scholarly journals Danegaptide Prevents TGFβ1-Induced Damage in Human Proximal Tubule Epithelial Cells of the Kidney

2021 ◽  
Vol 22 (6) ◽  
pp. 2809
Author(s):  
Paul E. Squires ◽  
Gareth W. Price ◽  
Ulrik Mouritzen ◽  
Joe A. Potter ◽  
Bethany M. Williams ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Proximal Tubule ◽  
Transforming Growth Factor ◽  
Atp Release ◽  
Paracellular Permeability ◽  
Antibody Array ◽  
Dye Uptake ◽  
Partial Restoration ◽  
And Function ◽  
Human Proximal Tubule

Chronic kidney disease (CKD) is a global health problem associated with a number of comorbidities. Recent evidence implicates increased hemichannel-mediated release of adenosine triphosphate (ATP) in the progression of tubulointerstitial fibrosis, the main underlying pathology of CKD. Here, we evaluate the effect of danegaptide on blocking hemichannel-mediated changes in the expression and function of proteins associated with disease progression in tubular epithelial kidney cells. Primary human proximal tubule epithelial cells (hPTECs) were treated with the beta1 isoform of the pro-fibrotic cytokine transforming growth factor (TGFβ1) ± danegaptide. qRT-PCR and immunoblotting confirmed mRNA and protein expression, whilst a cytokine antibody array assessed the expression/secretion of proinflammatory and profibrotic cytokines. Carboxyfluorescein dye uptake and ATP biosensing measured hemichannel activity and ATP release, whilst transepithelial electrical resistance was used to assess paracellular permeability. Danegaptide negated carboxyfluorescein dye uptake and ATP release and protected against protein changes associated with tubular injury. Blocking Cx43-mediated ATP release was paralleled by partial restoration of the expression of cell cycle inhibitors, adherens and tight junction proteins and decreased paracellular permeability. Furthermore, danegaptide inhibited TGFβ1-induced changes in the expression and secretion of key adipokines, cytokines, chemokines, growth factors and interleukins. The data suggest that as a gap junction modulator and hemichannel blocker, danegaptide has potential in the future treatment of CKD.

scholarly journals α1-Microglobulin (A1M) Protects Human Proximal Tubule Epithelial Cells from Heme-Induced Damage In Vitro

2020 ◽  
Vol 21 (16) ◽  
pp. 5825 ◽  
Author(s):  
Amanda Kristiansson ◽  
Sara Davidsson ◽  
Maria E. Johansson ◽  
Sarah Piel ◽  
Eskil Elmér ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Epithelial Cells ◽  
Proximal Tubule ◽  
Mitochondrial Function ◽  
Radical Scavenging ◽  
Related Factor ◽  
Protective Effects ◽  
Cellular Expression ◽  
Human Proximal Tubule

Oxidative stress is associated with many renal disorders, both acute and chronic, and has also been described to contribute to the disease progression. Therefore, oxidative stress is a potential therapeutic target. The human antioxidant α1-microglobulin (A1M) is a plasma and tissue protein with heme-binding, radical-scavenging and reductase activities. A1M can be internalized by cells, localized to the mitochondria and protect mitochondrial function. Due to its small size, A1M is filtered from the blood into the glomeruli, and taken up by the renal tubular epithelial cells. A1M has previously been described to reduce renal damage in animal models of preeclampsia, radiotherapy and rhabdomyolysis, and is proposed as a pharmacological agent for the treatment of kidney damage. In this paper, we examined the in vitro protective effects of recombinant human A1M (rA1M) in human proximal tubule epithelial cells. Moreover, rA1M was found to protect against heme-induced cell-death both in primary cells (RPTEC) and in a cell-line (HK-2). Expression of stress-related genes was upregulated in both cell cultures in response to heme exposure, as measured by qPCR and confirmed with in situ hybridization in HK-2 cells, whereas co-treatment with rA1M counteracted the upregulation. Mitochondrial respiration, analyzed with the Seahorse extracellular flux analyzer, was compromised following exposure to heme, but preserved by co-treatment with rA1M. Finally, heme addition to RPTE cells induced an upregulation of the endogenous cellular expression of A1M, via activation of the nuclear factor erythroid 2-related factor 2 (Nrf2)-pathway. Overall, data suggest that A1M/rA1M protects against stress-induced damage to tubule epithelial cells that, at least partly, can be attributed to maintaining mitochondrial function.

scholarly journals Transforming Growth Factor Beta 1 Drives a Switch in Connexin Mediated Cell-to-Cell Communication in Tubular Cells of the Diabetic Kidney

2018 ◽  
Vol 45 (6) ◽  
pp. 2369-2388 ◽  
Author(s):  
Claire Hills ◽  
Gareth William Price ◽  
Mark John Wall ◽  
Timothy John Kaufmann ◽  
Chi-Wai Tang ◽  
...  
Keyword(s):  
Diabetic Nephropathy ◽  
Epithelial Cells ◽  
Transforming Growth Factor ◽  
Cell Communication ◽  
Atp Release ◽  
Immunoblot Analysis ◽  
Biopsy Material ◽  
Junctional Conductance ◽  
Fibronectin Expression ◽  
Tgf Β1

Background/Aims: Changes in cell-to-cell communication have been linked to several secondary complications of diabetes, but the mechanism by which connexins affect disease progression in the kidney is poorly understood. This study examines a role for glucose-evoked changes in the beta1 isoform of transforming growth factor (TGFβ1), on connexin expression, gap-junction mediated intercellular communication (GJIC) and hemi-channel ATP release from tubular epithelial cells of the proximal renal nephron. Methods: Biopsy material from patients with and without diabetic nephropathy was stained for connexin-26 (CX26) and connexin-43 (CX43). Changes in expression were corroborated by immunoblot analysis in human primary proximal tubule epithelial cells (hPTECs) and model epithelial cells from human renal proximal tubules (HK2) cultured in either low glucose (5mmol/L) ± TGFβ1 (2-10ng/ml) or high glucose (25mmol/L) for 48h or 7days. Secretion of the cytokine was determined by ELISA. Paired whole cell patch clamp recordings were used to measure junctional conductance in control versus TGFβ1 treated (10ng/ml) HK2 cells, with carboxyfluorescein uptake and ATP-biosensing assessing hemi-channel function. A downstream role for ATP in mediating the effects of TGF-β1 on connexin mediated cell communication was assessed by incubating cells with ATPγS (1-100µM) or TGF-β1 +/- apyrase (5 Units/ml). Implications of ATP release were measured through immunoblot analysis of interleukin 6 (IL-6) and fibronectin expression. Results: Biopsy material from patients with diabetic nephropathy exhibited increased tubular expression of CX26 and CX43 (P<0.01, n=10), data corroborated in HK2 and hPTEC cells cultured in TGFβ1 (10ng/ml) for 7days (P<0.001, n=3). High glucose significantly increased TGFβ1 secretion from tubular epithelial cells (P<0.001, n=3). The cytokine (10ng/ml) reduced junctional conductance between HK2 cells from 4.5±1.3nS in control to 1.15±0.9nS following 48h TGFβ1 and to 0.42±0.2nS after 7days TGFβ1 incubation (P<0.05, n=5). Acute (48h) and chronic (7day) challenge with TGFβ1 produced a carbenoxolone (200µM)-sensitive increase in carboxyfluorescein loading, matched by an increase in ATP release from 0.29±0.06μM in control to 1.99±0.47μM after 48hr incubation with TGFβ1 (10ng/ml; P<0.05, n=3). TGF-β1 (2-10ng/ml) and ATPγs (1-100µM) increased expression of IL-6 (P<0.001 n=3) and fibronectin (P<0.01 n=3). The effect of TGF-β1 on IL-6 and fibronectin expression was partially blunted when preincubated with apyrase (n=3). Conclusion: These data suggest that chronic exposure to glucose-evoked TGFβ1 induce an increase in CX26 and CX43 expression, consistent with changes observed in tubular epithelia from patients with diabetic nephropathy. Despite increased connexin expression, direct GJIC communication decreases, whilst hemichannel expression/function and paracrine release of ATP increases, changes that trigger increased levels of expression of interleukin 6 and fibronectin. Linked to inflammation and fibrosis, local increases in purinergic signals may exacerbate disease progression and highlight connexin mediated cell communication as a future therapeutic target for diabetic nephropathy.

scholarly journals Proinflammatory Cytokines and Potassium Channels in the Kidney

2015 ◽  
Vol 2015 ◽  
pp. 1-8 ◽  
Author(s):  
Kazuyoshi Nakamura ◽  
Hikaru Hayashi ◽  
Manabu Kubokawa
Keyword(s):  
Proximal Tubule ◽  
Proinflammatory Cytokines ◽  
Transforming Growth Factor ◽  
Interleukin 1 ◽  
Transepithelial Transport ◽  
Channel Activity ◽  
Proximal Tubule Cells ◽  
Cell Functions ◽  
Tubule Cells ◽  
Human Proximal Tubule

Proinflammatory cytokines affect several cell functions via receptor-mediated processes. In the kidney, functions of transporters and ion channels along the nephron are also affected by some cytokines. Among these, alteration of activity of potassium ion (K+) channels induces changes in transepithelial transport of solutes and water in the kidney, since K+channels in tubule cells are indispensable for formation of membrane potential which serves as a driving force for the transepithelial transport. Altered K+channel activity may be involved in renal cell dysfunction during inflammation. Although little information was available regarding the effects of proinflammatory cytokines on renal K+channels, reports have emerged during the last decade. In human proximal tubule cells, interferon-γshowed a time-dependent biphasic effect on a 40 pS K+channel, that is, delayed suppression and acute stimulation, and interleukin-1βacutely suppressed the channel activity. Transforming growth factor-β1 activated KCa3.1 K+channel in immortalized human proximal tubule cells, which would be involved in the pathogenesis of renal fibrosis. This review discusses the effects of proinflammatory cytokines on renal K+channels and the causal relationship between the cytokine-induced changes in K+channel activity and renal dysfunction.

scholarly journals Caveolin- and clathrin-independent entry of BKPyV into primary human proximal tubule epithelial cells

Virology ◽  
2016 ◽  
Vol 492 ◽  
pp. 66-72 ◽  
Author(s):  
Linbo Zhao ◽  
Anthony T. Marciano ◽  
Courtney R. Rivet ◽  
Michael J. Imperiale
Keyword(s):  
Epithelial Cells ◽  
Proximal Tubule ◽  
Human Proximal Tubule

scholarly journals Human proximal tubule epithelial cells modulate autologous B-cell function

2015 ◽  
Vol 30 (10) ◽  
pp. 1674-1683 ◽  
Author(s):  
Sandeep Sampangi ◽  
Xiangju Wang ◽  
Kenneth W. Beagley ◽  
Travis Klein ◽  
Sadia Afrin ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Proximal Tubule ◽  
B Cell ◽  
Cell Function ◽  
B Cell Function ◽  
Human Proximal Tubule

scholarly journals Human proximal tubule epithelial cells modulate autologous dendritic cell function

2012 ◽  
Vol 28 (2) ◽  
pp. 303-312 ◽  
Author(s):  
Andrew J. Kassianos ◽  
Sandeep Sampangi ◽  
Xiangju Wang ◽  
Kathrein E. Roper ◽  
Ken Beagley ◽  
...  
Keyword(s):  
Dendritic Cell ◽  
Epithelial Cells ◽  
Proximal Tubule ◽  
Cell Function ◽  
Dendritic Cell Function ◽  
Autologous Dendritic Cell ◽  
Human Proximal Tubule

Beta-adrenergic receptor function in rat proximal tubule epithelial cells in culture

1995 ◽  
Vol 268 (4) ◽  
pp. F553-F560 ◽  
Author(s):  
A. S. Hanson ◽  
S. L. Linas
Keyword(s):  
Epithelial Cells ◽  
Adenylate Cyclase ◽  
Proximal Tubule ◽  
Adrenergic Receptors ◽  
Proximal Tubules ◽  
Sodium Reabsorption ◽  
Scatchard Analysis ◽  
Beta Adrenergic ◽  
Alpha Adrenergic Receptors ◽  
And Function

The adrenergic system is important in regulating proximal tubule sodium reabsorption. Although alpha-adrenergic receptors have been identified in proximal tubules, the presence and function of beta-adrenergic receptors (BAR) in proximal tubules is less certain. The purpose of our study was to determine whether functional BAR are present on apical or basolateral surfaces of proximal tubule epithelial cells (PTEC) of rat kidney. We specifically focused on BAR coupling to adenylate cyclase and on differences between requirements for apical and basolateral receptor coupling to adenylate cyclase. To determine BAR expression and function, primary cultures of rat PTECs were grown on permeable supports. Scatchard analysis of 125I-labeled cyanopindolol binding revealed a single class of receptors on both apical and basolateral surfaces. Apical isoproterenol (ISO) resulted in time- and concentration-dependent increases in adenosine 3',5'-cyclic monophosphate (cAMP) that were 50% of responses after basolateral ISO. Apical BAR-cAMP coupling was mediated by B1-adrenergic receptors (B1AR), since apical cAMP responses were abrogated with apical (but not basolateral) B1 but not B2 antagonists. Apical B1AR required endocytosis prior to adenylate cyclase activation, since increases in cAMP were prevented by phenylarsine oxide or colchicine. B1AR-adenylate cyclase coupling was independent of intra- or extracellular calcium, cyclooxygenase metabolites, and protein kinase C (PKC) and dependent on Gs guanine nucleotide regulatory protein. Prolonged exposure to ISO resulted in time- and concentration-dependent homologous desensitization of cAMP responses. Desensitization was independent of receptor sequestration, PKA, or PKC. We conclude the following: B1AR are present on both apical and basolateral surfaces of rat PTECs.(ABSTRACT TRUNCATED AT 250 WORDS)

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