Proximal Tubule p53 in Cold Storage/Transplantation-Associated Kidney Injury and Renal Graft Dysfunction

Kidney injury associated with cold storage/transplantation is a primary factor for delayed graft function and poor outcome of renal transplants. p53 contributes to both ischemic and nephrotoxic kidney injury, but its involvement in kidney cold storage/transplantation is unclear. Here, we report that p53 in kidney proximal tubules plays a critical role in cold storage/transplantation kidney injury and inhibition of p53 can effectively improve the histology and function of transplanted kidneys. In a mouse kidney cold storage/transplantation model, we detected p53 accumulation in proximal tubules in a cold storage time-dependent manner, which correlated with tubular injury and cell death. Pifithrin-α, a pharmacologic p53 inhibitor, could reduce acute tubular injury, apoptosis and inflammation at 24 h after cold storage/transplantation. Similar effects were shown by the ablation of p53 from proximal tubule cells. Notably, pifithrin-α also ameliorated kidney injury and improved the function of transplanted kidneys in 6 days when it became the sole life-supporting kidney in recipient mice. in vitro, cold storage followed by rewarming induced cell death in cultured proximal tubule cells, which was accompanied by p53 activation and suppressed by pifithrin-α and dominant-negative p53. Together, these results support a pathogenic role of p53 in cold storage/transplantation kidney injury and demonstrate the therapeutic potential of p53 inhibitors.

Download Full-text

Immunolocalization of NHE8 in rat kidney

AJP Renal Physiology ◽

10.1152/ajprenal.00229.2004 ◽

2005 ◽

Vol 288 (3) ◽

pp. F530-F538 ◽

Cited By ~ 65

Author(s):

Sunita Goyal ◽

SueAnn Mentone ◽

Peter S. Aronson

Keyword(s):

Proximal Tubule ◽

Brush Border ◽

Brush Border Membrane ◽

Rat Kidney ◽

Surface Membrane ◽

Proximal Tubules ◽

Intense Staining ◽

Proximal Tubule Cells ◽

Nephron Segment ◽

Tubule Cells

In situ hybridization studies demonstrated that Na+/H+ exchanger NHE8 is expressed in kidney proximal tubules. Although membrane fractionation studies suggested apical brush-border localization, precise membrane localization could not be definitively established. The goal of the present study was to develop isoform-specific NHE8 antibodies as a tool to directly establish the localization of NHE8 protein in the kidney by immunocytochemistry. Toward this goal, two sets of antibodies that label different NHE8 epitopes were developed. Monoclonal antibody 7A11 and polyclonal antibody Rab65 both specifically labeled NHE8 by Western blotting as well as by immunofluorescence microscopy. The immunolocalization pattern in the kidney seen with both antibodies was the same, thereby validating NHE8 specificity. In particular, NHE8 expression was observed on the apical brush-border membrane of all proximal tubules from S1 to S3. The most intense staining was evident in proximal tubules in the deeper cortex and medulla with a significant but somewhat weaker staining in superficial proximal tubules. Colocalization studies with γ-glutamyltranspeptidase and megalin indicated expression of NHE8 on both the microvillar surface membrane and the coated-pit region of proximal tubule cells, suggesting that NHE8 may be subject to endocytic retrieval and recycling. Although colocalizing in the proximal tubule with NHE3, no significant alteration in NHE8 protein expression was evident in NHE3-null mice. We conclude that NHE8 is expressed on the apical brush-border membrane of proximal tubule cells, where it may play a role in mediating or regulating ion transport in this nephron segment.

Download Full-text

Abstract P169: Antenatal Betamethasone Attenuates the Angiotensin-(1-7)/Nitric Oxide Axis in Adult Male but not Female Renal Proximal Tubule Cells

Hypertension ◽

10.1161/hyp.66.suppl_1.p169 ◽

2015 ◽

Vol 66 (suppl_1) ◽

Author(s):

Yixin Su ◽

Victor M Pulgar ◽

Jianli Bi ◽

Mark Chappell ◽

James C Rose

Keyword(s):

Nitric Oxide ◽

Proximal Tubule ◽

Adult Male ◽

Fetal Programming ◽

Cell Model ◽

Specific Effect ◽

Proximal Tubules ◽

Proximal Tubule Cells ◽

Tubule Cells ◽

Na Uptake

Our studies have revealed a sex-specific effect of fetal programming on sodium (Na+) excretion in adult sheep whereby the males exhibit reduced Na+ excretion and an attenuated natriuretic response to Ang-(1-7) as compared to the females. We hypothesize that the renal proximal tubules are a key target for the early programming effects of glucocorticoids exposure to regulate Na+ handling in the adult males. Therefore, we isolated and cultured cortical proximal tubule cells (RPTC) from adult male and female sheep antenatally exposed to betamethasone (Beta) or vehicle. Na+ uptake and nitric oxide (NO) were assessed with Sodium Green and DAF fluorescence prior to and following a low dose of Ang-(1-7) (1x10-11 M) in isolated RPTC from sheep at ~1.5 years of age. Data are expressed as % of basal uptake or area under the curve (AUC) for Na+ or % of control for NO. Male Beta RPTC exhibit greater Na+ uptake than male vehicle cells (427±32%, n=13, vs. 315±28%, n=14, p<0.05; however, Beta had no effect on Na+ uptake in the female cells (242±18%, n=9, vs. 250±15%, n=10, p>0.05). Ang-(1-7) inhibited Na+ uptake in RPTC from vehicle male (255±40%) and from both vehicle (191±14%) and Beta (209±11%) females (Figure 1B), but failed to attenuate Na+ uptake in Beta male cells (Figure 1A). Beta exposure also abolished NO stimulation by Ang-(1-7) in male but not female RPTC (Figure 1C). We conclude that an Ang-(1-7)-NO-dependent pathway contributes to the sex-dependent consequences of programming on Na+ regulation in the proximal tubules of the kidney. Moreover, the RPTC retain both the sex and Beta-induced phenotype of the adult and may reflect an appropriate cell model of fetal programming.

Download Full-text

Toxicoproteomic Analysis of Poly(ADP-Ribose)-Associated Proteins Induced by Oxidative Stress in Human Proximal Tubule Cells

Toxicological Sciences ◽

10.1093/toxsci/kfz131 ◽

2019 ◽

Vol 171 (1) ◽

pp. 117-131

Author(s):

Argel Islas-Robles ◽

Deepthi Yedlapudi ◽

Serrine S Lau ◽

Terrence J Monks

Keyword(s):

Oxidative Stress ◽

Cell Death ◽

Proximal Tubule ◽

Calcium Influx ◽

Regulation Of Transcription ◽

Loss Of Function ◽

Proximal Tubule Cells ◽

Associated Proteins ◽

Tubule Cells ◽

Parp 1

Abstract 2,3,5-Tris-(glutathion-S-yl)hydroquinone (TGHQ) is a nephrotoxic and nephrocarcinogenic metabolite of hydroquinone. TGHQ generates reactive oxygen species (ROS), causing DNA-strand breaks, hyperactivation of PARP-1, increases in intracellular calcium ([Ca2+]i), and cell death. PARP-1 catalyzes the attachment of ADP-ribose polymers (PAR) to target proteins. In human kidney proximal tubule cells, ROS-mediated PARP-1 hyperactivation and elevations in [Ca2+]i are reciprocally coupled. The molecular mechanism of this interaction is unclear. The aim of the present study was to identify ROS-induced PAR-associated proteins to further understand their potential role in cell death. PAR-associated proteins were enriched by immunoprecipitation, identified by LC-MS/MS, and relative abundance was obtained by spectral counting. A total of 356 proteins were PAR-modified following TGHQ treatment. A total of 13 proteins exhibited gene ontology annotations related to calcium. Among these proteins, the general transcription factor II-I (TFII-I) is directly involved in the modulation of [Ca2+]i. TFII-I binding to phospholipase C (PLC) leads to calcium influx via the TRPC3 channel. However, inhibition of TRPC3 or PLC had no effect on TGHQ-mediated cell death, suggesting that their loss of function may be necessary but insufficient to cause cell death. Nevertheless, TGHQ promoted a time-dependent translocation of TFII-I from the nucleus to the cytosol concomitant with a decrease in tyrosine phosphorylation in α/β-TFII-I. Therefore it is likely that ROS have an important impact on the function of TFII-I, such as regulation of transcription, and DNA translesion synthesis. Our data also shed light on PAR-mediated signaling during oxidative stress, and contributes to the development of strategies to prevent PAR-dependent cell death.

Download Full-text

Acute Kidney Injury in SARS-CoV-2 Infection: Direct Effect of Virus on Kidney Proximal Tubule Cells

International Journal of Molecular Sciences ◽

10.3390/ijms21093275 ◽

2020 ◽

Vol 21 (9) ◽

pp. 3275 ◽

Cited By ~ 10

Author(s):

Manoocher Soleimani

Keyword(s):

Acute Kidney Injury ◽

Severe Acute Respiratory Syndrome ◽

Proximal Tubule ◽

Rna Viruses ◽

Kidney Injury ◽

The Novel ◽

Proximal Tubule Cells ◽

Severe Respiratory Infection ◽

Tubule Cells ◽

Novel Coronavirus

Coronaviruses (CoVs), including Severe Acute Respiratory Syndrome (SARS), Middle East Respiratory Syndrome (MERS), and the novel coronavirus disease-2 (SARS-CoV-2) are a group of enveloped RNA viruses that cause a severe respiratory infection which is associated with a high mortality [...]

Download Full-text

Renal organic anion transporter 1 is maldistributed and diminishes in proximal tubule cells but increases in vasculature after ischemia and reperfusion

AJP Renal Physiology ◽

10.1152/ajprenal.90409.2008 ◽

2008 ◽

Vol 295 (6) ◽

pp. F1807-F1816 ◽

Cited By ~ 10

Author(s):

Osun Kwon ◽

Wei-Wei Wang ◽

Shane Miller

Keyword(s):

Proximal Tubule ◽

Ischemia Reperfusion ◽

Organic Anion ◽

Basolateral Membrane ◽

Kidney Injury ◽

Organic Anion Transporter ◽

Membrane Domain ◽

Proximal Tubule Cells ◽

Anion Transporter ◽

Tubule Cells

Renal solute clearances are reduced in ischemic acute kidney injury. However, the mechanisms explaining how solute clearance is impaired have not been clarified. Recently, we reported that cadaveric renal allografts exhibit maldistribution of organic anion transporter 1 (OAT1) in proximal tubule cells after ischemia and reperfusion, resulting in impairment of PAH clearance. In the present study, we characterized renal OAT1 in detail after ischemia-reperfusion using a rat model. We analyzed renal OAT1 using confocal microscopy with a three-dimensional reconstruction of serial optical images, Western blot, and quantitative real-time RT-PCR. OAT1 was distributed to basolateral membranes of proximal tubule cells in controls. With ischemia, OAT1 decreased in basolateral membrane, especially in the lateral membrane domain, and appeared diffusely in cytoplasm. After reperfusion following 60-min ischemia, OAT1 often formed cytoplasmic aggregates. The staining for OAT1 started reappearing in lateral membrane domain 1 h after reperfusion. The basolateral membrane staining was relatively well discernable at 240 h of reperfusion. Of note, a distinct increase in OAT1 expression was noted in vasculature early after ischemia and after reperfusion. The total amount of OAT1 protein expression in the kidney diminished after ischemia-reperfusion in a duration-dependent manner until 72 h, when they began to recover. However, even at 240 h, the amount of OAT1 did not reach control levels. The kidney tissues tended to show a remarkable but transient increase in mRNA expression for OAT1 at 5 min of ischemia. Our findings may provide insights of renal OAT1 in its cellular localization and response during ischemic acute kidney injury and recovery from it.

Download Full-text

Contribution of dedifferentiated proximal tubule cells to repair in acute kidney injury

Nature Reviews Nephrology ◽

10.1038/s41581-019-0235-3 ◽

2019 ◽

Vol 16 (2) ◽

pp. 65-65

Author(s):

Susan J. Allison

Keyword(s):

Acute Kidney Injury ◽

Proximal Tubule ◽

Kidney Injury ◽

Proximal Tubule Cells ◽

Tubule Cells

Download Full-text

6-Shogaol protects against ischemic acute kidney injury by modulating NF-κB and heme oxygenase-1 pathways

AJP Renal Physiology ◽

10.1152/ajprenal.00182.2019 ◽

2019 ◽

Vol 317 (3) ◽

pp. F743-F756 ◽

Cited By ~ 3

Author(s):

Sang Jun Han ◽

Mihwa Kim ◽

Vivette D. D’Agati ◽

H. Thomas Lee

Keyword(s):

Acute Kidney Injury ◽

Proximal Tubule ◽

Heme Oxygenase ◽

Kidney Injury ◽

Heme Oxygenase 1 ◽

Zinc Protoporphyrin ◽

Mrna Synthesis ◽

Proximal Tubule Cells ◽

Tnf Α ◽

Tubule Cells

Acute kidney injury (AKI) due to renal ischemia-reperfusion (I/R) is a major clinical problem without effective therapy. Ginger is one of the most widely consumed spices in the world, and 6-shogaol, a major ginger metabolite, has anti-inflammatory effects in neuronal and epithelial cells. Here, we demonstrate our novel findings that 6-shogaol treatment protected against renal I/R injury with decreased plasma creatinine, blood urea nitrogen, and kidney neutrophil gelatinase-associated lipocalin mRNA synthesis compared with vehicle-treated mice subjected to renal I/R. Additionally, 6-shogaol treatment reduced kidney inflammation (decreased proinflammatory cytokine and chemokine synthesis as well as neutrophil infiltration) and apoptosis (decreased TUNEL-positive renal tubular cells) compared with vehicle-treated mice subjected to renal I/R. In cultured human and mouse kidney proximal tubule cells, 6-shogaol significantly attenuated TNF-α-induced inflammatory cytokine and chemokine mRNA synthesis. Mechanistically, 6-shogaol significantly attenuated TNF-α-induced NF-κB activation in human renal proximal tubule cells by reducing IKKαβ/IκBα phosphorylation. Furthermore, 6-shogaol induced a cytoprotective chaperone heme oxygenase (HO)-1 via p38 MAPK activation in vitro and in vivo. Consistent with these findings, pretreatment with the HO-1 inhibitor zinc protoporphyrin IX completely prevented 6-shogaol-mediated protection against ischemic AKI in mice. Taken together, our study showed that 6-shogaol protects against ischemic AKI by attenuating NF-κB activation and inducing HO-1 expression. 6-Shogaol may provide a potential therapy for ischemic AKI during the perioperative period.

Download Full-text

Uptake and trafficking of fluorescent conjugates of folic acid in intact kidney determined using intravital two-photon microscopy

AJP Cell Physiology ◽

10.1152/ajpcell.00006.2004 ◽

2004 ◽

Vol 287 (2) ◽

pp. C517-C526 ◽

Cited By ~ 101

Author(s):

Ruben M. Sandoval ◽

Michael D. Kennedy ◽

Philip S. Low ◽

Bruce A. Molitoris

Keyword(s):

Folic Acid ◽

Proximal Tubule ◽

Rat Kidney ◽

Proximal Tubules ◽

Apical Surface ◽

Proximal Tubule Cells ◽

Two Photon Microscopy ◽

Two Photon ◽

Tubule Cells ◽

Folate Conjugate

Intravital two-photon microscopy was used to follow the uptake and trafficking of fluorescent conjugates of folic acid in the rat kidney. Intravenously administered folate-linked dye molecules quickly filled the plasma volume but not cellular components of the blood. Glomerular filtration occurred immediately and binding to proximal tubule cells was seen within seconds. Fluorescence from a pH-insensitive conjugate of folic acid, folate Texas red (FTR), was readily observed on the apical surface of the proximal tubules and in multiple cellular compartments, but little binding or uptake could be detected in any other kidney cells. Fluorescence from a pH-sensitive conjugate of folic acid, folate fluorescein, was seen only on the apical surface of proximal tubule cells, suggesting that internalized folate conjugates are localized to acidic compartments. The majority of the FTR conjugate internalized by proximal tubules accumulated within a lysosomal pool, as determined by colocalization studies. However, portions of FTR were also shown by electron microscopy to undergo transcytosis from apical to basal domains. Additional studies with colchicine, which is known to depolymerize microtubules and interrupt transcytosis, produced a marked reduction in endocytosis of FTR, with accumulation limited to the subapical region of the cell. No evidence of cytosolic release of either folate conjugate was observed, which may represent a key difference from the cytosolic deposition seen in neoplastic cells. Together, these data support the argument that folate conjugates (and, by extrapolation, physiological folate) bind to the apical surface of proximal tubule cells and are transported into and across the cells in endocytic compartments.

Download Full-text