Craig Slattery
University College Dublin, Science, Post-Doc
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Hepatocyte death is an important contributing factor in a number of diseases of the liver. PHD1 confers hypoxic sensitivity upon transcription factors including the hypoxia inducible factor (HIF) and nuclear factor-kappaB (NF-κB). Reduced... more
Hepatocyte death is an important contributing factor in a number of diseases of the liver. PHD1 confers hypoxic sensitivity upon transcription factors including the hypoxia inducible factor (HIF) and nuclear factor-kappaB (NF-κB). Reduced PHD1 activity is linked to decreased apoptosis. Here, we investigated the underlying mechanism(s) in hepatocytes. Basal NF-κB activity was elevated in PHD1(-/-) hepatocytes compared to wild type controls. ChIP-seq analysis confirmed enhanced binding of NF-κB to chromatin in regions proximal to the promoters of genes involved in the regulation of apoptosis. Inhibition of NF-κB (but not knock-out of HIF-1 or HIF-2) reversed the anti-apoptotic effects of pharmacologic hydroxylase inhibition. We hypothesize that PHD1 inhibition leads to altered expression of NF-κB-dependent genes resulting in reduced apoptosis. This study provides new information relating to the possible mechanism of therapeutic action of hydroxylase inhibitors that has been reported i...
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Research Interests: Computational Biology, Biology, Workflow, Risk assessment, Medicine, and 13 moreToxicogenomics, Transcriptome, Humans, Liver, Carcinogens, Observer Variation, Time Factors, Reproducibility of Results, Toxicogenetics, Risk Assessment, Gene Expression Regulation, Gene expression profiling, and Pharmacology and pharmaceutical sciences
Key points In cystinosis, a lysosomal storage disorder, an altered redox state has been suggested as contributing to cellular dysfunction. Ctns gene knockdown in a pancreatic β‐cell line caused increased cystine levels. Attenuated... more
Key points In cystinosis, a lysosomal storage disorder, an altered redox state has been suggested as contributing to cellular dysfunction. Ctns gene knockdown in a pancreatic β‐cell line caused increased cystine levels. Attenuated nutrient stimulated insulin secretion was observed after Ctns knockdown which may have been caused by an increase in oxidative stress. Oxidative stress may reduce ATP production in pancreatic β‐cells resulting in attenuated insulin release. The redox‐sensitive transcription factor NF‐κB was activated after Ctns knockdown which may contribute to the increased incidence of apoptosis. The pancreatic β‐cell has reduced antioxidant defences making it more susceptible to oxidative stress. In cystinosis, a lysosomal storage disorder, an altered redox state may contribute to cellular dysfunction. This rare disease is caused by an abnormal lysosomal cystine transporter, cystinosin, which causes excessive accumulation of cystine in the lysosome. Cystinosis associate...
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One of the main goals in functional genomics has been the development of tools that allow easy manipulation of gene expression levels that would be suitable for high throughput screening. RNA interference (RNAi) has emerged as one of the... more
One of the main goals in functional genomics has been the development of tools that allow easy manipulation of gene expression levels that would be suitable for high throughput screening. RNA interference (RNAi) has emerged as one of the preferred approaches to achieve this goal. It is an important biological mechanism in the regulation of gene expression in animals and plants. Here we evaluated the use of RNAi for knockdown of specific gene expression as an alternative to the production of transgenic animals. RNAi development may be in the early stages, however, the real and theoretical advantages of this system in reducing the use of animals merit further investigations.
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Research Interests: Biomarkers, Kidney diseases, Kidney transplantation, Medicine, Renal transplantation, and 13 moreHumans, Allografts, Renal Function, Biomarker, Urinary System, Proteinuria, Amino Acid Sequence, Graft Rejection, Clusterin, Delayed graft function, Molecular Sequence Data, Chronic Allograft Nephropathy, and Medical biochemistry and metabolomics
ClC-5 is a chloride/proton exchanger that plays an obligate role in albumin uptake by the renal proximal tubule. ClC-5 forms an endocytic complex with the albumin receptor megalin/cubilin. We have identified a novel ClC-5 binding partner,... more
ClC-5 is a chloride/proton exchanger that plays an obligate role in albumin uptake by the renal proximal tubule. ClC-5 forms an endocytic complex with the albumin receptor megalin/cubilin. We have identified a novel ClC-5 binding partner, aspartyl-aminopeptidase (DNPEP), that catalyses the release of N-terminal aspartate/glutamate residues. The physiological role of DNPEP remains largely unresolved. Mass spectrometric analysis of proteins binding to GST-ClC-5 C-terminus identified DNPEP as an interacting partner. Coimmunoprecipitation confirmed that DNPEP and ClC-5 also associated in cells. Experiments using purified GST-ClC-5 and His-DNPEP proteins demonstrated that the two proteins bound directly to each other. In opossum kidney (OK) cells, confocal immunofluorescence studies revealed that DNPEP co-localized with albumin containing endocytic vesicles. Over-expression of wild-type DNPEP increased cell-surface levels of ClC-5 and albumin uptake. Analysis of DNPEP immunoprecipitated ...
Research Interests: Physiology, Chemistry, Urology, Biology, Medicine, and 9 moreKidney, Endocytosis, Animals, Medical Physiology, Endosome, Rats, Albuminuria, CLC, and Cell Membrane(Kidney, Endocytosis, Animals, Medical Physiology, Endosome, Rats, Albuminuria, CLC, and Cell Membrane)
(Kidney, Endocytosis, Animals, Medical Physiology, Endosome, Rats, Albuminuria, CLC, and Cell Membrane)
This paper focuses on the role that mitogen-activated protein kinases (MAPKs) play in drug-induced kidney injury. The MAPKs, of which there are four major classes (ERK, p38, JNK, and ERK5/BMK), are signalling cascades which have been... more
This paper focuses on the role that mitogen-activated protein kinases (MAPKs) play in drug-induced kidney injury. The MAPKs, of which there are four major classes (ERK, p38, JNK, and ERK5/BMK), are signalling cascades which have been found to be broadly conserved across a wide variety of organisms. MAPKs allow effective transmission of information from the cell surface to the cytosolic or nuclear compartments. Cross talk between the MAPKs themselves and with other signalling pathways allows the cell to modulate responses to a wide variety of external stimuli. The MAPKs have been shown to play key roles in both mediating and ameliorating cellular responses to stress including xenobiotic-induced toxicity. Therefore, this paper will discuss the specific role of the MAPKs in the kidney in response to injury by a variety of xenobiotics and the potential for therapeutic intervention at the level of MAPK signalling across different types of kidney disease.
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Research Interests: Computational Biology, Biology, Cell Adhesion, Oxidative Stress, Apoptosis, and 15 moreMedicine, Cell line, Humans, Reactive Oxygen Species, Cyclosporine, Time Factors, Transforming Growth Factor Beta, Cell Proliferation, Nephrotoxicity, Mesangial Cells, Gene Expression Regulation, Gene expression profiling, Immunosuppressive Agents, renal insufficiency, and Pharmacology and pharmaceutical sciences(Medicine, Cell line, Humans, Reactive Oxygen Species, Cyclosporine, Time Factors, Transforming Growth Factor Beta, Cell Proliferation, Nephrotoxicity, Mesangial Cells, Gene Expression Regulation, Gene expression profiling, Immunosuppressive Agents, renal insufficiency, and Pharmacology and pharmaceutical sciences)
(Medicine, Cell line, Humans, Reactive Oxygen Species, Cyclosporine, Time Factors, Transforming Growth Factor Beta, Cell Proliferation, Nephrotoxicity, Mesangial Cells, Gene Expression Regulation, Gene expression profiling, Immunosuppressive Agents, renal insufficiency, and Pharmacology and pharmaceutical sciences)
Research Interests: Principal Component Analysis, Fluorescence Microscopy, Biology, Metabolomics, Magnetic Resonance Spectroscopy, and 15 moreMedicine, Cell Culture, Cell line, Humans, Journal Article, Epithelial cells, Diclofenac, Environmental Pollutants, Toxicant, Nifedipine, Metabolite, Metabolome, Biochemistry and cell biology, Mannitol, and Ochratoxins
Molecular O(2) and CO(2) are the primary substrate and product of aerobic metabolism, respectively. Levels of these physiologic gases in the cell microenvironment vary dramatically both in health and in diseases, such as chronic... more
Molecular O(2) and CO(2) are the primary substrate and product of aerobic metabolism, respectively. Levels of these physiologic gases in the cell microenvironment vary dramatically both in health and in diseases, such as chronic inflammation, ischemia, and cancer, in which metabolism is significantly altered. The identification of the hypoxia-inducible factor led to the discovery of an ancient and direct link between tissue O(2) and gene transcription. In this study, we demonstrate that mammalian cells (mouse embryonic fibroblasts and others) also sense changes in local CO(2) levels, leading to altered gene expression via the NF-κB pathway. IKKα, a central regulatory component of NF-κB, rapidly and reversibly translocates to the nucleus in response to elevated CO(2). This response is independent of hypoxia-inducible factor hydroxylases, extracellular and intracellular pH, and pathways that mediate acute CO(2)-sensing in nematodes and flies and leads to attenuation of bacterial LPS-i...
Research Interests: Immunology, Carbon Dioxide, Fluorescence Microscopy, Biology, Inflammation, and 15 moreCell Biology, Immunology of the Gut, Confocal Microscopy, Innate immunity, Medicine, Gene expression, Signal Transduction, Humans, Mice, Animals, NF κB, Protein Transport, Gene Expression Regulation, Innate Immune System, and reverse transcriptase polymerase chain reaction
Research Interests: Chemistry, Biology, Cell Biology, Confocal Microscopy, Medicine, and 15 moreRNA interference, Endocytosis, Animals, Medical Physiology, Protein Interaction, Rats, Amino Acid Sequence, Protein Binding, Proximal Tubule, Fusion Protein, Binding Site, Scaffold Proteins, Molecular Sequence Data, PDZ domain, and binding sites
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The primary cilium is an immotile sensory and signaling organelle found on the majority of mammalian cell types. Of the multitude of roles that the primary cilium performs, perhaps some of the most important include maintenance of... more
The primary cilium is an immotile sensory and signaling organelle found on the majority of mammalian cell types. Of the multitude of roles that the primary cilium performs, perhaps some of the most important include maintenance of differentiation, quiescence, and cellular polarity. Given that the progression of cancer requires disruption of all of these processes, we have investigated the effects of several carcinogens on the primary cilium of the RPTEC/TERT1 human proximal tubular epithelial cell line. Using both scanning electron microscopy and immunofluorescent labeling of the ciliary markers acetylated tubulin and Arl13b, we confirmed that RPTEC/TERT1 cells express primary cilium upon reaching confluence. Treatment with the carcinogens ochratoxin A (OTA) and potassium bromate (KBrO3) caused a significant reduction in the number of ciliated cells, while exposure to nifedipine, a noncarcinogenic renal toxin, had no effect on primary cilium expression. Flow cytometric analysis of t...
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The immunosuppressant drugs cyclosporine A (CsA) and sirolimus (SRL) used in combination demonstrated beneficial effects in organ transplantation, but this combination can also result in increased adverse effects. We previously showed... more
The immunosuppressant drugs cyclosporine A (CsA) and sirolimus (SRL) used in combination demonstrated beneficial effects in organ transplantation, but this combination can also result in increased adverse effects. We previously showed that not only CsA treatment but also its combination with SRL decreased paracellular permeability in renal proximal tubular cells by modification of the tight junction proteins, claudins, through ERK1/2 signaling pathway. In this present study, evidence is presented that not only CsA but also the combination of CsA/SRL may have adverse effects on the barrier function of renal proximal cells, at least in part, through the expression of the cytokine transforming growth factor (TGF)-β1. CsA treatment upregulated TGF-β1gene expression and this upregulation was enhanced when CsA and SRL were applied together. Addition of TGF-β1(5 ng/ml) altered the barrier function with increased transepithelial electrical resistance (TER) and claudin-1 expression. Use of a...
Research Interests: Physiology, Biology, Membrane Proteins, Enzyme Inhibitors, Medicine, and 15 moreGene expression, Signal Transduction, Cell line, Kidney, Nitriles, Animals, American, Cyclosporine, Medical Physiology, Epithelial cells, Sirolimus, Electric Impedance, Claudin, Smad proteins, and Immunosuppressive Agents
Background: High-mobility group box protein 1 (HMGB-1) is a chromatin-binding protein that bends DNA, thereby facilitating gene transcription. HMGB-1 has also been observed as an extracellular secreted protein in serum of patients with... more
Background: High-mobility group box protein 1 (HMGB-1) is a chromatin-binding protein that bends DNA, thereby facilitating gene transcription. HMGB-1 has also been observed as an extracellular secreted protein in serum of patients with sepsis and has putative intracellular signalling effects regulating the production of interleukin-1 and tumour necrosis factor in a number of inflammatory conditions. Methods: We established a model of immune-mediated epithelial-mesenchymal transition (EMT) in human proximal tubular epithelial cells (PTECs). PTECs were cultured with conditioned medium containing supernatant from activated peripheral blood mononuclear cells (aPBMCs). The model was characterized using phenotypic and transcriptomic approaches and suppression subtractive hybridisation was performed to identify differentially regulated genes. Results: Activation of PBMCs resulted in increased secretion of HMGB-1. In addition, treatment of PTECs with aPBMC-conditioned medium resulted in sig...
Research Interests: Biology, Cell Migration, Cell Biology, Medicine, Humans, and 15 moreNucleic acid hybridization, Phenotype, Clinical Sciences, Epithelial cells, Analysis of Variance, Receptor for Advanced Glycation End Products, Protein Binding, Renal Fibrosis, PBMC, Blood cells, Secretion, Gene Expression Regulation, Transforming Growth Factor, Cadherins, and Conditioned medium
The natural compound curcumin has been shown to have therapeutic potential against a wide range of diseases such as cancer. Curcumin reduces cell viability of renal cell carcinoma (RCC) cells when combined with TNF-related... more
The natural compound curcumin has been shown to have therapeutic potential against a wide range of diseases such as cancer. Curcumin reduces cell viability of renal cell carcinoma (RCC) cells when combined with TNF-related apoptosis-inducing ligand (TRAIL), a cytokine that specifically targets cancer cells, by helping overcome TRAIL resistance. However, the therapeutic effects of curcumin are limited by its low bioavailability. Similar compounds to curcumin with higher bioavailability, such as demethoxycurcumin (DMC) and 3,5-bis(2-fluorobenzylidene)-4-piperidone (EF24), can potentially have similar anticancer effects and show a similar synergy with TRAIL, thus reducing RCC viability. This study aims to show the effects of DMC and EF24 in combination with TRAIL at reducing ACHN cell viability and ACHN cell migration. It also shows the changes in death receptor 4 (DR4) expression after treatment with these compounds individually and in combination with TRAIL, which can play a role in ...
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Cyclosporine A, which has been the foremost immunosuppressive agent since the early 1980's, significantly improves the success of organ transplantation. However, common complications of cyclosporine A therapy, such as severe renal... more
Cyclosporine A, which has been the foremost immunosuppressive agent since the early 1980's, significantly improves the success of organ transplantation. However, common complications of cyclosporine A therapy, such as severe renal tubulointerstitial fibrosis, limit the drug's clinical use. Although the exact mechanisms driving cyclosporine A-induced tubulointerstitial fibrosis remain elusive, we hypothesized that epithelial-mesenchymal transition (EMT) may play a major role. We investigated this in vitro by treating human proximal tubular cells with cyclosporine A. Morphological changes were observed after cyclosporine A treatment, including cell elongation (with a large degree of detachment), cytoskeletal rearrangement, and junctional disruption. In addition, expression of the myofibroblast-specific marker α-smooth muscle actin was detected in treated cells. These observations are consistent with events described during EMT. Using Affymetrix gene microarrays, we identified 128 genes that were differentially regulated in renal tubular cells after cyclosporine A treatment, including known profibrotic factors, oncogenes, and transcriptional regulators. Cyclosporine A induced a dose-dependent increase in transforming growth factor-β secretion from proximal tubular cells. Subsequent functional studies revealed that protein kinase C-β isoforms play a key role in cyclosporine A-induced effects. These findings provide novel insights into cyclosporine A-induced renal fibrosis and the molecular mechanisms underlying EMT, events that may be relevant in other disease states.
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The calcineurin inhibitor cyclosporine A (CsA) is a widely used immunosuppressive agent. However, nephrotoxicity is a serious side effect observed in patients which limits clinical use of CsA. CsA nephrotoxicity is associated with... more
The calcineurin inhibitor cyclosporine A (CsA) is a widely used immunosuppressive agent. However, nephrotoxicity is a serious side effect observed in patients which limits clinical use of CsA. CsA nephrotoxicity is associated with tubulointerstitial injury progressing to nephropathy. This is typically diagnosed by invasive renal biopsy and is often only detected when the disease process is well advanced. Therefore identification of novel, early indicators of CsA nephrotoxicity could be clinically advantageous. This study aimed to establish a murine model of CsA nephrotoxicity and to identify urinary proteins that may indicate the onset of CsA-induced nephropathy using 2-D gel electrophoresis. CsA nephrotoxicity was induced in CD-1 mice by daily CsA administration for 4 weeks. By week 4, elevated serum creatinine and proteinuria were observed after CsA treatment indicating significant renal dysfunction. Decreased cadherin-1, increased α-smooth muscle actin and fibroblast specific protein 1 in kidney tissue indicated disruption of normal tubular architecture. Alterations in podocin and uromodulin were also observed which may indicate damage to other segments of the nephron. Proteomic analysis of urine identified a number of differentially regulated proteins that may be involved in early CsA nephropathy including cadherin 1, superoxide dismutase and vinculin. These findings suggest novel mechanisms of CsA nephrotoxicity and identify novel potential markers of the disease.
Research Interests: Identification, Toxicity, Kidney, Mice, Animals, and 16 moreMale, Animal Model, Cyclosporine, Superoxide Dismutase, Serum Creatinine, Mouse Model, Urinary System, Transforming Growth Factor Beta, Proteome analysis, Side Effect, Renal biopsy, Gel electrophoresis, Cadherin, Murine Model, Renal Dysfunction, and Glomerulonephritis
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Cyclosporine A (CsA) significantly improves the success of organ transplantation, however renal fibrosis, characterised by severe tubulointerstitial fibrosis is a complication of CsA therapy. Previously we have reported the involvement of... more
Cyclosporine A (CsA) significantly improves the success of organ transplantation, however renal fibrosis, characterised by severe tubulointerstitial fibrosis is a complication of CsA therapy. Previously we have reported the involvement of PKC-β isoforms in a model of CsA-induced tubulointerstitial fibrosis and we have now further elucidated this role. Treatment of human proximal tubular epithelial cells with CsA resulted in increased fibronectin production which coincided with increased PKC activity. To delineate the respective contributions of the two PKC-β isoforms in fibrotic events, we overexpressed PKC-βI, -βII, or both in combination. Overexpression of the two PKC-β isoforms induced morphological alterations, secretion of the profibrotic cytokine TGF-β1, and fibronectin release from proximal tubular cells however PKC-βII induced more significant effects in all parameters examined. Inhibition of PKC-β completely abrogated the CsA-induced increase in fibronectin secretion demonstrating a direct antifibrotic effect of PKC-β inhibition. Further studies also identified a role for the p44/42 mitogen activated kinase signalling pathway in CsA-induced fibrotic effects downstream of PKC-β. Overall, these findings demonstrate a central role for PKC-β, and particularly PKC-βII in the development of tubulointerstitial fibrosis and suggest that PKC-β may be a viable therapeutic target in CsA nephropathy.
Research Interests: Signal Transduction, Cell line, Humans, Nitriles, Organ Transplantation, and 13 moreCyclosporine, Fibrosis, Phenotype, Mitogen Activated Protein Kinase, Fibronectin, Epithelial cells, CSA, Isoenzymes, Pyridines, Cell Shape, Transforming Growth Factor Beta, Imidazoles, and Biochemistry and cell biology
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The E2A gene encodes two distinct basic helix-loop-helix transcription factors, E12 and E47. E12 and E47 modulate expression of their target genes through formation of homodimers and heterodimers with other basic helix-loop-helix... more
The E2A gene encodes two distinct basic helix-loop-helix transcription factors, E12 and E47. E12 and E47 modulate expression of their target genes through formation of homodimers and heterodimers with other basic helix-loop-helix transcription factors. E2A proteins are thought to play critical roles in regulation of cell commitment, growth and differentiation in a range of cell types including lymphocytes, muscle cells and neurons. Emerging evidence suggests that E2A proteins also play key roles in the process of epithelial mesenchymal transition, a mechanism which contributes significantly to kidney disease progression and tumour metastasis. Further understanding of the diverse effects of E2A proteins may lead to novel therapeutic approaches to targeting important disease process.
Research Interests: Gene expression, Epithelial-Mesenchymal Transition, Humans, Animals, Alternative splicing, and 10 morePhenotype, Transcription Factor, Embryonic Development, Alternative Splicing, Cell Proliferation, Amino Acid Sequence, Lymphocytes, Biochemistry and cell biology, Dimerization, and Molecular Sequence Data
Epithelial–mesenchymal transition (EMT), a process whereby renal tubular epithelial cells lose phenotype and gain fibroblast-like characteristics, has been demonstrated to contribute significantly to the development of renal fibrosis. The... more
Epithelial–mesenchymal transition (EMT), a process whereby renal tubular epithelial cells lose phenotype and gain fibroblast-like characteristics, has been demonstrated to contribute significantly to the development of renal fibrosis. The immunosuppressant cyclosporine A (CsA) has been shown to induce renal fibrosis, a major complication of CsA therapy. The mechanisms that drive CsA-induced fibrosis remain undefined, however, CsA has been demonstrated to induce EMT in human renal proximal tubular epithelial cells (RPTEC). E2A transcription factors were identified as being upregulated by CsA treatment. To further examine the role of E2A proteins in EMT, E12 and E47 were overexpressed, alone and in combination, in human RPTEC. Both E12 and E47 elicited EMT effects on tubular epithelial cells with E47 more potent in inducing the fibroblast-like phenotype. These results indicate the important role of the E2A gene products in the progression of CsA-induced EMT and provide novel insights into CsA-induced renal fibrosis.