Biochemical characterization and osmolytes in papillary collecting ducts from pig and dog kidneys

1988 ◽  
Vol 66 (10) ◽  
pp. 1282-1290 ◽  
Author(s):  
Yvan Boulanger ◽  
Pascale Legault ◽  
Alberto Tejedor ◽  
Patrick Vinay ◽  
Yves Theriault
Keyword(s):  
Oxidative Phosphorylation ◽  
Biochemical Characterization ◽  
Collecting Duct ◽  
Proton Nuclear Magnetic Resonance ◽  
Collecting Ducts ◽  
Papillary Collecting Duct ◽  
Biochemical Measurements ◽  
Sugar Phosphates

Papillary collecting duct tubules were prepared in gram quantities from the papillae of dog and pig kidneys. Measurements of substrate and oxygen utilizations by these tubules under both aerobic and anaerobic conditions showed the potential for both glycolysis and oxidative phosphorylation. Oxygen is not necessary to maintain a normal adenosine 5′-triphosphate concentration, but oxidative phosphorylation contributes to more than 65% of the metabolism under aerobic conditions in the two species. Both phosphorus-31 and proton nuclear magnetic resonance spectra recorded from extracts of dog cortex, red medulla, and papilla showed a clear gradient from cortex to papilla for osmolytes, such as glycerophosphorylcholine, sorbitol, inositol, betaine, and sugar phosphates. Other molecules identified in the spectra included glucose, sorbitol, mannitol, lactate, glutamine, alanine, threonine, and adenosine 5′-triphosphate. Conventional biochemical measurements supported these findings. An increase in osmolality from 300 to 600 mosmol/kg H2O for 120 min did not increase the glycerophosphorylcholine and sorbitol concentrations of dog papillary collecting ducts in vitro, but a small effect of a 24-h dehydration was detected in vivo.

Tubular prostaglandin E2 production and its role in urinary hypotonicity after release of ureteral occlusion in the rat

1987 ◽  
Vol 73 (4) ◽  
pp. 395-399 ◽  
Author(s):  
Shozo Torikai
Keyword(s):  
Prostaglandin E2 ◽  
Collecting Duct ◽  
Urine Osmolality ◽  
Urine Concentration ◽  
Pge2 Production ◽  
Endogenous Prostaglandin ◽  
Collecting Ducts ◽  
Ureteral Occlusion

1. In order to explore the involvement of endogenous prostaglandin E2 (PGE2) in the urine concentration defect after ureteral occlusion, PGE2 production by isolated collecting ducts in vitro and effects of indomethacin on urine osmolality in vivo were examined. 2. Twenty-four hours ureter obstruction caused increased PGE2 production by the medullary collecting ducts, which was maintained at a high level on the day after release of obstruction (0.8 ± 0.2 pg/mm normal, 8.1 ± 0.9 pg/mm 24 h obstruction, and 6.6 ± 1.0 pg/mm post-obstruction, mean ± sem). An enhanced PGE2 production was also observed for papillary collecting duct on the day after release of 24 h ureteral occlusion (3.9 ± 0.5 pg/mm normal and 7.7 ± 1.2 pg/mm post-obstruction). 3. Administration of indomethacin to the unilateral post-obstructive rats slightly raised the urine osmolality of the post-obstructed kidney (from 339 ± 17 to 390 ± 22 mosmol/kg H2O), while it had a greater effect on the contralateral intact kidney (from 1569 ± 138 to 2567 ± 198 mosmol/kg H2O). 4. Our data may indicate that the urine concentration defect after 24 h ureteral occlusion is ascribable mainly to a mechanism other than increased endogenous PGE2.

Net acid transport by isolated perfused inner medullary collecting ducts

1990 ◽  
Vol 258 (1) ◽  
pp. F75-F84 ◽  
Author(s):  
S. M. Wall ◽  
J. M. Sands ◽  
M. F. Flessner ◽  
H. Nonoguchi ◽  
K. R. Spring ◽  
...  
Keyword(s):  
Collecting Duct ◽  
Intercalated Cells ◽  
Acid Transport ◽  
Concentration Gradients ◽  
Collecting Ducts ◽  
Acidic Form ◽  
Medullary Collecting Duct ◽  
Stop Flow

The isolated perfused tubule technique was used to study net acid transport in rat terminal inner medullary collecting duct (IMCD) segments. The stop-flow luminal pH [measured fluorometrically with the acidic form of the pH-sensitive dye 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein in the lumen] fell 0.35 units below the bath pH in tubules from control rats and 0.53 units below the bath in tubules from deoxycorticosterone-treated rats. Tubules from control rats absorbed bicarbonate and secreted ammonium against concentration gradients, although at low rates. In control rats, 10(-8) M vasopressin added to the bath increased bicarbonate absorption almost threefold. Treatment of rats in vivo with deoxycorticosterone significantly increased the rate of bicarbonate absorption in vitro. In vivo NH4Cl loading also significantly increased bicarbonate absorption. Staining microdissected tubules with acridine orange confirmed that the perfused segments lacked intercalated cells. We conclude that the terminal IMCD spontaneously acidifies the lumen despite an absence of intercalated cells. Bicarbonate absorption appears to be regulated by the same factors that affect net acidification in other collecting duct segments.

The in vivo distribution and dynamics of DNA topoisomerase II in Drosophila embryonic nuclei and chromosomes

1993 ◽  
Vol 51 ◽  
pp. 74-75
Author(s):  
Jason R. Swedlow ◽  
Neil Osheroff ◽  
Tim Karr ◽  
John W. Sedat ◽  
David A. Agard
Keyword(s):  
Topoisomerase Ii ◽  
Biochemical Characterization ◽  
Developmental Cycle ◽  
Dna Topoisomerase Ii ◽  
Chromosomal Distribution ◽  
Dna Topoisomerase ◽  
Ideal System ◽  
Dnase Treatment

DNA topoisomerase II is an ATP-dependent double-stranded DNA strand-passing enzyme that is necessary for full condensation of chromosomes and for complete segregation of sister chromatids at mitosis in vivo and in vitro. Biochemical characterization of chromosomes or nuclei after extraction with high-salt or detergents and DNAse treatment showed that topoisomerase II was a major component of this remnant, termed the chromosome scaffold. The scaffold has been hypothesized to be the structural backbone of the chromosome, so the localization of topoisomerase II to die scaffold suggested that the enzyme might play a structural role in the chromosome. However, topoisomerase II has not been studied in nuclei or chromosomes in vivo. We have monitored the chromosomal distribution of topoisomerase II in vivo during mitosis in the Drosophila embryo. This embryo forms a multi-nucleated syncytial blastoderm early in its developmental cycle. During this time, the embryonic nuclei synchronously progress through 13 mitotic cycles, so this is an ideal system to follow nuclear and chromosomal dynamics.

scholarly journals Let-7a regulates EV secretion and mitochondrial oxidative phosphorylation by targeting SNAP23 in colorectal cancer

2021 ◽  
Vol 40 (1) ◽  
Author(s):  
You Dong Liu ◽  
Xiao Peng Zhuang ◽  
Dong Lan Cai ◽  
Can Cao ◽  
Qi Sheng Gu ◽  
...  
Keyword(s):  
Oxidative Phosphorylation ◽  
Metabolic Reprogramming ◽  
Luciferase Reporter ◽  
Mitochondrial Oxidative Phosphorylation ◽  
In Vivo Assays ◽  
Reporter Assays ◽  
Extracellular Mirna

Abstract Background MicroRNAs (miRNAs) are abundant in tumor-derived extracellular vesicles (EVs) and the functions of extracellular miRNA to recipient cells have been extensively studied with tumorigenesis. However, the role of miRNA in EV secretion from cancer cells remains unknown. Methods qPCR and bioinformatics analysis were applied for determining extracellular let-7a expression from CRC patient serum and cells. Nanosight particle tracking analysis was performed for investigating the effect of let-7a on EV secretion. Luciferase reporter assays was used for identifying targeted genes synaptosome-associated protein 23 (SNAP23). In vitro and in vivo assays were used for exploring the function of let-7a/SNAP23 axis in CRC progression. Bioenergetic assays were performed for investigating the role of let-7a/SNAP23 in cellular metabolic reprogramming. Results let-7a miRNA was elevated in serum EVs from CRC patients and was enriched in CRC cell-derived EVs. We determined that let-7a could suppress EV secretion directly targeting SNAP23. In turn, SNAP23 promotes EV secretion of let-7a to downregulate the intracellular let-7a expression. In addition, we found a novel mechanism of let-7a/SNAP23 axis by regulating mitochondrial oxidative phosphorylation (OXPHOS) through Lin28a/SDHA signaling pathway. Conclusions Let-7a plays an essential role in not only inhibiting EV secretion, but also suppressing OXPHOS through SNAP23, resulting in the suppression of CRC progression, suggesting that let-7a/SNAP23 axis could provide not only effective tumor biomarkers but also novel targets for tumor therapeutic strategies.

scholarly journals Enzymatic activity in turkey, duck, quail and chicken liver microsomes against four human cytochrome P450 prototype substrates and aflatoxin B1

2011 ◽  
Vol 1 (1) ◽  
pp. 4 ◽  
Author(s):  
Hansen W. Murcia ◽  
Gonzalo J. Díaz ◽  
Sandra Milena Cepeda
Keyword(s):  
Cytochrome P450 ◽  
Aflatoxin B1 ◽  
Enzyme Activities ◽  
High Performance ◽  
Biochemical Characterization ◽  
Liver Microsomes ◽  
Cytochrome P450 Enzymes ◽  
Catalytic Rate

Cytochrome P450 enzymes (CYP) are a group of monooxygenases able to biotransform several kinds of xenobiotics including aflatoxin B1 (AFB1), a highly toxic mycotoxin. These enzymes have been widely studied in humans and others mammals, but there is not enough information in commercial poultry species about their biochemical characteristics or substrate specificity. The aim of the present study was to identify CYPs from avian liver microsomes with the use of prototype substrates specific for human CYP enzymes and AFB1. Biochemical characterization was carried out in vitro and biotransformation products were detected by high-performance liquid chromatography (HPLC). Enzymatic constants were calculated and comparisons between turkey, duck, quail and chicken activities were done. The results demonstrate the presence of four avian ortholog enzyme activities possibly related with a CYP1A1, CYP1A2, CYP2A6 (activity not previously identified) and CYP3A4 poultry orthologs, respectively. Large differences in enzyme kinetics specific for prototype substrates were found among the poultry species studied. Turkey liver microsomes had the highest affinity and catalytic rate for AFB1 whereas chicken enzymes had the lowest affinity and catalytic rate for the same substrate. Quail and duck microsomes showed intermediate values. These results correlate well with the known in vivo sensitivity for AFB1 except for the duck. A high correlation coefficient between 7-ethoxyresorufin-Odeethylase (EROD) and 7-methoxyresorufin- O-deethylase (MROD) activities was found in the four poultry species, suggesting that these two enzymatic activities might be carried out by the same enzyme. The results of the present study indicate that four prototype enzyme activities are present in poultry liver microsomes, possibly related with the presence of three CYP avian orthologs. More studies are needed in order to further characterize these enzymes.

Is the function of the renal papilla coupled exclusively to an anaerobic pattern of metabolism?

1979 ◽  
Vol 236 (5) ◽  
pp. F423-F433 ◽  
Author(s):  
J. J. Cohen
Keyword(s):  
Oxidative Metabolism ◽  
Aerobic Glycolysis ◽  
Collecting Duct ◽  
High Rate ◽  
O2 Uptake ◽  
Duct Cells ◽  
Collecting Duct Cells ◽  
Mitochondrial Oxidative Metabolism

It is widely accepted that in vivo the function of the papilla of the mammalian kidney is supported primarily by anaerobic metabolism. As a result, the major source of energy for support of function in the papilla is considered to be derived from glycolysis. This orientation originates from two concepts: 1) that in vivo the gaseous environment of the papilla has such a low PO2 that O2 availability limits O2 consumption, and 2) that papillary tissue has a high rate of glycolysis when compared with either cortical tissue or extrarenal tissues. It has also been tacitly assumed that papillary tissue has a "low" O2 uptake. Review of the measurements of PO2 of papillary tissue and of urine PO2 indicates that the PO2 of papillary tissue should not limit its aerobic mitochondrial oxidative metabolism. While the rate of aerobic glycolysis in papillary tissue is high, simultaneously papillary tissue has a rate of O2 uptake similar to that of liver and higher than that of muscle. The major (two-thirds) source of energy for papillary tissue in vitro is from O2 uptake. That papillary tissue is not exclusively dependent on glucose for its energy requirements is indicated by the greater stimulation of papillary tissue QO2 by succinate than by glucose. Thus, papillary tissue has both a high aerobic mitochondrial oxidative metabolism and a high aerobic glycolytic metabolism. It is suggested that the mechanism for the high rate of aerobic glycolysis in the presence of an adequate O2 supply is due to the relatively small mass of mitochondria in papillary tissue in relation to the amount of work done by the tissue. As a result of the limited rate of ATP production by the mitochondrial electron transport chain, the phosphorylation state ([ATP]/[ADP][Pi]) is reduced and the cytoplasmic redox state ([NAD+]/[NADH]) of the papillary collecting duct cells also becomes more reduced; changes in both ratios enhance the rate of glycolysis. This limited metabolic capacity of the collecting duct cells may permit an excess volume of solute and water to be excreted during volume expansion diuresis. The metabolic characteristics of the papilla, when compared to cortex, also provide a basis for the observed differences in substrate selectivity of cortex and medulla with respect to utilization of glucose and lactate. The experimental approaches that may provide information bearing on the suggested mechanisms for regulation of papillary metabolism in relation to tubular work functions are indicated.

scholarly journals IL-32 is a metabolic regulator promoting survival and proliferation of malignant plasma cells

2021 ◽  
Author(s):  
Kristin Roseth Aass ◽  
Robin Mjelle ◽  
Martin H. Kastnes ◽  
Synne S. Tryggestad ◽  
Luca M. van den Brink ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Oxidative Phosphorylation ◽  
Plasma Cells ◽  
Inflammatory Diseases ◽  
Mitochondrial Respiratory Chain ◽  
Gene Signature ◽  
Growth And Survival ◽  
Novel Concept

AbstractIL-32 is a non-classical cytokine expressed in cancers, inflammatory diseases and infections. IL-32 can have both extracellular and intracellular functions, and its receptor is not identified. We here demonstrate that endogenously expressed, intracellular IL-32 binds to components of the mitochondrial respiratory chain and promotes oxidative phosphorylation. Knocking out IL-32 in malignant plasma cells significantly reduced survival and proliferation in vitro and in vivo. High throughput transcriptomic and MS-metabolomic profiling of IL-32 KO cells revealed that loss of IL-32 leads to profound perturbations in metabolic pathways, with accumulation of lipids, pyruvate precursors and citrate, indicative of reduced mitochondrial function. IL-32 is expressed in a subgroup of multiple myeloma patients with an inferior prognosis. Primary myeloma cells expressing IL-32 were characterized by a plasma cell gene signature associated with immune activation, proliferation and oxidative phosphorylation. We propose a novel concept for regulation of metabolism by an intracellular cytokine and identify IL-32 as an endogenous growth and survival factor for malignant plasma cells. IL-32 is a potential prognostic biomarker and a treatment target in multiple myeloma.

scholarly journals Collecting Duct Is a Site of Sodium Retention in PAN Nephrosis: A Rationale for Amiloride Therapy

2001 ◽  
Vol 12 (3) ◽  
pp. 598-601 ◽  
Author(s):  
GEORGES DESCHÊNES ◽  
MONIKA WITTNER ◽  
ANTONIO DI STEFANO ◽  
SYLVIE JOUNIER ◽  
ALAIN DOUCET
Keyword(s):  
Collecting Duct ◽  
Urinary Sodium Excretion ◽  
Sodium Balance ◽  
Sodium Reabsorption ◽  
Puromycin Aminonucleoside ◽  
Aldosterone Receptor ◽  
Collecting Tubules ◽  
A Site

Abstract. Micropuncture studies of the distal nephron and measurements of Na,K-ATPase activity in microdissected collecting tubules have suggested that renal retention of sodium in puromycin aminonucleoside (PAN) nephrotic rats originates in the collecting duct. The present study demonstrated this hypothesis by in vitro microperfusion and showed that amiloride was able to restore sodium balance. Indeed, isolated perfused cortical collecting ducts from PAN-treated rats exhibited an abnormally high transepithelial sodium reabsorption that was abolished by amiloride, and in vivo administration of amiloride fully prevented decreased urinary sodium excretion and positive sodium balance in nephrotic rats. As expected from the aldosterone independence of Na+ retention in PAN nephrotic rats, blockade of aldosterone receptor by potassium canrenoate did not alter urinary Na+ excretion, Na+ balance, or ascites formation in PAN nephrotic rats.

scholarly journals Cell-specific qRT-PCR of renal epithelial cells reveals a novel innate immune signature in murine collecting duct

2018 ◽  
Vol 315 (4) ◽  
pp. F812-F823 ◽  
Author(s):  
Vijay Saxena ◽  
David S. Hains ◽  
John Ketz ◽  
Melinda Chanley ◽  
John D. Spencer ◽  
...  
Keyword(s):  
Urinary Tract ◽  
Mrna Expression ◽  
Collecting Duct ◽  
Immune Defense ◽  
Innate Immune ◽  
Targeted Analysis ◽  
Collecting Tubule ◽  
Tubule Cells

The urinary tract is usually culture negative despite its close proximity to microbial flora. The precise mechanism by which the kidneys and urinary tract defends against infection is not well understood. The initial kidney cells to encounter ascending pathogens are the collecting tubule cells that consist of principal cells (PCs) that express aquaporin 2 (AQP2) and intercalated cells (ICs) that express vacuolar H+-ATPase (V-ATPase, B1 subunit). We have previously shown that ICs are involved with the human renal innate immune defense. Here we generated two reporter mice, VATPase B1-cre+tdT+mice to fluorescently label ICs and AQP2-cre+tdT+mice to fluorescently label PCs, and then performed flow sorting to enrich PCs and ICs for analysis. Isolated ICs and PCs along with proximal tubular cells were used to measure antimicrobial peptide (AMP) mRNA expression. ICs and PCs were significantly enriched for AMPs. Isolated ICs responded to uropathogenic Escherichia coli (UPEC) challenge in vitro and had higher RNase4 gene expression than control while both ICs and PCs responded to UPEC challenge in vivo by upregulating Defb1 mRNA expression. To our knowledge, this is the first report of isolating murine collecting tubule cells and performing targeted analysis for multiple classes of AMPs.

scholarly journals 3D kidney organoids for bench-to-bedside translation

2020 ◽  
Author(s):  
Navin Gupta✉ ◽  
Emre Dilmen ◽  
Ryuji Morizane
Keyword(s):  
Collecting Duct ◽  
Developmental Toxicity ◽  
Human Kidney ◽  
Great Promise ◽  
Duct Systems ◽  
Recent Developments ◽  
Induced Pluripotent ◽  
Kidney Organoids

Abstract The kidneys are essential organs that filter the blood, removing urinary waste while maintaining fluid and electrolyte homeostasis. Current conventional research models such as static cell cultures and animal models are insufficient to grasp the complex human in vivo situation or lack translational value. To accelerate kidney research, novel research tools are required. Recent developments have allowed the directed differentiation of induced pluripotent stem cells to generate kidney organoids. Kidney organoids resemble the human kidney in vitro and can be applied in regenerative medicine and as developmental, toxicity, and disease models. Although current studies have shown great promise, challenges remain including the immaturity, limited reproducibility, and lack of perfusable vascular and collecting duct systems. This review gives an overview of our current understanding of nephrogenesis that enabled the generation of kidney organoids. Next, the potential applications of kidney organoids are discussed followed by future perspectives. This review proposes that advancement in kidney organoid research will be facilitated through our increasing knowledge on nephrogenesis and combining promising techniques such as organ-on-a-chip models.

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