Cortisol alters carbonic anhydrase-mediated renal sulfate secretion

Active transepithelial sulfate secretion rate by winter flounder renal proximal tubule epithelium in primary culture (fPTC) is dependent on intracellular carbonic anhydrase (CA) and enhanced by cortisol. To further evaluate this relationship, a partial cDNA clone (327 bp) of carbonic anhydrase II (CAII) with high sequence similarity to CAII from numerous species including fish, chicken, and human was obtained from fPTCs. The majority of CA activity and CAII protein was present in the cytosol of fPTCs; however, significant amounts of both (in addition to SDS-resistant CA activity, i.e., CAIV-like isoform) were present in concentrated plasma membranes. CAII from concentrated membranes migrated differently than purified CAII on nondenaturing PAGE gels, suggesting that CAII associates with another membrane component. Treatment of fPTCs with the cell-soluble CA inhibitor methazolamide (100 μM) caused a 58% reduction in active transepithelial SO42- secretion. fPTCs that were previously cultured under high-cortisol concentrations, when subjected to 5 days of low physiological levels of cortisol, had decreased CA activity (28%), CAII protein abundance (65%), and net active SO42- secretion (28%), with no effect on epithelial differentiation. Methazolamide and low-cortisol treatment in combination inhibited net active SO42- secretion 56%, which was not different than the effect of methazolamide treatment alone. These data indicate that cortisol directly increases renal CA activity, CAII protein abundance, and CA-dependent SO42- secretion in the marine teleost renal proximal tubule.

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Stimulation of renal sulfate secretion by metabolic acidosis requires Na+/H+ exchange induction and carbonic anhydrase

AJP Renal Physiology ◽

10.1152/ajprenal.00468.2004 ◽

2005 ◽

Vol 289 (1) ◽

pp. F208-F216 ◽

Cited By ~ 6

Author(s):

Ryan M. Pelis ◽

Susan L. Edwards ◽

Stan C. Kunigelis ◽

James B. Claiborne ◽

J. Larry Renfro

Keyword(s):

Metabolic Acidosis ◽

Carbonic Anhydrase ◽

Proximal Tubule ◽

Brush Border ◽

Primary Cultures ◽

Acute Effect ◽

Renal Proximal Tubule ◽

Ussing Chambers ◽

Renal Tubular ◽

Nhe Isoforms

The acute effect of metabolic acidosis on SO42− secretion by the marine teleost renal proximal tubule was examined. Metabolic acidosis was mimicked in primary cultures of winter flounder renal proximal tubule epithelium (fPTCs) mounted in Ussing chambers by reducing interstitial pH to 7.1 (normally 7.7). fPTCs with metabolic acidosis secreted SO42− at a net rate that was 40% higher than in paired isohydric controls (pH 7.7 on interstitium). The stimulation was completely blocked by the carbonic anhydrase inhibitor methazolamide (100 μM). Although Na+/H+ exchange (NHE) isoforms 1, 2, and 3 were identified in fPTCs by immunoblotting, administering EIPA (20 μM) to the interstitial and luminal bath solutions had no effect on net SO42− secretion by fPTCs with a normal interstitial pH of 7.7. However, EIPA (20 μM) blocked most of the stimulation caused by acidosis when applied to the lumen but not interstitium, demonstrating that induction of brush-border NHE activity is important. In the intact flounder, serum pH dropped 0.4 pH units (pH 7.7 to 7.3, at 2–3 h) when environmental pH was lowered from 7.8 to ∼4.3. Whereas serum [SO42−] was not altered by acidosis, renal tubular SO42− secretion rate was elevated 200%. Thus metabolic acidosis strongly stimulates renal sulfate excretion most likely by a direct effect on active renal proximal tubule SO42− secretion. This stimulation appears to be dependent on inducible brush-border NHE activity.

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A novel murine protein with no effect on iron homoeostasis is homologous with transferrin and is the putative inhibitor of carbonic anhydrase

Biochemical Journal ◽

10.1042/bj20070384 ◽

2007 ◽

Vol 406 (1) ◽

pp. 85-95 ◽

Cited By ~ 9

Author(s):

Fudi Wang ◽

Adam P. Lothrop ◽

Nicholas G. James ◽

Tanya A. M. Griffiths ◽

Lisa A. Lambert ◽

...

Keyword(s):

Carbonic Anhydrase ◽

Gene Product ◽

Sequence Similarity ◽

Expression System ◽

Duplication Event ◽

Coding Region ◽

High Sequence Similarity ◽

Bhk Cells ◽

Major Function ◽

Yeast Expression System

In a search for genes that modify iron homoeostasis, a gene (1300017J02Rik) was located immediately upstream of the murine TF (transferrin) gene. However, expression of the 1300017J02Rik gene product was not responsive to a number of modulators of iron metabolism. Specifically, expression was not altered in mouse models of iron disorders including mice with deficiencies in the haemochromatosis protein Hfe, the recombination-activating protein, Rag, β2-microglobulin, TF, ceruloplasmin or Hb, or in mice with microcytic anaemia. Additionally, neither lipopolysaccharide nor hypoxia treatment resulted in any significant changes in the 1300017J02Rik expression level. The genomic DNA sequence suggested that the 1300017J02Rik gene product might be a protein equivalent to the pICA {porcine ICA [inhibitor of CA (carbonic anhydrase)]}. The coding region for the murine 1300017J02Rik gene was placed into the pNUT expression vector. Transformed BHK cells (baby-hamster kidney cells) were transfected with this plasmid, resulting in secretion of recombinant mICA (murine ICA) into the tissue culture medium. Following purification to homogeneity, the yield of mICA from the BHK cells was found to be considerably greater (at least 4-fold) than the yield of pICA from a previously reported Pichia pastoris (yeast) expression system. MS showed that the recombinant mICA was a glycoprotein that associated with CA in a 1:1 stoichiometry. Despite its high sequence similarity to TF, titration experiments showed that mICA was unable to bind iron specifically. Although enzymatic assays revealed that mICA was able to inhibit CA, it is unclear if this is its sole or even its major function since, to date, humans and other primates appear to lack functional ICA. Lastly, we note that this member of the TF superfamily is a relatively recent addition resulting from a tandem duplication event.

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Renal Na/H exchanger NHE-3 and Na-PO4 cotransporter NaPi-2 protein expression in glucocorticoid excess and deficient states.

Journal of the American Society of Nephrology ◽

10.1681/asn.v991560 ◽

1998 ◽

Vol 9 (9) ◽

pp. 1560-1567

Author(s):

J Loffing ◽

M Lötscher ◽

B Kaissling ◽

J Biber ◽

H Murer ◽

...

Keyword(s):

Protein Expression ◽

Proximal Tubule ◽

Apical Membrane ◽

Renal Proximal Tubule ◽

Current Data ◽

Bilateral Adrenalectomy ◽

Thick Ascending Limb ◽

Protein Abundance ◽

Intrarenal Distribution

Administration of pharmacologic doses of glucocorticoid in vivo increases renal proximal tubule apical membrane Na/H exchange and decreases Na/PO4 cotransport activity (1). Current data suggest that the NHE-3 and NaPi-2 proteins mediate significant fractions of proximal tubule apical membrane Na/H exchange and Na/PO4 cotransport, respectively. This study examines whether glucocorticoid excess or deficiency affects NHE-3 and NaPi-2 protein abundance and the intrarenal distribution of these transporters. Protein abundance of NHE-3 and NaPi-2 in control rats was compared to rats rendered glucocorticoid-deficient by bilateral adrenalectomy, and to rats receiving pharmacologic doses of dexamethasone using immunoblots and immunohistochemistry. Adrenalectomy had modest effects on NHE-3 protein abundance, but dexamethasone administration to either adrenalectomized or sham-operated rats significantly increased NHE-3 protein abundance in both the proximal tubule and thick ascending limb, but not the thin descending limb. Adrenalectomy increased NaPi-2 protein abundance in the proximal tubule, whereas dexamethasone administration dramatically suppressed NaPi-2 protein on the apical membrane in both adrenalectomized and sham-operated animals. No significant reciprocal increase in subapical NaPi-2 staining was seen in the dexamethasone-treated rats. The present study shows that glucocorticoids regulate proximal tubule apical membrane Na/H exchange and NaPi cotransport by changes in protein abundance of NHE-3 and NaPi-2, respectively.

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Acetazolamide causes renal HCO3− wasting but inhibits ammoniagenesis and prevents the correction of metabolic acidosis by the kidney

AJP Renal Physiology ◽

10.1152/ajprenal.00501.2019 ◽

2020 ◽

Vol 319 (3) ◽

pp. F366-F379

Author(s):

Perwez Alam ◽

Sihame Amlal ◽

Charuhas V. Thakar ◽

Hassane Amlal

Keyword(s):

Metabolic Acidosis ◽

Carbonic Anhydrase ◽

Proximal Tubule ◽

Glutamate Dehydrogenase ◽

Ammonia Synthesis ◽

Similar Degree ◽

Protein Abundance ◽

Text Generation ◽

Glutamine Transporter ◽

Renal Adaptation

Carbonic anhydrase (CAII) binds to the electrogenic basolateral Na+-[Formula: see text] cotransporter (NBCe1) and facilitates [Formula: see text] reabsorption across the proximal tubule. However, whether the inhibition of CAII with acetazolamide (ACTZ) alters NBCe1 activity and interferes with the ammoniagenesis pathway remains elusive. To address this issue, we compared the renal adaptation of rats treated with ACTZ to NH4Cl loading for up to 2 wk. The results indicated that ACTZ-treated rats exhibited a sustained metabolic acidosis for up to 2 wk, whereas in NH4Cl-loaded rats, metabolic acidosis was corrected within 2 wk of treatment. [Formula: see text] excretion increased by 10-fold in NH4Cl-loaded rats but only slightly (1.7-fold) in ACTZ-treated rats during the first week despite a similar degree of acidosis. Immunoblot experiments showed that the protein abundance of glutaminase (4-fold), glutamate dehydrogenase (6-fold), and SN1 (8-fold) increased significantly in NH4Cl-loaded rats but remained unchanged in ACTZ-treated rats. Na+/H+ exchanger 3 and NBCe1 proteins were upregulated in response to NH4Cl loading but not ACTZ treatment and were rather sharply downregulated after 2 wk of ACTZ treatment. ACTZ causes renal [Formula: see text] wasting and induces metabolic acidosis but inhibits the upregulation of glutamine transporter and ammoniagenic enzymes and thus suppresses ammonia synthesis and secretion in the proximal tubule, which prevented the correction of acidosis. This effect is likely mediated through the inhibition of the CA-NBCe1 metabolon complex, which results in cell alkalinization. During chronic ACTZ treatment, the downregulation of both NBCe1 and Na+/H+ exchanger 3, along with the inhibition of ammoniagenesis and [Formula: see text] generation, contributes to the maintenance of metabolic acidosis.

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Immunologic characterization of plasma membranes from the renal proximal tubule of the dog

Kidney International ◽

10.1038/ki.1977.52 ◽

1977 ◽

Vol 11 (5) ◽

pp. 348-356 ◽

Cited By ~ 15

Author(s):

Sue Chant ◽

Melvin Silverman

Keyword(s):

Proximal Tubule ◽

Plasma Membranes ◽

Renal Proximal Tubule

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The Fine Structure of Rat Renal Microbodies and Cytoplasmic Bodies Following Perfusion Fixation

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100073544 ◽

1973 ◽

Vol 31 ◽

pp. 620-621

Author(s):

J. M. Barrett ◽

P. M. Heidger

Keyword(s):

Fine Structure ◽

Proximal Tubule ◽

Rat Kidney ◽

Renal Proximal Tubule ◽

Convincing Evidence ◽

Cytoplasmic Bodies ◽

Rat Renal Proximal Tubule ◽

Renal Proximal Tubule Cells ◽

Perfusion Fixation

Microbodies have received extensive morphological and cytochemical investigation since they were first described by Rhodin in 1954. To our knowledge, however, all investigations of microbodies and cytoplasmic bodies of rat renal proximal tubule cells have employed immersion fixation. Tisher, et al. have shown convincing evidence of fine structural alteration of microbodies in rhesus monkey kidney following immersion fixation; these alterations were not encountered when in vivo intravascular perfusion was employed. In view of these studies, and the fact that techniques for perfusion fixation have been established specifically for the rat kidney by Maunsbach, it seemed desirable to employ perfusion fixation to study the fine structure and distribution of microbodies and cytoplasmic bodies within the rat renal proximal tubule.

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