Calcium-Mediated Mechanisms of Cystic Expansion

In this review, we will discuss several well-accepted signaling pathways toward calcium-mediated mechanisms of cystic expansion. The second messenger calcium ion has contributed to a vast diversity of signal transduction pathways. We will dissect calcium signaling as a possible mechanism that contributes to renal cyst formation. Because cytosolic calcium also regulates an array of signaling pathways, we will first discuss cilia-induced calcium fluxes, followed by Wnt signaling that has attributed to much-discussed planar cell polarity. We will then look at the relationship between cytosolic calcium and cAMP as one of the most important aspects of cyst progression. The signaling of cAMP on MAPK and mTOR will also be discussed. We infer that while cilia-induced calcium fluxes may be the initial signaling messenger for various cellular pathways, no single signaling mediator or pathway is implicated exclusively in the progression of the cystic expansion. This article is part of a Special Issue entitled: Polycystic Kidney Disease

Polycystic Diseases in Visceral Organs

Primary cilia are nonmotile, microtubule-based, antenna-like organelles projecting from the apical surface of most mammalian cells. Elegant studies have established the importance of ciliary structure and function in signal transduction and the sensory roles of cilia in maintaining healthy cellular state. In particular, dysfunctional cilia have been implicated in a large number of diseases mainly characterized by the presence of fluid-filled cysts in various organs. Aside from polycystic kidney disease (PKD), however, the roles of cilia in polycystic liver disease (PLD), polycystic pancreas disease (PPD), and polycystic ovarian syndrome (PCOS) are still very vague. In addition, although gender and sex hormones are known to regulate cyst formation, their roles in regulating physiological functions of cilia need to be further explored

Cryptorchidism and Infertility in Rats with Targeted Disruption of the Adamts16 Locus

A Disintegrin And Metalloproteinase with ThromboSpondin motifs16 (ADAMTS-16) is a member of a family of metalloproteinases. Using a novel zinc-finger nuclease based gene-edited rat model harboring a targeted mutation of the Adamts16 locus, we previously reported this gene to be linked to blood pressure regulation. Here we document our observation with this model that Adamts16 is essential for normal development of the testis. Absence of Adamts16 in the homozygous Adamts16(mutant) males resulted in cryptorchidism and male sterility. Heterozygous Adamts16(mutant) males were normal, indicating that this is a recessive trait. Testes of homozygous Adamts16(mutant) males were significantly smaller with significant histological changes associated with the lack of sperm production. Temporal histological assessments of the testis demonstrated that the seminiferous tubules did not support active spermatogenesis, but progressively lost germ cells, accumulated vacuoles and did not have any sperm. These observations, taken together with our previous report of renal abnormalities observed with the same Adamts16(mutant) rats, suggest an important mechanistic link between Adamts16 and the functioning of the male genitourinary system

Cilostazol Prevents Endothelin-Induced Smooth Muscle Constriction and Proliferation

Cilostazol is a phosphodiesterase inhibitor that has been shown to inhibit platelet activation. Endothelin is known to be the most potent endogenous growth promoting and vasoactive peptide. In patients and animal models with stroke, the level of circulating endothelin increases and complicates the recovery progress contributed by vascular constriction (an immediate pathology) and vascular proliferation (a long-term pathology). However, the effects of cilostazol on endothelin have not been explored. To demonstrate the dual-antagonizing effects of cilostazol on vasoconstriction and cell proliferation induced by endothelin, we used primary culture of mouse vascular smooth muscle cells in vitro, mouse femoral artery ex vivo, and intracranial basilar artery ex vivo. We show that the dual-inhibition effects of cilostazol are mediated by blocking endothelin-induced extracellular calcium influx. Although cilostazol does not inhibit endothelin-induced intraorganellar calcium release, blockade of extracellular calcium influx is sufficient to blunt endothelin-induced vasoconstriction. We also show that cilostazol inhibits endothelin-induced cellular proliferation by blocking extracellular calcium influx. Inhibition of cAMP-dependent protein kinase (PKA) can block anti-proliferation activity of cilostazol, confirming the downstream role of PKA in cellular proliferation. To further demonstrate the selectivity of the dual-antagonizing effects of cilostazol, we used a different phosphodiesterase inhibitor. Interestingly, sildenafil inhibits endothelin-induced vasoconstriction but not cellular proliferation in smooth muscle cells. For the first time, we show selective dual-antagonizing effects of cilostazol on endothelin. We propose that cilostazol is an excellent candidate to treat endothelin-associated diseases, such as stroke

Cilostazol Prevents Endothelin-Induced Smooth Muscle Constriction and Proliferation

Cilostazol is a phosphodiesterase inhibitor that has been shown to inhibit platelet activation. Endothelin is known to be the most potent endogenous growth promoting and vasoactive peptide. In patients and animal models with stroke, the level of circulating endothelin increases and complicates the recovery progress contributed by vascular constriction (an immediate pathology) and vascular proliferation (a long-term pathology). However, the effects of cilostazol on endothelin have not been explored. To demonstrate the dual-antagonizing effects of cilostazol on vasoconstriction and cell proliferation induced by endothelin, we used primary culture of mouse vascular smooth muscle cells in vitro, mouse femoral artery ex vivo, and intracranial basilar artery ex vivo. We show that the dual-inhibition effects of cilostazol are mediated by blocking endothelin-induced extracellular calcium influx. Although cilostazol does not inhibit endothelin-induced intraorganellar calcium release, blockade of extracellular calcium influx is sufficient to blunt endothelin-induced vasoconstriction. We also show that cilostazol inhibits endothelin-induced cellular proliferation by blocking extracellular calcium influx. Inhibition of cAMP-dependent protein kinase (PKA) can block anti-proliferation activity of cilostazol, confirming the downstream role of PKA in cellular proliferation. To further demonstrate the selectivity of the dual-antagonizing effects of cilostazol, we used a different phosphodiesterase inhibitor. Interestingly, sildenafil inhibits endothelin-induced vasoconstriction but not cellular proliferation in smooth muscle cells. For the first time, we show selective dual-antagonizing effects of cilostazol on endothelin. We propose that cilostazol is an excellent candidate to treat endothelin-associated diseases, such as stroke

Ciliotherapy: A Novel Intervention in Polycystic Kidney Disease

Background Ciliopathies are a group of diseases associated with abnormal structure or function of primary cilia. Ciliopathies include polycystic kidney disease (PKD), a pathology associated with vascular hypertension. We previously showed that cilia length regulates cilia function, and cilia function is required for nitric oxide (NO) biosynthesis in endothelial cells. Because patients with PKD show abnormal sensory cilia function, the aim of our current study was to search for a targeted therapy focused on primary cilia, which we refer to as ‘cilio-therapy’. Methods and Results In the present studies, our in vitro analyses refined fenoldopam as an equipotent and more specific dopa- minergic agonist to regulate cilia length and function

Dopamine Receptor Type 5 in the Primary Cilia Has Dual Chemo- and Mechano-Sensory Roles

Polycystic kidney disease is characterized by cardiovascular irregularities, including hypertension. Dopamine, a circulating hormone, is implicated in essential hypertension in humans and animal models. Vascular endothelial primary cilia are known to function as mechano-sensory organelles. Although both primary cilia and dopamine receptors play important roles in vascular hypertension, their relationship has never been explored. To determine the roles of the dopaminergic system and mechano-sensory cilia, we studied the effects of dopamine on ciliary length and function in wild-type and mechano-insensitive polycystic mutant cells (Pkd1−/− and Tg737orpk/orpk). We show for the first time that mouse vascular endothelia exhibit dopamine receptor-type 5 (DR5), which colocalizes to primary cilia in cultured cells and mouse arteries in vivo. DR5 activation increases cilia length in arteries and endothelial cells through cofilin and actin polymerization. DR5 activation also restores cilia function in the mutant cells. In addition, silencing DR5 completely abolishes mechano-ciliary function in WT cells. We found that DR5 plays very important roles in ciliary length and function. Furthermore, the chemo-sensory function of cilia can alter the mechano-sensory function through changes in sensitivity to fluid-shear stress. We propose that ciliary DR5 has functional chemo- and mechano-sensory roles in endothelial cells

Temporal histological changes in rat testes.

<p>Testes from n = 3 animals each from the various groups of animals were observed under (a) 10X and (b) 40X magnification post PSH staining as described under the methods section. Representative images are shown.</p

Detection of <i>Adamts16</i> mRNA within the rat testes.

<p>Presence of <i>Adamts16</i> mRNA in the testes of age-matched 30 day old animals was determined by <i>in-situ</i> hybridization as described under the methods section. <i>β-actin</i> was used as a control.</p