. the bile duct), P2 receptors activate ion secretion, whereas in absorptive epithelia (e.g. the renal tubule), they inhibit absorption. Thus, the overall functional consequence of epithelial P2 receptor stimulation is to promote an increased amount of fluid in the tubular compartment. The flow-induced secretion of ATP now found in renal and bile duct epithelia has a fascinating general perspective: flow changes per se, via local puringeric signalling, regulate the amount of ion and water transport in many epithelia. This novel concept of flow-sensing and ion transport regulation implies that local physical conditions define the final quantity and composition of the transported fluids.

Signal transduction and activator of transcription 3 (STAT3) is a key transcription factor implicated in the pathogenesis of kidney fibrosis. Although Stat3 deletion in tubular epithelial cells is known to protect mice from fibrosis, vFoxd1 cells remains unclear. Using Foxd1-mediated Stat3 knockout mice, CRISPR, and inhibitors of STAT3, we investigate its function. STAT3 is phosphorylated in tubular epithelial cells in acute kidney injury, whereas it is expanded to interstitial cells in fibrosis in mice and humans. Foxd1-mediated deletion of Stat3 protects mice from folic-acid- and aristolochic-acid-induced kidney fibrosis. Mechanistically, STAT3 upregulates the inflammation and differentiates pericytes into myofibroblasts. STAT3 activation increases migration and profibrotic signaling in genome-edited, pericyte-like cells. Conversely, blocking Stat3 inhibits detachment, migration, and profibrotic signaling. Furthermore, STAT3 binds to the Collagen1a1 promoter in mouse kidneys and cells. Together, our study identifies a previously unknown function of STAT3 that promotes kidney fibrosis and has therapeutic value in fibrosis.

Most renal tubular epithelial cells possess primary cilia that protrude into the tubular lumen, where they are exposed to urine flow. The cilia of the epithelia curve with the urine flow within the renal tubules. The primary cilium of renal epithelium has been assumed to be a mechano-sensor of the urine flow. However, the mechanical characteristics of the cilium of an each cell are not well assessed. In this study, we measured Young's modulus of each primary cilium by using optical tweezers. First, we developed a system to manipulate each primary cilium of living tubular epithelial cells in situ by using optical tweezers. We adopted flexible-substratum technique to visualize individual micron-sized primary cilia of the Madin-Darby canine kidney (MDCK) cell lines. Through the use of the flexible-substratum technique, the primary cilia of MDCK cells could be clearly imaged from the side. We showed that we could apply minute force to polystyrene microspheres stuck on the tip of the cilia by using optical tweezers. The Young's modulus of each primary cilium was successfully examined by measuring the displacement of the microsphere. We concluded that this methodology enables us to manipulate each primary cilium minutely and to estimate the elasticity of an individual primary cilium.

Primary cilia are sensory organelles that extend from the cell surface and sense extracellular signals. Endothelial primary cilia protruding from the inner surface of blood vessel walls sense changes in blood flow and convert this mechanosensation into an intracellular biochemical/molecular signal, which triggers a cellular response. Primary endothelial cilia dysfunction may contribute to the impairment of this response and thus be directly implicated in the development of vascular abnormalities such as hypertension and aneurysms. Using both in vitro techniques as well as in vivo animal models, we and others have investigated fluid flow mechanosensory functions of endothelial cilia in cultured cells, animal models and autosomal dominant polycystic kidney disease (ADPKD) patients. More in-depth studies directed at identification of the mechanisms of fluid flow sensing will further enhance our knowledge of cilia-dependent vascular pathology. Although the current treatments aimed at treating the cardiovascular symptoms in ADPKD patients successfully slowed the progression of cyst growth, there is growing evidence which suggests that drugs which interfere with primary cilia function or structure could reduce cardiovascular complications in ADPKD. This review is to summarize the most recent studies on primary endothelial cilia function in the vascular system and to present primary cilia as a novel therapeutic target for vascular hypertension.

Fabry disease is an X-linked recessive lysosomal storage disease that is caused by deficient activity of the lysosomal enzyme α-galactosidase A (α-Gal A), an enzyme that cleaves terminal α-galactosyl residues. This deficiency results in progressive lysosomal accumulation of glycosphingolipid with

ERK-mediated suppression of cilia in cisplatin-induced tubular cell apoptosis and acute kidney injury

Tubular epithelial cells in the kidney are continuously exposed to urinary fluid shear stress (FSS) generated by urine movement and recent in vitro studies suggest that changes of FSS could contribute to kidney injury. However it is unclear whether FSS alters the epithelial characteristics of the renal tubule. Here, we evaluated in vitro and in vivo the influence of FSS on epithelial characteristics of renal proximal tubular cells taking the organization of junctional complexes and the presence of the primary cilium as markers of epithelial phenotype. Human tubular cells (HK-2) were subjected to FSS (0.5 Pa) for 48h. Control cells were maintained under static conditions. Markers of tight junctions (Claudin-2, ZO-1), Par polarity complex (Pard6), adherens junctions (E-Cadherin, β-Catenin) and the primary cilium (α-acetylated Tubulin) were analysed by quantitative PCR, Western blot or immunocytochemistry. In response to FSS, Claudin-2 disappeared and ZO-1 displayed punctuated and discontinuous staining in the plasma membrane. Expression of Pard6 was also decreased. Moreover, E-Cadherin abundance was decreased, while its major repressors Snail1 and Snail2 were overexpressed, and β-Catenin staining was disrupted along the cell periphery. Finally, FSS subjected-cells exhibited disappeared primary cilium. Results were confirmed in vivo in a uninephrectomy (8 months) mouse model where increased FSS induced by adaptive hyperfiltration in remnant kidney was accompanied by both decreased epithelial gene expression including ZO-1, E-cadherin and β-Catenin and disappearance of tubular cilia. In conclusion, these results show that proximal tubular cells lose an important number of their epithelial characteristics after long term exposure to FSS both in vitro and in vivo. Thus, the changes in urinary FSS associated with nephropathies should be considered as potential insults for tubular cells leading to disorganization of the tubular epithelium.

The calcium-activated chloride channel Anoctamin 1 contributes to the regulation of renal function

Many cilium types have at their proximal base a particulated membrane structure, the so-called ciliary necklace. Necklaces of primary and secondary cilia of olfactory receptor cells and ciliated respiratory cells, and of primary cilia of olfactory supporting cells were studied as a function of embryonic age. Strand numbers in necklaces of primary cilia of these cell types do not differ, but they differ significantly from those of necklaces of secondary cilia. Primary cilia have 2 to 4, but most commonly 3, necklace strands. This is true for necklaces of primary cilia of 8 different nasal cell types: olfactory epithelial basal and glandular cells, vomeronasal receptor and supporting cells, and microvillous respiratory epithelial cells, in addition to the 3 cell types mentioned above. Comparison with other systems suggests that primary cilia resemble flagella of eukaryotic flagellates and spermatozoa of some invertebrates with respect to their number of necklace strands. Average numbers of necklace strands in secondary olfactory cilia increase from 3-4 at the 16th and 17th gestational days to 6-7 in adults. Those in secondary respiratory cilia increase from 2-3 at the 18th and 19th gestational days to 5-6 in adults. Longer cilia have more strands than shorter ones. Necklaces often have free strand endings, also in primary cilia, suggesting that they spiral. Comparing the present data with those in the literature suggests that necklace features occurring during reciliation differ from those of de novo ciliogenesis. Primary and secondary cilia share the following qualities: 1) Membrane regions above necklace strands can differ quite drastically from those below the strands.(ABSTRACT TRUNCATED AT 250 WORDS)