PPARγ Agonists: Blood Pressure and Edema

Peroxisome proliferator activated receptor γ (PPARγ) agonists are widely used in the treatment of type 2 diabetes. Side effects of drug treatment include both fluid retention and a lowering of blood pressure. Data from animal and human studies suggest that these effects arise, at least in part, from drug-induced changes in the kidney. In order to capitalize on the positive aspect (lowering of blood pressure) and exclude the negative one (fluid retention), it is necessary to understand the mechanisms of action underlying each of the effects. When interpreted with known physiological principles, current hypotheses regarding potential mechanisms produce enigmas that are difficult to resolve. This paper is a summary of the current understanding of PPARγ agonist effects on both blood pressure and fluid retention from a renal perspective and concludes with the newest studies that suggest alternative pathways within the kidney that could contribute to the observed drug-induced effects

THE ROLE OF THE PHOSPHOINOSITIDE PATHWAY IN HORMONAL REGULATION OF THE EPITHELIAL SODIUM CHANNEL

In summary, insulin and aldosterone stimulate phosphatidylinositol phosphorylation, thus indicating the existence of a regulated protein at or before the PI3-kinase step. Aldosterone induces the synthesis of sgk, a downstream element of the PI pathway. Sgk is necessary, but not rate-limiting, for aldosterone- and insulin-stimulated Na+ transport. However, the enzyme appears to be rate-limiting for the natriferic action of ADH. Insulin-stimulated Na+ transport, an acute response, is dependent on PI3-kinase activity but the magnitude of the response is not altered by a cellular excess of sgk. ADH-stimulated transport is not dependent on PI3-kinase but is potentiated by an excess of sgk. The foregoing data indicate that the PI pathway is involved in several steps of the natriferic action of hormones and intersects with other pathways which regulate ENaC. Furthermore, the data are consistent with the hypothesis that activation of PI3-kinase may ultimately stimulate channel insertion as well as regulate channel endocytosis. Both of these phenomena can result in an increase of ENaC-mediated Na+ transport

Phosphoinositide lipid second messengers: new paradigms for transepithelial signal transduction

Multiple forms of phosphatidylinositol are generated by differential phosphorylation of the inositol headgroup. These phosphoinositides, specifically PI(4,5)P2, have been implicated as modulators in a variety of transport processes. The data indicate that phosphoinositides can modulate transporters directly or via the activation of down-stream signaling components. The phosphoinositide pathway has been linked to changes in transporter kinetics, intracellular signaling, membrane targeting and membrane stability. Recent results obtained for several of the well-characterized transport systems suggest the need to reassess the role of PI(4,5)P2 and question whether lower abundance forms of the phosphoinositides, notably PI(3,4,5)P3 (PIP3) and PI(3,4)P2, are the pertinent transport regulators. In contrast to PI(4,5)P2, these latter forms represent a dynamic, regulated pool, the characteristics of which are more compatible with the nature of signaling intermediates. A recently described, novel transepithelial signaling pathway has been demonstrated for PIP3 in which a signal initiated on the basolateral membrane is transduced to the apical membrane entirely within the planar face of the inner leaflet of the plasma membrane. The new paradigms emerging from recent studies may be widely applicable to transporter regulation in other cell types and are particularly relevant for signaling in polarized cells

Modulation of basal and peptide hormone-stimulated Na transport by membrane cholesterol content in the A6 epithelial cell line

These studies examined the effect of altering plasma membrane cholesterol on basal Na+ flux as well as on the natriferic responses to the peptide hormones, insulin and anti-diuretic hormone (ADH) in the A6 model renal cell line. Membrane cholesterol concentrations were depleted or enriched using methyl-beta-cyclodextrin (MbetaCD) or a MbetaCD/cholesterol inclusion complex respectively. Effects of changes in the apical and basolateral plasma membranes were examined independently. Apical membrane cholesterol removal or supplementation had no effect on the basal Na+ transport rate. Short-term apical membrane cholesterol supplementation also had no effect on insulin-stimulated Na+ transport or on the initial phase of the ADH response. Interestingly, the additional apical membrane cholesterol had an inhibitory effect on the ADH response after 30 minutes. Apical membrane cholesterol depletion partially inhibited the responses to both insulin and ADH. Conversely, supplementation of basolateral cholesterol caused a significant increase in basal Na+ flux. Removal of cholesterol from the basolateral plasma membrane caused a decrease in basal Na+ flux with a time course analogous to channel turnover and completely inhibited peptide hormone responses. None of the changes in membrane cholesterol content decreased transcellular resistance. These results indicate an important role for membrane cholesterol content in the regulation of ENaC-mediated Na+ uptake

Acetylation of albumin by low doses of aspirin

Aspirin has a variety of pharmacologic actions, which are expressed at different doses of the drug. An effect on platelet function occurs at very low doses of aspirin (1,2). Indeed, a large number of clinical trials have been carried out to assess whether low to moderate doses of aspirin (180 to 1500 mg per day) taken prophylactically will affect the natural history of a variety of diseases in which thrombosis is thought to play a role (3)

Secretory Activity of Human Cyst Fluid Isolated From Polycystic Kidney Disease Patients

Polycystic kidney disease (PKD) is characterized by the slow growth of fluid-filled cysts in kidney tubules. Kidney function is relatively normal in the first 5 decades of life despite substantial cyst development but thereafter the decline in function is precipitous leading to complete renal failure in five years in 50% of patients. As cysts increase in size, the probability of rupture and release of cyst fluid becomes increasingly likely. Cyst fluid has been shown to cause an additional secretory Cl- flux that leads to the expansion of the remaining intact cysts. We have previously shown that the active component of the cyst fluid is lysophosphatidic acid (LPA). Electrophysiological techniques were used to measure human cyst fluid stimulation of ion flux in the mpkCCDcl4 cell line, a model of the cell type that lines renal cysts. The Cl- secretory response is due to activation of both CFTR and a calcium activated chloride channel. Interestingly, the cyst fluid effect was not accompanied by an increase in cAMP but rather via a LPA receptor mediated activation of phospholipase C followed by the stimulation of the tyrosine kinase Pyk. These results suggest novel targets for treatment of late stage PKD

Hormonal regulation of the epithelial Na+ channel: From amphibians to mammals

High-resistance epithelia derived from amphibian sources such as frog skin, toad urinary bladder, and the A6 Xenopus laevis kidney cell line have been widely used to elucidate the underlying mechanisms involved in the regulation of vectorial ion transport. More recently, the isolation of high-resistance mammalian cell lines has provided model systems in which to study differences and similarities between the regulation of ion transporter function in amphibian and mammalian renal epithelia. In the present study, we have compared the natriferic (Na+ retaining) responses to aldosterone, insulin, and vasotocin/vasopressin in the A6 and mpkCCDcl4 (mouse principal cells of the kidney cortical collecting duct) cell lines. The functional responses of the epithelial Na+ channel (ENaC) to hormonal stimulation were remarkably similar in both the amphibian and mammalian lines. In addition, insulin- and aldosterone-stimulated, reabsorptive Na+ transport in both cell lines requires the presence of functional PI3-kinase

Characterization of hormone-stimulated Na+ transport in a high-resistance clone of the MDCK cell line

The Madin-Darby canine kidney (MDCK) cell line forms an epithelial monolayer which expresses many of the morphological and functional properties of the renal collecting duct. The C7 subclone of the parent line forms an epithelium which expresses many of the characteristics of principal cells. The MDCK-C7 subclone forms a high-resistance epithelium that is capable of vectorial ion transport. We have found that this epithelium responds to aldosterone, antidiuretic hormone (ADH) and insulin like growth factor 1 (IGF1) with increases in amiloride-sensitive Na+ transport. The responses to aldosterone and ADH follow time-courses that are consistent with the action of these hormones in vivo. This is the first demonstration of IGF1-induced Na+ reabsorption in a mammalian model system. Interestingly, a maximal response to any one of these natriferic factors does not inhibit a subsequent response to another hormone. These studies indicate that the C7 subclone retains many of the natriferic responses of the native principal cells and is an ideal model for studying hormonal modulation of Na+ transport

Effects of Lysophosphatidic Acid (LPA) and Antidiuretic Hormone (ADH) on Cl- Secretory Responses in Polycystic Kidney Disease (PKD)

poster abstractPolycystic kidney disease (PKD) is a genetic disease that causes the formation of fluid-filled cysts in the kidney and other organs such as the liver and pancreas. Kidney function is seemingly unaltered despite substantial cyst development over the first four to six decades of life, but then the decline in renal function is precipitous often leading to complete renal failure in 5 years. Antidiuretic hormone (ADH) causes an increase in Cl- secretion into the cyst lumen, and one of the drugs in human clinical trials for treatment of PKD is an ADH receptor antagonist. The hormone works by stimulating cAMP production, which leads to the Cl- secretion. Interestingly, we have found that cyst fluid from human patients also causes a secretory Cl- flux that can lead to the growth of the remaining intact cysts. The active component of the cyst fluid is LPA, a phospholipid that acts as an extracellular signaling molecule. This secretion is important in late stage disease when large cysts are likely to leak or burst contributing to the rapid decline in renal function. Electrophysiological techniques were implemented to compare the ion fluxes stimulated by ADH and LPA. In the mpkCCDc14 (mouse principal cells of the cortical collecting duct clone 4) cell line we found that the Cl- secretory pathways stimulated by the two factors are separate and independent. Further indication of this separation is our finding that LPA stimulation does not increase cAMP levels. Therefore we have identified an additional target for potential pharmaceutical intervention in the treatment of PKD

Bioinformatic Analysis of Proteomic Changes That Occur in an Airway Epithelial Cell Line in Response to Exposure to Physiologically Relevant Concentrations of Carbon Nanotubes

poster abstractCarbon nanomaterials are widely produced and used in industry, medicine and scientific research. To examine the impact of acute exposure to nanoparticles on human health, the human airway epithelial cell line, Calu-3, was used to evaluate potential alterations in cellular function of airway epithelia after 24 hours exposure to different concentrations of two common carbon nanoparticles, single- and multi-wall carbon nanotubes (SWCNT, MWCNT). After exposure to the nanoparticles, label-free quantitative mass spectrometry (LFQMS) was used to study the differential protein expression in Calu-3 cells. Ingenuity Pathway Analysis (IPA) was used to conduct a bioinformatic analysis of proteins identified in LFQMS. Changes in protein abundance generated in response to 100 ng/ml exposure of both MWCNT and SWCNT suggest that cell functions of cell death and survival, cell-to-cell signaling and interaction, cellular assembly and organization, cellular growth and proliferation, infectious disease, molecular transport and protein synthesis are predicted to be effected. The majority of the protein changes represent a decrease in amount suggesting a shut down of metabolism to protect cells. The STRING database was used to analyze the protein networks in different functions. Interestingly some proteins like cadherin 1 (CDH1), signal transducer and activator of transcription 1 (STAT1), junction plakoglobin (JUP), apoptosis-associated speck-like protein containing a CARD (PYCARD), appear in several functions and tend to be in the center of the networks, which suggest they may play important roles in the cell function and activity