NFAT5 contributes to osmolality-induced MCP-1 expression in mesothelial cells.

Increased expression of the C-C chemokine monocyte chemoattractant protein-1 (MCP-1) in mesothelial cells in response to high glucose concentrations and/or high osmolality plays a crucial role in the development of peritoneal fibrosis during continuous ambulatory peritoneal dialysis (CAPD). Recent studies suggest that in kidney cells osmolality-induced MCP-1 upregulation is mediated by the osmosensitive transcription factor, nuclear factor of activated T cells 5 (NFAT5). The present study addressed the question of whether activation of NFAT5 by hyperosmolality, as present in PD fluids, contributes to MCP-1 expression in the mesothelial cell line Met5A.

Cyclooxygenase-2-dependent phosphorylation of the pro-apoptotic protein Bad inhibits tonicity-induced apoptosis in renal medullary cells.

During antidiuresis, cell survival in the renal medulla requires cyclooxygenase-2 (COX-2) activity. We have recently found that prostaglandin E2 (PGE2) promotes cell survival by phosphorylation and, hence, inactivation of the pro-apoptotic protein Bad during hypertonic stress in Madin-Darby canine kidney (MDCK) cells in vitro. Here we determine the role of COX-2-derived PGE(2) on phosphorylation of Bad and medullary apoptosis in vivo using COX-2-deficient mice.

EGF receptor signaling is involved in expression of osmoprotective TonEBP target gene aldose reductase under hypertonic conditions.

Renal medullary cells adapt to their hyperosmotic environment by enhanced expression of various osmoprotective genes. Although it is clearly established that TonEBP contributes to the expression of these genes, neither the precise signaling mechanism by which hypertonicity activates TonEBP is completely understood, nor is it known whether a membrane-bound osmosenser, corresponding to yeast and bacteria, is present in mammalian cells. We found evidence that metalloproteinase (MMP)-dependent activation of the epidermal growth factor receptor (EGFR) signals to TonEBP and stimulates the expression of the TonEBP target gene aldose reductase (AR) under hypertonic conditions.

PGE2 potentiates tonicity-induced COX-2 expression in renal medullary cells in a positive feedback loop involving EP2-cAMP-PKA signaling.

Cyooxygenase-2 (COX-2)-derived PGE2 is critical for the integrity and function of renal medullary cells during antidiuresis. The present study extended our previous finding that tonicity-induced COX-2 expression is further stimulated by the major COX-2 product PGE2 and investigated the underlying signaling pathways and the functional relevance of this phenomenon. Hyperosmolality stimulated COX-2 expression and activity in Madin-Darby canine kidney (MDCK) cells, a response that was further increased by PGE2-cAMP signaling, suggesting the existence of a positive feedback loop.

Osmoadaptation of Mammalian cells - an orchestrated network of protective genes.

In mammals, the cells of the renal medulla are physiologically exposed to interstitial osmolalities several-fold higher that found in any other tissue. Nevertheless, these cells not only have the ability to survive in this harsh environment, but also to function normally, which is critical for maintenance of systemic electrolyte and fluid homeostasis. Over the last two decades, a substantial body of evidence has accumulated, indicating that sequential and well orchestrated genomic responses are required to provide tolerance to osmotic stress.

Nitric oxide decreases expression of osmoprotective genes via direct inhibition of TonEBP transcriptional activity.

During antidiuresis, renal medullary cells adapt to the hyperosmotic interstitial environment by increased expression of osmoprotective genes, which is driven by a common transcriptional activator, tonicity-responsive enhancer binding protein (TonEBP). Because nitric oxide (NO) is abundantly produced in the renal medulla, the present studies addressed the effect of NO on expression of osmoprotective genes and TonEBP activation in MDCK cells. Several structurally unrelated NO donors blunted tonicity-induced up-regulation of TonEBP target genes involved in intracellular accumulation of organic osmolytes.

Ectodomain shedding of pro-TGF-alpha is required for COX-2 induction and cell survival in renal medullary cells exposed to osmotic stress.

In the renal medulla, cyclooxygenase (COX)-2 is induced by osmotic stress as present in this kidney region during antidiuresis. Increasing evidence suggests that EGF receptor (EGFR) signaling is involved in this process. The aim of the present study was to examine the mechanisms responsible for COX-2 expression and PGE(2) production during hypertonic conditions and to identify potential autocrine/paracrine EGFR ligands.

Prostaglandin E2 stimulates expression of osmoprotective genes in MDCK cells and promotes survival under hypertonic conditions.

The cells of the renal medulla produce large amounts of prostaglandin E2 (PGE2) via cyclooxygenases (COX)-1 and -2. PGE2 is well known to play a critical role in salt and water balance and maintenance of medullary blood flow. Since renal medullary PGE2 production increases in antidiuresis, and since COX inhibition is associated with damage to the renal medulla during water deprivation, PGE2 may promote the adaptation of renal papillary cells to high interstitial solute concentrations.

Cell volume regulation in the renal papilla.

Urine osmolality and interstitial solute concentration in the renal medulla are determined by the body's hydration state. This implies that in the renal medulla extracellular solute concentrations, and hence the volume of medulla-resident cells, will vary drastically with the state of hydration. Medullary cells regulate their volume primarily by adjusting accumulation and release of low-molecular weight organic osmolytes appropriately.

Survival in hostile environments: strategies of renal medullary cells.

Cells in the renal medulla exist in a hostile milieu characterized by wide variations in extracellular solute concentrations, low oxygen tensions, and abundant reactive oxygen species. This article reviews the strategies adopted by these cells to allow them to survive and fulfill their functions under these extreme conditions.

Production of ammonium by Helicobacter pylori mediates occludin processing and disruption of tight junctions in Caco-2 cells.

Tight junctions, paracellular permeability barriers that define epithelial cell polarity, play an essential role in transepithelial transport, cell-cell adhesion and lymphocyte transmigration. They are also important for the maintenance of innate immune defence and intestinal antigen uptake. Ammonium (NH4+) is elevated in the gastric aspirates of Helicobacter pylori-infected patients and has been implicated in the disruption of tight-junction functional integrity and the induction of gastric mucosal damage during H.

Response of renal medullary cells to osmotic stress.

In antidiuresis renal medullary cells are exposed to high NaCl and urea concentrations. Long-term adaptation of renal medullary cells to high extracellular NaCl concentrations is accomplished by intracellular accumulation of organic osmolytes. The underlying mechanisms include enhanced uptake from the extracellular space (betaine, myo-inositol and amino acids), increased intracellular production [sorbitol and glycerophosphorylcholine (GPC)] and reduced intracellular degradation (GPC).

Differential expression of heat shock protein 27 and 70 in renal papillary collecting duct and interstitial cells - implications for urea resistance.

The adaptation of renal medullary cells to their hyperosmotic environment involves the accumulation of compatible organic osmolytes and the enhanced synthesis of heat shock proteins (HSP) 27 and 70. While the mechanisms leading to osmolyte accumulation are similar in papillary collecting duct (PCD) and papillary interstitial (PI) cells, the present data demonstrate that HSP27 and HSP70 are expressed differentially in these cells both in vivo and in vitro. HSP70 is abundant in PCD, but not expressed in PI cells in the papilla in situ, while HSP27 is expressed in both PCD and PI cells.

Cell survival in the hostile environment of the renal medulla.

The countercurrent system in the medulla of the mammalian kidney provides the basis for the production of urine of widely varying osmolalities, but necessarily entails extreme conditions for medullary cells, i.e., high concentrations of solutes (mainly NaCl and urea) in antidiuresis, massive changes in extracellular solute concentrations during the transitions from antidiuresis to diuresis and vice versa, and low oxygen tension.

Effect of ammonium on the expression of osmosensitive genes in Madin-Darby canine kidney cells.

The cells of the kidney medulla are exposed routinely to high extracellular concentrations of various solutes including NaCl, urea and ammonium (NH4+). Although it is well established that the expression of a variety of osmosensitive genes and proteins, which confer cytoprotection on renal medullary cells, is induced by high NaCl concentrations, the role of NH4+ in these cellular responses is unclear. This study thus addressed the effect of NH4+ on the expression of the betaine/GABA transporter (BGT-1), the sodium/myo-inositol cotransporter (SMIT), aldose reductase (AR), and heat shock protein 70 (HSP70) in Madin-Darby canine kidney (MDCK) cells, using Northern and Western blot analyses and enzyme-linked immunosorbent assay (ELISA).

Chronic COX-2 inhibition reduces medullary HSP70 expression and induces papillary apoptosis in dehydrated rats.

Background: Papillary cells adapt to their hyperosmotic environment by accumulating organic osmolytes and by enhanced synthesis of heat shock protein 70 (HSP70), which protect against high-solute concentrations. Because cyclooxygenase-2 (COX-2) is expressed abundantly in the renal papilla and is induced by dehydration, and because HSP70 expression is stimulated by specific prostaglandins, COX-2 inhibition may interfere with cellular osmoadaptation.

Methods: In vivo, rats received rofecoxib before water deprivation.

Intracellular Na concentration and Rb uptake in proximal convoluted tubule cells and abundance of Na/K-ATPase alpha1-subunit in NHE3-/- mice.

Proximal solute and fluid absorption is greatly reduced in mice in which the gene encoding the Na/H exchanger isoform 3 has been ablated (NHE3-/-). To obtain information on the intracellular functional consequences of such selective NHE3 deficiency, Na, Cl and K concentrations and cell Rb uptake were measured using electron microprobe analysis after a 30-s infusion of Rb (an index of basolateral Na/K-ATPase activity) in proximal convoluted tubule (PCT) cells of NHE3-/- and wild-type (NHE3+/+) mice. In addition, the relative abundance of the alpha1-subunit of the Na/K-ATPase in the outer cortex was determined by Western blot analysis.

Heat shock protein 72, a chaperone abundant in renal papilla, counteracts urea-mediated inhibition of enzymes.

Urea, at concentrations routinely observed in the renal inner medulla during antidiuresis in many mammals, is a potent protein destabilizing agent that reduces the activity of many enzymes. The molecular chaperone heat shock protein 72 (HSP72) is expressed at about 5 ng/ micro g protein in the renal papilla and is thus 40 times more abundant than in the isosmotic cortex and may counteract the deleterious effects of high urea concentrations in the inner medulla. To test this hypothesis, we examined the effect of recombinant HSP72 on lactate dehydrogenase activity in the presence of 0.

Regulation of TonEBP transcriptional activator in MDCK cells following changes in ambient tonicity.

In response to ambient hypertonicity, TonEBP (tonicity-responsive enhancer binding protein) stimulates certain genes including those encoding cytokines, transporters for organic solutes, and a molecular chaperone. TonEBP is regulated in a bidirectional manner, upregulated by an increase in ambient tonicity while downregulated by a decrease. To investigate the role of intracellular ionic strength in the activity of TonEBP, we subjected Madin-Darby canine kidney cells to a variety of conditions.

Relationship between intracellular ionic strength and expression of tonicity-responsive genes in rat papillary collecting duct cells.

Intracellular ionic strength may play an important role in regulating the expression of genes encoding osmolyte-accumulating molecules. To establish whether a strict relation exists between these variables, intracellular ionic strength (sum of Na+, Cl- and K+ concentrations) and the relative abundance of mRNA derived from various tonicity-sensitive genes was examined using electron microprobe analysis and Northern blots on primary cultures of rat papillary collecting duct (PCD) cells following acute or long-term alterations in medium tonicity. Hypertonic medium (450 mosmol kg(-1)) evoked an initial rise in intracellular ionic strength (269 +/- 5 vs.

Deafness and renal tubular acidosis in mice lacking the K-Cl co-transporter Kcc4.

Hearing depends on a high K(+) concentration bathing the apical membranes of sensory hair cells. K(+) that has entered hair cells through apical mechanosensitive channels is transported to the stria vascularis for re-secretion into the scala media(). K(+) probably exits outer hair cells by KCNQ4 K(+) channels(), and is then transported by means of a gap junction system connecting supporting Deiters' cells and fibrocytes() back to the stria vascularis.

Regulated overexpression of heat shock protein 72 protects Madin-Darby canine kidney cells from the detrimental effects of high urea concentrations.

Exposure of renal medullary cells to elevated extracellular NaCl concentrations is associated with increased heat shock protein 72 (HSP72) expression and improved resistance to subsequent exposure to a high urea concentration (600 mM). To establish a causal relationship between HSP72 expression and protection against high urea concentrations, HSP72 was inducibly overexpressed in Madin-Darby canine kidney (MDCK) cells, in the absence of hypertonic stress before urea exposure. For this purpose, the human stress-inducible HSP72 gene was cloned downstream from a dexamethasone (DEX)-inducible promoter in the eukaryotic expression vector pLKneo.