scholarly journals Urinary miRNA Biomarkers of Drug-Induced Kidney Injury and Their Site Specificity Within the Nephron

2020 ◽  
Author(s):  
Brian N Chorley ◽  
Heidrun Ellinger-Ziegelbauer ◽  
Michael Tackett ◽  
Frank J Simutis ◽  
Alison H Harrill ◽  
...  
Keyword(s):  
Proximal Tubule ◽  
Laser Capture Microdissection ◽  
Collecting Duct ◽  
Unmet Need ◽  
Kidney Injury ◽  
Thick Ascending Limb ◽  
Loop Of Henle ◽  
Drug Induced ◽  
Nephron Segments ◽  
Laser Capture

Abstract Drug-induced kidney injury (DIKI) is a major concern in both drug development and clinical practice. There is an unmet need for biomarkers of glomerular damage and more distal renal injury in the loop of Henle and the collecting duct (CD). A cross-laboratory program to identify and characterize urinary microRNA (miRNA) patterns reflecting tissue- or pathology-specific DIKI was conducted. The overall goal was to propose miRNA biomarker candidates for DIKI that could supplement information provided by protein kidney biomarkers in urine. Rats were treated with nephrotoxicants causing injury to distinct nephron segments: the glomerulus, proximal tubule, thick ascending limb (TAL) of the loop of Henle and CD. Meta-analysis identified miR-192-5p as a potential proximal tubule-specific urinary miRNA candidate. This result was supported by data obtained in laser capture microdissection nephron segments showing that miR-192-5p expression was enriched in the proximal tubule. Discriminative miRNAs including miR-221-3p and -222-3p were increased in urine from rats treated with TAL versus proximal tubule toxicants in accordance with their expression localization in the kidney. Urinary miR-210-3p increased up to 40-fold upon treatment with TAL toxicants and was also enriched in laser capture microdissection samples containing TAL and/or CD versus proximal tubule. miR-23a-3p was enriched in the glomerulus and was increased in urine from rats treated with doxorubicin, a glomerular toxicant, but not with toxicants affecting other nephron segments. Taken together these results suggest that urinary miRNA panels sourced from specific nephron regions may be useful to discriminate the pathology of toxicant-induced lesions in the kidney, thereby contributing to DIKI biomarker development needs for industry, clinical, and regulatory use.

scholarly journals Evidence for Basolateral P2Y6Receptors along the Rat Proximal Tubule: Functional and Molecular Characterization

2001 ◽  
Vol 12 (8) ◽  
pp. 1640-1647 ◽  
Author(s):  
MATTHEW A. BAILEY ◽  
MARTINE IMBERT-TEBOUL ◽  
CLARE TURNER ◽  
S. KAILA SRAI ◽  
GEOFFREY BURNSTOCK ◽  
...  
Keyword(s):  
Proximal Tubule ◽  
Intracellular Calcium ◽  
Calcium Concentration ◽  
Inositol Phosphates ◽  
Collecting Duct ◽  
Intracellular Calcium Concentration ◽  
Thick Ascending Limb ◽  
Nephron Segments ◽  
Receptor Mrna ◽  
Concentration Changes

Abstract. In this study, the distribution of P2Y6receptor mRNA in rat nephron segments was investigated and a functional approach was used to analyze basolateral protein expression. Reverse transcription-PCR studies revealed more intense expression of P2Y6receptor mRNA in the proximal tubule and the thick ascending limb of Henle's loop, less intense expression in the thin descending limb and the cortical and outer medullary collecting ducts, and no detectable expression in either the thin ascending limb or the inner medullary collecting duct. Dose-dependent calcium responses to basolateral administration of UDP (a selective agonist for the P2Y6receptor) were observed in the proximal tubule but not in any of the other segments studied. In the proximal tubule, intracellular calcium concentration changes induced by UDP were associated with increased production of inositol phosphates, as were those induced by ATP and norepinephrine. However, UDP-induced intracellular calcium concentration changes were different, exhibiting no plateau after the initial peak; moreover, a single stimulation with a high concentration of UDP induced full desensitization of the UDP-sensitive calcium pathway but did not alter the responsiveness of the proximal tubule to ADP (a specific P2Y1receptor agonist), ATP or norepinephrine. In summary, this report demonstrates that P2Y6receptor mRNA is expressed in most segments of the rat nephron but that basolateral expression of the protein is restricted to the proximal tubule, where the receptor is coexpressed with the P2Y1receptor. The differences in the distributions of P2Y6receptor mRNA and UDP responses may indicate the presence of luminal receptors in other nephron segments.

scholarly journals Extracellular 2′,3′-cAMP-adenosine pathway in proximal tubular, thick ascending limb, and collecting duct epithelial cells

2013 ◽  
Vol 304 (1) ◽  
pp. F49-F55 ◽  
Author(s):  
Edwin K. Jackson ◽  
Delbert G. Gillespie
Keyword(s):  
Epithelial Cells ◽  
Collecting Duct ◽  
Phosphodiesterase Inhibitor ◽  
Kidney Injury ◽  
Thick Ascending Limb ◽  
Extracellular Adenosine ◽  
Nephron Segments ◽  
Proximal Tubular Epithelial Cells ◽  
Proximal Tubular ◽  
Commercial Source

In a previous study, we demonstrated that human proximal tubular epithelial cells obtained from a commercial source metabolized extracellular 2′,3′-cAMP to 2′-AMP and 3′-AMP and extracellular 2′-AMP and 3′-AMP to adenosine (the extracellular 2′,3′-cAMP-adenosine pathway; extracellular 2′,3′-cAMP → 2′-AMP + 3′-AMP → adenosine). The purpose of this study was to investigate the metabolism of extracellular 2′,3′-cAMP in proximal tubular vs. thick ascending limb vs. collecting duct epithelial cells freshly isolated from their corresponding nephron segments obtained from rat kidneys. In epithelial cells from all three nephron segments, 1) extracellular 2′,3′-cAMP was metabolized to 2′-AMP and 3′-AMP, with 2′-AMP > 3′-AMP, 2) the metabolism of extracellular 2′,3′-cAMP to 2′-AMP and 3′-AMP was not inhibited by either 3-isobutyl-1-methylxanthine (phosphodiesterase inhibitor) or 1,3-dipropyl-8-p-sulfophenylxanthine (ecto-phosphodiesterase inhibitor), 3) extracellular 2′,3′-cAMP increased extracellular adenosine levels, 4) 3′-AMP and 2′-AMP were metabolized to adenosine with an efficiency similar to that of 5′-AMP, and 5) the metabolism of 5′-AMP, 3′-AMP, and 2′-AMP was not inhibited by α,β-methylene-adenosine-5′-diphosphate (CD73 inhibitor). These results support the conclusion that renal epithelial cells all along the nephron can metabolize extracellular 2′,3′-cAMP to 2′-AMP and 3′-AMP and can efficiently metabolize extracellular 2′-AMP and 3′-AMP to adenosine and that the metabolic enzymes involved are not the classical phosphodiesterases nor ecto-5′-nucleotidase (CD73). Because 2′,3′-cAMP is released by injury and because previous studies demonstrate that the extracellular 2′,3′-cAMP-adenosine pathway stimulates epithelial cell proliferation via adenosine A2B receptors, the present results suggest that the extracellular 2′,3′-cAMP-adenosine pathway may help restore epithelial cells along the nephron following kidney injury.

Dependency of Microdissected Nephron Segments upon Oxidative Phosphorylation and Exogenous Substrates: A Relationship between Tubular Anatomical Location in the Kidney and Metabolic Activity

1989 ◽  
Vol 77 (3) ◽  
pp. 287-295 ◽  
Author(s):  
Shozo Torikai
Keyword(s):  
Collecting Duct ◽  
Anatomical Location ◽  
Thick Ascending Limb ◽  
Tubular Cells ◽  
Renal Tubular Cells ◽  
Nephron Segments ◽  
Henle's Loop ◽  
Renal Tubular ◽  
Substrate Deprivation ◽  
Exogenous Substrates

1. In order to examine the possibility of heterogeneity in the dependence of renal tubular cells upon oxidative phosphorylation and exogenous substrates, the effects of antimycin A and substrate deprivation on adenosine 5′-triphosphate (ATP) content were examined in isolated rat nephron segments in vitro at 37°C. 2. Antimycin A (5 μmol/l) caused varying decrements in cell ATP level within 5 min in the following order: proximal tubules > cortical thick ascending limb of Henle's loop (cTAL) > cortical collecting duct (cCD) in the cortex, and thin descending limb of Henle's loop (TDL) > medullary thick ascending limb of Henle's loop (mTAL) > outer medullary collecting duct (omCD) in the inner stripe of the outer medulla. In the thick ascending limb and the collecting duct, the segments located in the cortex were more sensitive than those in the medulla. 3. Substrate deprivation for 30 min markedly decreased the cell ATP content in cortical and medullary proximal tubules and also in medullary TDL, whereas it caused only a slight decrease in cTAL and mTAL with no change in cCD and omCD. 4. Media made hypertonic by the addition of 200 mmol/l NaCl under aerobic conditions, increased the requirement for exogenous substrates in TDL and mTAL, but not in omCD. This stimulation was seen to a lesser extent in media made hypertonic by the addition of mannitol instead of NaCl. 5. Taking into consideration a knowledge of rat kidney architecture and intrarenal gradient of oxygen partial pressure, it is likely that the observed dependency upon both oxygen and exogenous substrates in the renal tubular cells reflects adaptation of such cells to their anatomical location, and to the availability of those substances in situ. Furthermore, extracellular sodium concentration and osmolarity stimulate metabolic requirements to a different extent among the nephron segments.

Intrarenal and Cellular Localization of CLC-K2 Protein in the Mouse Kidney

2001 ◽  
Vol 12 (7) ◽  
pp. 1327-1334 ◽  
Author(s):  
KATSUKI KOBAYASHI ◽  
SHINICHI UCHIDA ◽  
SHUKI MIZUTANI ◽  
SEI SASAKI ◽  
FUMIAKI MARUMO
Keyword(s):  
Cellular Localization ◽  
Collecting Duct ◽  
Type A ◽  
Mouse Kidney ◽  
Thick Ascending Limb ◽  
Intercalated Cells ◽  
Basolateral Membranes ◽  
Nephron Segments ◽  
Henle's Loop ◽  
Distal Tubules

Abstract. CLC-K2, a kidney-specific member of the CLC chloride channel family, is thought to play an important role in the transepithelial Cl- transport in the kidney. This consensus was first reached shortly after it was demonstrated that the mutations of the human CLCNKB gene resulted in Bartter's syndrome type III. To clarify the pathogenesis, the exact intrarenal and cellular localization of CLC-K2 by immunohistochemistry of the Clcnk1-/- mouse kidney were investigated by use of an anti-CLC-K antibody that recognized both CLC-K1 and CLC-K2. CLC-K2 is expressed in the thick ascending limb of Henle's loop and distal tubules, where it is localized to the basolateral membranes. The localization of CLC-K2 to these nephron segments strongly implies that CLC-K2 confers the basolateral chloride conductance in the thick ascending limb of Henle's loop and distal tubules, where Cl- is taken up by the bumetanide-sensitive Na-K-2Cl cotransporter or the thiazide-sensitive Na-Cl cotransporter at the apical membranes. CLC-K2 expression was also shown to extend into the connecting tubule in the basolateral membrane. CLC-K2 was found in basolateral membranes of the type A intercalated cells residing along the collecting duct. This localization strongly suggests that CLC-K2 confers the basolateral conductance in the type A intercalated cells where Cl- is taken up by the anion exchanger in exchange for HCO3- at the basolateral membranes. These aspects of CLC-K2 localization suggest that CLC-K2 is important in Cl- transport in the distal nephron segments.

scholarly journals More actors in ammonia absorption by the thick ascending limb

2012 ◽  
Vol 302 (3) ◽  
pp. F293-F297 ◽  
Author(s):  
Pascal Houillier ◽  
Soline Bourgeois
Keyword(s):  
Current Knowledge ◽  
Collecting Duct ◽  
Ammonia Excretion ◽  
Hydrogen Ion ◽  
Thick Ascending Limb ◽  
Loop Of Henle ◽  
Transport Mechanisms ◽  
Sodium Hydrogen ◽  
Ammonia Transport ◽  
Ammonia Absorption

This review will briefly summarize current knowledge on the basolateral ammonia transport mechanisms in the thick ascending limb (TAL) of the loop of Henle. This segment transports ammonia against a concentration gradient and is responsible for the accumulation of ammonia in the medullary interstitium, which, in turn, favors ammonia secretion across the collecting duct. Experimental data indicate that the sodium/hydrogen ion exchanger isoform 4 (NHE4; Scl9a4) is a sodium/ammonia exchanger and plays a major role in this process. Disruption of murine NHE4 leads to metabolic acidosis with inappropriate urinary ammonia excretion and decreases the ability of the TAL to absorb ammonia and to build the corticopapillary ammonia gradient. However, NHE4 does not account for the entirety of ammonia absorption by the TAL, indicating that, at least, one more transporter is involved.

scholarly journals Differential Expression Patterns of Claudins, Tight Junction Membrane Proteins, in Mouse Nephron Segments

2002 ◽  
Vol 13 (4) ◽  
pp. 875-886 ◽  
Author(s):  
Yumiko Kiuchi-Saishin ◽  
Shimpei Gotoh ◽  
Mikio Furuse ◽  
Akiko Takasuga ◽  
Yasuo Tano ◽  
...  
Keyword(s):  
Membrane Proteins ◽  
Tight Junction ◽  
Polyclonal Antibodies ◽  
Collecting Duct ◽  
Expression Patterns ◽  
Distal Tubule ◽  
Northern Blotting ◽  
Thick Ascending Limb ◽  
Specific Expression ◽  
Nephron Segments

ABSTRACT. As the first step in understanding the physiologic functions of claudins (tight junction integral membrane proteins) in nephrons, the expression of claudin-1 to -16 in mouse kidneys was examined by Northern blotting. Among these claudins, only claudin-6, -9, -13, and -14 were not detectable. Claudin-5 and -15 were detected only in endothelial cells. Polyclonal antibodies specific for claudin-7 and -12 were not available. Therefore, the distributions of claudin-1, -2, -3, -4, -8, -10, -11, and -16 in nephron segments were examined with immunofluorescence microscopy. For identification of individual segments, antibodies specific for segment markers were used. Immunofluorescence microscopic analyses of serial frozen sections of mouse kidneys with polyclonal antibodies for claudins and segment markers revealed that claudins demonstrated very complicated, segment-specific, expression patterns in nephrons, i.e., claudin-1 and -2 in Bowman’s capsule, claudin-2, -10, and -11 in the proximal tubule, claudin-2 in the thin descending limb of Henle, claudin-3, -4, and -8 in the thin ascending limb of Henle, claudin-3, -10, -11, and -16 in the thick ascending limb of Henle, claudin-3 and -8 in the distal tubule, and claudin-3, -4, and -8 in the collecting duct. These segment-specific expression patterns of claudins are discussed, with special reference to the physiologic functions of tight junctions in nephrons.

Abstract 376: The Expression of (pro)renin Receptor is Upregulated in the Kidney by High Salt Diet and No Inhibition

Hypertension ◽  
2012 ◽  
Vol 60 (suppl_1) ◽  
Author(s):  
Rong Rong ◽  
Osamu Ito ◽  
Nobuyoshi Mori ◽  
Yuma Tamura ◽  
Akihiro Sakuyama ◽  
...  
Keyword(s):  
Collecting Duct ◽  
High Salt ◽  
Kidney Cortex ◽  
Control Group ◽  
Thick Ascending Limb ◽  
High Salt Diet ◽  
Salt Diet ◽  
Protein Levels ◽  
Nephron Segments ◽  
Sd Rats

The (pro)renin receptor ((P)RR)-bound (pro)renin not only causes the generation of angiotensin II via the increased enzymatic activity, it also activates the receptor’s own intracellular signaling pathways up-regulating the expression of the profibrotic proteins. To clarify the regulation of (P)RR expression, the present study examined the effects of high salt diet and nitric oxide synthase (NOS) inhibition on the (P)RR expression in the kidney. The nephron segments were isolated from male Sprague-Dawley (SD) rats by microdissection and bulk isolation technique, and the (P)RR mRNA and protein expressions were examined by using reverse transcription polymerase chain reaction (RT-PCR) and Western blot analysis. In adiition, 5 week-old, male SD rats were randomly divided into 3 groups: a control group, a high salt diet (HS) group and a Nω-Nitro-L-arginine (L-NAME) group, and each group was treated with vehicle, high salt diet (8%, NaCl), or L-NAME (600mg/ml in drinking water), respectively. After 4 weeks, the (P)RR expression in the kidney was compared among these groups. The (P)RR mRNA was expressed in the glomerulus (Glm), the proximal convoluted and straight tubule, the cortical and medullary thick ascending limb (TAL) and collecting duct. The (P)RR protein as well as mRNA was expressed widely in the nephron segments; the preglomerular arteriole, the Glm, the proximal tubules (PT), the medullary TAL (mTAL) and inner medullary collecting duct (IMCD). Compared with the control group, the (P)RR protein levels significantly increased in the kidney cortex of both HS group and L-NAME group by 96% (p<0.01) and 506% (p<0.01) and in the inner medulla of L-NAME group by 148% (p<0.05), but did not significantly change in the outer medulla of HS group or L-NAME group. HS increased the (P)RR protein levels in the Glm and PT by 48% (p<0.05) and 39% (p<0.01), but did not affect them in other nephron segments. These results indicated that (P)RR is expressed widely in the nephron segments and that HS and NOS inhibition upregulate the (P)RR expression in the kidney, suggesting roles of (P)RR in hypertensive kidney disorder.

Cell-specific expression of amiloride-sensitive, Na(+)-conducting ion channels in the kidney

1996 ◽  
Vol 271 (4) ◽  
pp. C1303-C1315 ◽  
Author(s):  
F. Ciampolillo ◽  
D. E. McCoy ◽  
R. B. Green ◽  
K. H. Karlson ◽  
A. Dagenais ◽  
...  
Keyword(s):  
Cell Lines ◽  
Collecting Duct ◽  
Extracellular Fluid ◽  
Cation Channel ◽  
Thick Ascending Limb ◽  
Distal Nephron ◽  
Rt Pcr ◽  
Specific Expression ◽  
Nephron Segments ◽  
Medullary Collecting Duct

Amiloride-sensitive, electrogenic Na+ absorption across the distal nephron plays a vital role in regulating extracellular fluid volume and blood pressure. Recently, two amiloride-sensitive, Na(+)-conducting ion channel cDNAs were cloned. One, an epithelial Na(+)-selective channel (ENaC), is responsible for Na+ absorption throughout the distal nephron. The second, a guanosine 3',5'-cyclic monophosphate (cGMP)-inhibitable cation channel, is conductive to Na+ and Ca2+ and contributes to Na+ absorption across the inner medullary collecting duct (IMCD). As a first step toward understanding the segment-specific contributions(s) of cGMP-gated cation channels and ENaC to Na+ and Ca2+ uptake along the nephron, we used in situ reverse transcription-polymerase chain reaction (RT-PCR) hybridization, solution-phase RT-PCR, and Western blot analysis to examine the nephron and cell-specific expression of these channels in mouse kidney cell lines and/or dissected nephron segments. cGMP-gated cation channel mRNA was detected in proximal tubule, medullary thick ascending limb (mTAL), distal convoluted tubule (DCT), cortical collecting duct (CCD), outer medullary collecting duct (OMCD), and IMCD. cGMP-gated cation channel protein was detected in DCT, CCD, and IMCD cell lines. These observations suggest that hormones that modulate intracellular cGMP levels may regulate Na+, and perhaps Ca2+, uptake throughout the nephron. mRNA for alpha-mENaC, a subunit of the mouse ENaC, was detected in mTAL, DCT, CCD, OMCD, and IMCD. Coexpression of alpha-mENaC and cGMP-gated cation channel mRNAs in mTAL, DCT, CCD, OMCD, and IMCD suggests that both channels may contribute to Na+ absorption in these nephron segments.

Differential localization of two glucose transporter isoforms in rat kidney

1990 ◽  
Vol 259 (2) ◽  
pp. C286-C294 ◽  
Author(s):  
B. Thorens ◽  
H. F. Lodish ◽  
D. Brown
Keyword(s):  
Proximal Tubule ◽  
Glucose Transporter ◽  
Collecting Duct ◽  
Rat Kidney ◽  
Basolateral Membrane ◽  
Staining Intensity ◽  
Intercalated Cells ◽  
Brain Glucose ◽  
Nephron Segments ◽  
Glucose Transporter Isoforms

The localization of two glucose transporter isoforms was mapped in the rat kidney: the high-Michaelis constant (Km; 15-20 mM) low-affinity "liver" transporter and the low-Km (1-2 mM) high-affinity "erythroid/brain" transporter. Both are basolateral membrane proteins, but the liver transporter was present exclusively in the S1 part of the proximal tubule, whereas the erythroid/brain transporter was expressed at variable levels in different nephron segments. Staining intensity was low in the straight proximal tubule (S3), intermediate in the medullary thin and thick ascending limbs, and highest in connecting segments and collecting ducts. In the collecting duct, the erythroid/brain glucose transporter was expressed at the highest level in intercalated cells; less was present in principal cells. In the papilla, only intercalated cells expressed this transporter isoform. These results suggest specific involvements of each transporter isoform in transepithelial glucose reabsorption by different segments of the proximal tubule. They also indicate that while the liver glucose transporter is present in gluconeogenic cells, there is a good correlation between the level of expression of the erythroid/brain glucose transporter and the glycolytic activity of the different nephron segments.

scholarly journals Evaluation of Novel Biomarkers of Nephrotoxicity in Two Strains of Rat Treated with Cisplatin

2010 ◽  
Vol 38 (6) ◽  
pp. 943-956 ◽  
Author(s):  
Jean-Charles Gautier ◽  
Björn Riefke ◽  
Jakob Walter ◽  
Petra Kurth ◽  
Lou Mylecraine ◽  
...  
Keyword(s):  
Proximal Tubule ◽  
Collecting Duct ◽  
Kidney Injury ◽  
Renal Proximal Tubule ◽  
Tubular Injury ◽  
Sprague Dawley ◽  
Glutathione S Transferase ◽  
Proximal Tubular ◽  
Novel Biomarkers ◽  
Noninvasive Biomarkers

Cisplatin is an anticancer agent that induces renal proximal tubule lesions in many species. Studies were conducted in Sprague-Dawley and Han-Wistar rats to evaluate the utility of novel preclinical biomarkers of nephrotoxicity for renal lesions caused by this compound. Groups of 10 males of each strain were given a single intraperitoneal injection of 0.3, 1, or 3 mg/kg cisplatin and were sacrificed on days 2, 3, and 5. The novel biomarkers α-glutathione-S-transferase (α-GST) (for proximal tubular injury), μ-glutathione-S-transferase (μ-GST) (for distal tubular injury), clusterin (for general kidney injury), and renal papillary antigen-1 (RPA-1) (for collecting duct injury) were measured in urine by enzyme immunoassay. Histologically, degeneration and necrosis of the S3 segment of the renal proximal tubule were observed on day 2 (Han-Wistar) and days 3 and 5 (both strains) at 1 and 3 mg/kg. Results showed that in both strains of rats, urinary α-GST and clusterin can be detected in urine soon after injury, are more sensitive than BUN and serum creatinine, and therefore are usable as noninvasive biomarkers of proximal tubule injury. Changes in both μ-GST or RPA-1 were considered to represent secondary minor effects of proximal tubular injury on distal segments of the nephron.

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