Acute phosphate depletion inhibits the Na+/H+ antiporter in a cultured renal cell line

1993 ◽  
Vol 265 (3) ◽  
pp. F440-F448
Author(s):  
J. Green ◽  
O. Foellmer ◽  
C. R. Kleeman ◽  
M. M. Basic
Keyword(s):  
Cell Line ◽  
Time Course ◽  
Maximal Activity ◽  
Cell Recovery ◽  
Opossum Kidney ◽  
Opossum Kidney Cell ◽  
Proximal Tubular ◽  
Pi Deficit

We studied the effect of acute Pi depletion on the regulation of intracellular pH (pHi) in the OK opossum kidney cell line by using the pH-sensitive dye 2'7'-bis(carboxyethyl)-5(6)-carboxyfluorescein (BCECF). Cell recovery from an NH4Cl acid load in HCO3-free buffer disclosed an Na(+)-dependent component blocked by amiloride and a smaller Na(+)-independent component that increased on exposure of the cells to a high-K+ buffer. After 24-h incubation of the cells in phosphate-free medium, pHi recovery by the Na+/H+ exchanger was markedly inhibited, whereas the Na(+)-independent pHi recovery was not affected. The inhibition of Na+/H+ exchange was reversible on correction of cellular Pi deficit. A similar phenomenon was observed when cellular Pi depletion was induced by acute exposure (min) to fructose. Pi depletion shifted the pHi dependence of the exchanger and also reduced its maximal activity. Time-course studies revealed that the effect of Pi depletion could not be attributed to attenuation of Na(+)-K(+)-adenosinetriphosphatase activity and resultant diminution of the transmembrane gradient for the Na+ influx. We conclude that acute Pi depletion in cultured proximal tubular cells leads to reversible inhibition of the Na+/H+ exchanger. This in vitro finding may relate to the in vivo observation of impaired HCO3 reabsorption and bicarbonaturia in acute Pi depletion.

Autocrine/paracrine regulation of renal Na(+)-phosphate cotransport by dopamine

1993 ◽  
Vol 264 (4) ◽  
pp. F618-F622 ◽  
Author(s):  
R. P. Glahn ◽  
M. J. Onsgard ◽  
G. M. Tyce ◽  
S. L. Chinnow ◽  
F. G. Knox ◽  
...  
Keyword(s):  
In Vivo Model ◽  
Acid Decarboxylase ◽  
System Support ◽  
Opossum Kidney ◽  
Amino Acid Decarboxylase ◽  
Ok Cells ◽  
Proximal Tubular ◽  
Phosphate Cotransport

We tested the hypothesis that dopamine (DA) acts as an autocrine/paracrine regulator of Na(+)-Pi symport in proximal tubules, using opossum kidney (OK) cells as an in vivo model. Both DA and parathyroid hormone (PTH) increased adenosine 3',5'-cyclic monophosphate (cAMP) and inhibited Na(+)-gradient-dependent uptake of 32P but not that of L-[3H]-alanine. Incubation of OK cells with L-dopa, a DA precursor, resulted in accumulation of DA (7.4 nM), a ninefold increase of cAMP in the medium, and an inhibition (-10%) of Na(+)-Pi uptake. Carbidopa, an inhibitor of aromatic-L-amino acid decarboxylase, prevented the formation of DA from L-dopa, the increase in cAMP, and the inhibition of Na(+)-Pi cotransport. Pi-replete OK cells produced more DA (+15%) from L-dopa than Pi-deprived cells; however, the endogenous DA inhibited Na(+)-Pi cotransport both in Pi-deprived and in Pi-replete cells. Thus OK cells can synthesize DA from L-dopa in a quantity sufficient to elicit both the maximum DA-stimulated cAMP accumulation and inhibition of Na(+)-Pi cotransport in the same cell population. Our data, obtained on an in vitro system, support the hypothesis proposing that DA generated in proximal tubular cells can modulate, via cAMP, the Na(+)-Pi symport in the same or adjacent cells. If present in the kidney, this pathway might represent an autocrine/paracrine system that can contribute to regulation of renal Pi homeostasis.

scholarly journals Molecular signature of anastasis for reversal of apoptosis

F1000Research ◽  
2017 ◽  
Vol 6 ◽  
pp. 43 ◽  
Author(s):  
Ho Man Tang ◽  
C. Conover Talbot Jr ◽  
Ming Chiu Fung ◽  
Ho Lam Tang
Keyword(s):  
Gene Expression ◽  
Time Course ◽  
Molecular Mechanisms ◽  
Expression Profiles ◽  
Cancer Cell Line ◽  
Molecular Signature ◽  
Cell Recovery ◽  
Therapeutic Implications

Anastasis (Greek for "rising to life") is a cell recovery phenomenon that rescues dying cells from the brink of cell death. We recently discovered anastasis to occur after the execution-stage of apoptosis in vitro and in vivo. Promoting anastasis could in principle preserve injured cells that are difficult to replace, such as cardiomyocytes and neurons. Conversely, arresting anastasis in dying cancer cells after cancer therapies could improve treatment efficacy. To develop new therapies that promote or inhibit anastasis, it is essential to identify the key regulators and mediators of anastasis – the therapeutic targets. Therefore, we performed time-course microarray analysis to explore the molecular mechanisms of anastasis during reversal of ethanol-induced apoptosis in mouse primary liver cells. We found striking changes in transcription of genes involved in multiple pathways, including early activation of pro-cell survival, anti-oxidation, cell cycle arrest, histone modification, DNA-damage and stress-inducible responses, and at delayed times, angiogenesis and cell migration. Validation with RT-PCR confirmed similar changes in the human liver cancer cell line, HepG2, during anastasis. Here, we present the time-course whole-genome gene expression dataset revealing gene expression profiles during the reversal of apoptosis. This dataset provides important insights into the physiological, pathological, and therapeutic implications of anastasis.

Internalization and catabolism of insulin by an established renal cell line

1988 ◽  
Vol 254 (6) ◽  
pp. C822-C828 ◽  
Author(s):  
C. Yagil ◽  
B. H. Frank ◽  
R. Rabkin
Keyword(s):  
Cell Line ◽  
Degradation Products ◽  
Renal Tubules ◽  
Insulin Metabolism ◽  
Opossum Kidney ◽  
Opossum Kidney Cell ◽  
Proximal Tubular ◽  
Specific Receptors ◽  
Membrane Bound ◽  
Renal Cell Line

Proximal renal tubules are a key site of insulin metabolism. To explore the kinetics and metabolic requirements of insulin internalization and catabolism, we used the opossum kidney cell line, which has proximal tubular-like features and possesses insulin-specific receptors. Internalization was determined by separating membrane-bound insulin from intracellular insulin by exposure to an acidified medium. Internalization of membrane-bound insulin was rapid, and half-maximal internalization occurred within 2.5 min. Degradation products did not accumulate in the cell but appeared in the medium after a delay of 5 min from the onset of internalization. In other experiments, addition of KCN (2 mM) or omission of glucose did not alter degradation, but KCN, combined with the omission of glucose, inhibited degradation by 64%. This was associated with a 240% increase in membrane-bound insulin and an 81% decrease in intracellular insulin. Accordingly, it appears that under these circumstances impaired degradation was a consequence of impaired internalization. In contrast, although 0.1 mM chloroquine, an endosomal-lysosomal inhibitor, also depressed degradation (by 57%), intracellular insulin increased fourfold, indicating failure of intracellular processing. We conclude that these cultured kidney cells rapidly internalize and degrade insulin and that internalization, a prerequisite for degradation, is dependent on energy that can be derived from anaerobic glycolysis or oxidative metabolism. Furthermore, the intracellular degradative processing of insulin by these cells involves a chloroquine-sensitive pathway.

CD36 mediates proximal tubular binding and uptake of albumin and is upregulated in proteinuric nephropathies

2012 ◽  
Vol 303 (7) ◽  
pp. F1006-F1014 ◽  
Author(s):  
Richard J. Baines ◽  
Ravinder S. Chana ◽  
Matthew Hall ◽  
Maria Febbraio ◽  
David Kennedy ◽  
...  
Keyword(s):  
Proximal Tubule ◽  
Renal Biopsy ◽  
Renal Diseases ◽  
Creatinine Ratio ◽  
Proximal Tubular Cells ◽  
Opossum Kidney ◽  
Tubular Cells ◽  
Proximal Tubular

Dysregulation of renal tubular protein handling in proteinuria contributes to the development of chronic kidney disease. We investigated the role of CD36 as a novel candidate mediator of albumin binding and endocytosis in the kidney proximal tubule using both in vitro and in vivo approaches, and in nephrotic patient renal biopsy samples. In CD36-transfected opossum kidney proximal tubular cells, both binding and uptake of albumin were substantially enhanced. A specific CD36 inhibitor abrogated this effect, but receptor-associated protein, which blocks megalin-mediated endocytosis of albumin, did not. Mouse proximal tubular cells expressed CD36 and this was absent in CD36 null animals, whereas expression of megalin was equal in these animals. Compared with wild-type mice, CD36 null mice demonstrated a significantly increased urinary protein-to-creatinine ratio and albumin-to-creatinine ratio. Proximal tubular cells expressed increased CD36 when exposed to elevated albumin concentrations in culture medium. Expression of CD36 was studied in renal biopsy tissue obtained from adult patients with heavy proteinuria due to minimal change disease, membranous nephropathy, or focal segmental glomerulosclerosis. Proximal tubular CD36 expression was markedly increased in proteinuric individuals. We conclude that CD36 is a novel mediator influencing binding and uptake of albumin in the proximal tubule that is upregulated in proteinuric renal diseases. CD36 may represent a potential therapeutic target in proteinuric nephropathy.

scholarly journals Molecular signature of anastasis for reversal of apoptosis

F1000Research ◽  
2017 ◽  
Vol 6 ◽  
pp. 43 ◽  
Author(s):  
Ho Man Tang ◽  
C. Conover Talbot Jr ◽  
Ming Chiu Fung ◽  
Ho Lam Tang
Keyword(s):  
Gene Expression ◽  
Cell Death ◽  
Time Course ◽  
Molecular Mechanisms ◽  
Expression Profiles ◽  
Molecular Signature ◽  
Cell Recovery ◽  
Therapeutic Implications

Apoptosis is a type of programmed cell death that is essential for normal organismal development and homeostasis of multicellular organisms by eliminating unwanted, injured, or dangerous cells. This cell suicide process is generally assumed to be irreversible. However, accumulating studies suggest that dying cells can recover from the brink of cell death. We recently discovered an unexpected reversibility of the execution-stage of apoptosis in vitro and in vivo, and proposed the term anastasis (Greek for “rising to life”) to describe this cell recovery phenomenon. Promoting anastasis could in principle preserve injured cells that are difficult to replace, such as cardiomyocytes and neurons. Conversely, arresting anastasis in dying cancer cells after cancer therapies could improve treatment efficacy. To develop new therapies that promote or inhibit anastasis, it is essential to identify the key regulators and mediators of anastasis – the therapeutic targets. Therefore, we performed time-course microarray analysis to explore the molecular mechanisms of anastasis during reversal of ethanol-induced apoptosis in mouse primary liver cells. We found striking changes in transcription of genes involved in multiple pathways, including early activation of pro-survival genes, cell cycle arrest, stress-inducible responses, and at delayed times, cell migration and angiogenesis. Here, we present the time-course whole-genome gene expression dataset revealing gene expression profiles during the reversal of apoptosis. This dataset provides important insights into the physiological, pathological, and therapeutic implications of anastasis.

Comparison of High Purity Factor IX Concentrates and a Prothrombin Complex Concentrate in a Canine Model of Thrombogenicity

1991 ◽  
Vol 66 (05) ◽  
pp. 609-613 ◽  
Author(s):  
I R MacGregor ◽  
J M Ferguson ◽  
L F McLaughlin ◽  
T Burnouf ◽  
C V Prowse
Keyword(s):  
High Purity ◽  
Time Course ◽  
Prothrombin Complex Concentrate ◽  
Degradation Products ◽  
Canine Model ◽  
Factor Ix ◽  
In Vitro Tests ◽  
Albumin Infusion

SummaryA non-stasis canine model of thrombogenicity has been used to evaluate batches of high purity factor IX concentrates from 4 manufacturers and a conventional prothrombin complex concentrate (PCC). Platelets, activated partial thromboplastin time (APTT), fibrinogen, fibrin(ogen) degradation products and fibrinopeptide A (FPA) were monitored before and after infusion of concentrate. Changes in FPA were found to be the most sensitive and reproducible indicator of thrombogenicity after infusion of batches of the PCC at doses of between 60 and 180 IU/kg, with a dose related delayed increase in FPA occurring. Total FPA generated after 100-120 IU/kg of 3 batches of PCC over the 3 h time course was 9-12 times that generated after albumin infusion. In contrast the amounts of FPA generated after 200 IU/kg of the 4 high purity factor IX products were in all cases similar to albumin infusion. It was noted that some batches of high purity concentrates had short NAPTTs indicating that current in vitro tests for potential thrombogenicity may be misleading in predicting the effects of these concentrates in vivo.

A Novel Triazole Derivative Drug Presenting In Vitro and In Vivo Anticancer Properties

2018 ◽  
Vol 18 (17) ◽  
pp. 1483-1493
Author(s):  
Ricardo Imbroisi Filho ◽  
Daniel T.G. Gonzaga ◽  
Thainá M. Demaria ◽  
João G.B. Leandro ◽  
Dora C.S. Costa ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cancer Cells ◽  
Cell Line ◽  
Cancer Cell ◽  
Cancer Cell Line ◽  
Hepatic Function ◽  
Anticancer Properties ◽  
Mcf 7

Background: Cancer is a major cause of death worldwide, despite many different drugs available to treat the disease. This high mortality rate is largely due to the complexity of the disease, which results from several genetic and epigenetic changes. Therefore, researchers are constantly searching for novel drugs that can target different and multiple aspects of cancer. Experimental: After a screening, we selected one novel molecule, out of ninety-four triazole derivatives, that strongly affects the viability and proliferation of the human breast cancer cell line MCF-7, with minimal effects on non-cancer cells. The drug, named DAN94, induced a dose-dependent decrease in MCF-7 cells viability, with an IC50 of 3.2 ± 0.2 µM. Additionally, DAN94 interfered with mitochondria metabolism promoting reactive oxygen species production, triggering apoptosis and arresting the cancer cells on G1/G0 phase of cell cycle, inhibiting cell proliferation. These effects are not observed when the drug was tested in the non-cancer cell line MCF10A. Using a mouse model with xenograft tumor implants, the drug preventing tumor growth presented no toxicity for the animal and without altering biochemical markers of hepatic function. Results and Conclusion: The novel drug DAN94 is selective for cancer cells, targeting the mitochondrial metabolism, which culminates in the cancer cell death. In the end, DAN94 has been shown to be a promising drug for controlling breast cancer with minimal undesirable effects.

Studies on the mechanism of acute inhibition of thyroglobulin hydrolysis by iodine

1985 ◽  
Vol 108 (4) ◽  
pp. 511-517 ◽  
Author(s):  
Nandalal Bagchi ◽  
Birdie Shivers ◽  
Thomas R. Brown
Keyword(s):  
Time Course ◽  
Period 2 ◽  
Iodotyrosine Deiodinase ◽  
Rate Of Hydrolysis ◽  
Underlying Mechanisms ◽  
Excess Iodide ◽  
Sites Of Action ◽  
Hydrolysis Of

Abstract. Iodine in excess is known to acutely inhibit thyroidal secretion. In the present study we have characterized the time course of the iodine effect in vitro and investigated the underlying mechanisms. Labelled thyroid glands were cultured in vitro in medium containing mononitrotyrosine, an inhibitor of iodotyrosine deiodinase. The rate of hydrolysis of labelled thyroglobulin was measured as the proportion of labelled iodotyrosines and iodothyronines recovered at the end of culture and was used as an index of thyroidal secretion. Thyrotrophin (TSH) administered in vivo acutely stimulated the rate of thyroglobulin hydrolysis. Addition of Nal to the culture medium acutely inhibited both basal and TSH-stimulated thyroglobulin hydrolysis. The effect of iodide was demonstrable after 2 h, maximal after 6 h and was not reversible upon removal of iodide. Iodide abolished the dibutyryl cAMP induced stimulation of thyroglobulin hydrolysis. Iodide required organic binding of iodine for its effect but new protein or RNA synthesis was not necessary. The inhibitory effects of iodide and lysosomotrophic agents such as NH4C1 and chloroquin on thyroglobulin hydrolysis were additive suggesting different sites of action. Iodide added in vitro altered the distribution of label in prelabelled thyroglobulin in a way that suggested increased coupling in the thyroglobulin molecule. These data indicate that 1) the iodide effect occurs progressively over a 6 h period, 2) continued presence of iodide is not necessary once the inhibition is established, 3) iodide exerts its action primarily at a post cAMP, prelysosomal site and 4) the effect requires organic binding of iodine, but not new RNA or protein synthesis. Our data are consistent with the hypothesis that excess iodide acutely inhibits thyroglobulin hydrolysis by increasing the resistance of thyroglobulin to proteolytic degradation through increased iodination and coupling.

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