Type 1 Natural Killer T-cells (NKT1 cells) play a critical role in mediating hepatic ischemia-rep... more Type 1 Natural Killer T-cells (NKT1 cells) play a critical role in mediating hepatic ischemia-reperfusion injury (IRI). Although hepatic steatosis is a major risk factor for preservation type injury, how NKT cells impact this is understudied. Given NKT1 cell activation by phospholipid ligands recognized presented by CD1d, we hypothesized that NKT1 cells are key modulators of hepatic IRI because of the increased frequency of activating ligands in the setting of hepatic steatosis. We first demonstrate that IRI is exacerbated by a high-fat diet (HFD) in experimental murine models of warm partial ischemia. This is evident in the evaluation of ALT levels and Phasor-Fluorescence Lifetime (Phasor-FLIM) Imaging for glycolytic stress. Polychromatic flow cytometry identified pronounced increases in CD45+CD3+NK1.1+NKT1 cells in HFD fed mice when compared to mice fed a normal diet (ND). This observation is further extended to IRI, measuring ex vivo cytokine expression in the HFD and ND. Much hi...
HIV disease remains prevalent in the USA and chronic kidney disease remains a major cause of morb... more HIV disease remains prevalent in the USA and chronic kidney disease remains a major cause of morbidity in HIV-1-positive patients. Host double-stranded RNA (dsRNA)-activated protein kinase (PKR) is a sensor for viral dsRNA, including HIV-1. We show that PKR inhibition by compound C16 ameliorates the HIV-associated nephropathy (HIVAN) kidney phenotype in the Tg26 transgenic mouse model, with reversal of mitochondrial dysfunction. Combined analysis of single-nucleus RNA-seq and bulk RNA-seq data revealed that oxidative phosphorylation was one of the most downregulated pathways and identified signal transducer and activator of transcription (STAT3) as a potential mediating factor. We identified in Tg26 mice a novel proximal tubular cell cluster enriched in mitochondrial transcripts. Podocytes showed high levels of HIV-1 gene expression and dysregulation of cytoskeleton-related genes; and these cells dedifferentiated and were lost from the glomerular tuft. In injured proximal tubules, c...
All forms of progressive renal diseases develop a final pathway of tubulointerstitial fibrosis an... more All forms of progressive renal diseases develop a final pathway of tubulointerstitial fibrosis and glomerulosclerosis. Renal fibrosis is usually quantified using histological staining, a process that is time-consuming and pathologist dependent. Here we develop a fast and operator-independent method to measure fibrosis utilizing the murine unilateral ureteral obstruction model which manifests a time-dependent fibrotic increase in obstructed kidneys while the contralateral kidneys are used as controls. After ureteral obstruction, kidneys were analyzed at 7, 14, and 21 days. Fibrosis was quantified using fluorescence lifetime imaging (FLIM) and second harmonic generation (SHG) in a Deep Imaging via Enhanced photon Recovery deep tissue imaging microscope. This microscope was developed for deep tissue along with second and third harmonic generation imaging and has extraordinary sensitivity toward harmonic generation. SHG data suggest the presence of more fibrillar collagen in the obstruc...
Sodium glucose cotransporter, type 2 inhibitors, such as Empagliflozin, are protective of the kid... more Sodium glucose cotransporter, type 2 inhibitors, such as Empagliflozin, are protective of the kidneys by unclear mechanisms. Our aim was to determine how Empagliflozin affected kidney cortical metabolome and lipidome in mice. Adult male TALLYHO mice (prone to obesity) were treated with a high-milk-fat diet, or this diet containing Empagliflozin (0.01%), for 8 weeks. Targeted and untargeted metabolomics and lipidomics were conducted on kidney cortex by liquid chromatography followed by tandem mass-spectroscopy. Metabolites were statistically analyzed by MetaboAnalyst 5.0, LipidSig (lipid species only) and/or CEU Mass Mediator (untargeted annotation). In general, volcano plotting revealed oppositely skewed patterns for targeted metabolites (primarily hydrophilic) and lipids (hydrophobic) in that polar metabolites showed a larger number of decreased species, while non-polar (lipids) had a greater number of increased species (>20% changed and/or raw p-value < 0.05). The top three ...
Traditionally, the prevention and management of chronic complications in individuals with type... more Traditionally, the prevention and management of chronic complications in individuals with type 1 (T1D) and type 2 diabetes (T2D) has been focused on of nephropathy, retinopathy, neuropathy, and atherosclerotic cardiovascular disease (including ischemic heart disease, stroke or peripheral vascular disease) (1). However, heart failure (HF) has been recognized as a common complication of diabetes, with a prevalence of up to 22% in individuals with diabetes and increasing incidence rates (2–4). This recognition stems in part from trials focused on cardiovascular safety of newer drugs to treat diabetes. Data also suggest HF may develop in individuals with diabetes even in the absence of hypertension, coronary heart disease, or valvular heart disease and, as such, represents a major cardiovascular complication in this vulnerable population (5). Given that during the past decade, the prevalence of diabetes (particularly T2D) has risen by 30% globally (6) (with prevalence expected to inc...
Angiotensin II can cause oxidative stress and increased blood pressure that result in long term c... more Angiotensin II can cause oxidative stress and increased blood pressure that result in long term cardiovascular pathologies. Here we evaluated the contribution of cellular senescence to the effect of chronic exposure to low dose angiotensin II in a model that mimics long term tissue damage. We utilized the INK-ATTAC (p16Ink4a–Apoptosis Through Targeted Activation of Caspase 8) transgenic mouse model that allows for conditional elimination of p16Ink4a -dependent senescent cells by administration of AP20187. Angiotensin II treatment for 3 weeks induced ATTAC transgene expression in kidneys but not in lung, spleen and brain tissues. In the kidneys increased expression of ATM, p15 and p21 matched with angiotensin II induction of senescence-associated secretory phenotype genes MMP3, FGF2, IGFBP2, and tPA. Senescent cells in the kidneys were identified as endothelial cells by detection of GFP expressed from the ATTAC transgene and increased expression of angiopoietin 2 and von Willebrand F...
Journal of Clinical and Translational Science, 2021
IMPACT: This work may lead to new treatments for crystalline nephropathies. OBJECTIVES/GOALS: Thi... more IMPACT: This work may lead to new treatments for crystalline nephropathies. OBJECTIVES/GOALS: This study investigated obeticholic acid (OCALIVA ®) as a potential treatment for 2,8-dihydroxyadenine (2,8-DHA) nephropathy using a mouse model. The treatment was investigated in both sexes at two timepoints. METHODS/STUDY POPULATION: Male and female C57BL/6J mice (12 weeks of age) were fed chow (Research Diets D19120401i) or chow admixed with adenine (0.2% w/w) ad lib for either 3.5 or 7 weeks. Mice were treated with either vehicle (corn oil) or obeticholic acid (10 mg/kg BW) by gavage 5 days per week. Each of the 16 combinations of sex/diet/timepoint/treatment groups had an n = 6 (96 mice in total). Food and body weights were measured twice per week, and 24-hour urines were collected prior to euthanasia. Serum and organs were collected and processed for biochemical and histopathological analyses. RESULTS/ANTICIPATED RESULTS: At both the 3.5-week and 7-week timepoints, dietary adenine rob...
Journal of the American Society of Nephrology, 1998
Administration of pharmacologic doses of glucocorticoid in vivo increases renal proximal tubule a... more Administration of pharmacologic doses of glucocorticoid in vivo increases renal proximal tubule apical membrane Na/H exchange and decreases Na/PO4 cotransport activity (1). Current data suggest that the NHE-3 and NaPi-2 proteins mediate significant fractions of proximal tubule apical membrane Na/H exchange and Na/PO4 cotransport, respectively. This study examines whether glucocorticoid excess or deficiency affects NHE-3 and NaPi-2 protein abundance and the intrarenal distribution of these transporters. Protein abundance of NHE-3 and NaPi-2 in control rats was compared to rats rendered glucocorticoid-deficient by bilateral adrenalectomy, and to rats receiving pharmacologic doses of dexamethasone using immunoblots and immunohistochemistry. Adrenalectomy had modest effects on NHE-3 protein abundance, but dexamethasone administration to either adrenalectomized or sham-operated rats significantly increased NHE-3 protein abundance in both the proximal tubule and thick ascending limb, but ...
Type 1 Natural Killer T-cells (NKT1 cells) play a critical role in mediating hepatic ischemia-rep... more Type 1 Natural Killer T-cells (NKT1 cells) play a critical role in mediating hepatic ischemia-reperfusion injury (IRI). Although hepatic steatosis is a major risk factor for preservation type injury, how NKT cells impact this is understudied. Given NKT1 cell activation by phospholipid ligands recognized presented by CD1d, we hypothesized that NKT1 cells are key modulators of hepatic IRI because of the increased frequency of activating ligands in the setting of hepatic steatosis. We first demonstrate that IRI is exacerbated by a high-fat diet (HFD) in experimental murine models of warm partial ischemia. This is evident in the evaluation of ALT levels and Phasor-Fluorescence Lifetime (Phasor-FLIM) Imaging for glycolytic stress. Polychromatic flow cytometry identified pronounced increases in CD45+CD3+NK1.1+NKT1 cells in HFD fed mice when compared to mice fed a normal diet (ND). This observation is further extended to IRI, measuring ex vivo cytokine expression in the HFD and ND. Much hi...
HIV disease remains prevalent in the USA and chronic kidney disease remains a major cause of morb... more HIV disease remains prevalent in the USA and chronic kidney disease remains a major cause of morbidity in HIV-1-positive patients. Host double-stranded RNA (dsRNA)-activated protein kinase (PKR) is a sensor for viral dsRNA, including HIV-1. We show that PKR inhibition by compound C16 ameliorates the HIV-associated nephropathy (HIVAN) kidney phenotype in the Tg26 transgenic mouse model, with reversal of mitochondrial dysfunction. Combined analysis of single-nucleus RNA-seq and bulk RNA-seq data revealed that oxidative phosphorylation was one of the most downregulated pathways and identified signal transducer and activator of transcription (STAT3) as a potential mediating factor. We identified in Tg26 mice a novel proximal tubular cell cluster enriched in mitochondrial transcripts. Podocytes showed high levels of HIV-1 gene expression and dysregulation of cytoskeleton-related genes; and these cells dedifferentiated and were lost from the glomerular tuft. In injured proximal tubules, c...
All forms of progressive renal diseases develop a final pathway of tubulointerstitial fibrosis an... more All forms of progressive renal diseases develop a final pathway of tubulointerstitial fibrosis and glomerulosclerosis. Renal fibrosis is usually quantified using histological staining, a process that is time-consuming and pathologist dependent. Here we develop a fast and operator-independent method to measure fibrosis utilizing the murine unilateral ureteral obstruction model which manifests a time-dependent fibrotic increase in obstructed kidneys while the contralateral kidneys are used as controls. After ureteral obstruction, kidneys were analyzed at 7, 14, and 21 days. Fibrosis was quantified using fluorescence lifetime imaging (FLIM) and second harmonic generation (SHG) in a Deep Imaging via Enhanced photon Recovery deep tissue imaging microscope. This microscope was developed for deep tissue along with second and third harmonic generation imaging and has extraordinary sensitivity toward harmonic generation. SHG data suggest the presence of more fibrillar collagen in the obstruc...
Sodium glucose cotransporter, type 2 inhibitors, such as Empagliflozin, are protective of the kid... more Sodium glucose cotransporter, type 2 inhibitors, such as Empagliflozin, are protective of the kidneys by unclear mechanisms. Our aim was to determine how Empagliflozin affected kidney cortical metabolome and lipidome in mice. Adult male TALLYHO mice (prone to obesity) were treated with a high-milk-fat diet, or this diet containing Empagliflozin (0.01%), for 8 weeks. Targeted and untargeted metabolomics and lipidomics were conducted on kidney cortex by liquid chromatography followed by tandem mass-spectroscopy. Metabolites were statistically analyzed by MetaboAnalyst 5.0, LipidSig (lipid species only) and/or CEU Mass Mediator (untargeted annotation). In general, volcano plotting revealed oppositely skewed patterns for targeted metabolites (primarily hydrophilic) and lipids (hydrophobic) in that polar metabolites showed a larger number of decreased species, while non-polar (lipids) had a greater number of increased species (>20% changed and/or raw p-value < 0.05). The top three ...
Traditionally, the prevention and management of chronic complications in individuals with type... more Traditionally, the prevention and management of chronic complications in individuals with type 1 (T1D) and type 2 diabetes (T2D) has been focused on of nephropathy, retinopathy, neuropathy, and atherosclerotic cardiovascular disease (including ischemic heart disease, stroke or peripheral vascular disease) (1). However, heart failure (HF) has been recognized as a common complication of diabetes, with a prevalence of up to 22% in individuals with diabetes and increasing incidence rates (2–4). This recognition stems in part from trials focused on cardiovascular safety of newer drugs to treat diabetes. Data also suggest HF may develop in individuals with diabetes even in the absence of hypertension, coronary heart disease, or valvular heart disease and, as such, represents a major cardiovascular complication in this vulnerable population (5). Given that during the past decade, the prevalence of diabetes (particularly T2D) has risen by 30% globally (6) (with prevalence expected to inc...
Angiotensin II can cause oxidative stress and increased blood pressure that result in long term c... more Angiotensin II can cause oxidative stress and increased blood pressure that result in long term cardiovascular pathologies. Here we evaluated the contribution of cellular senescence to the effect of chronic exposure to low dose angiotensin II in a model that mimics long term tissue damage. We utilized the INK-ATTAC (p16Ink4a–Apoptosis Through Targeted Activation of Caspase 8) transgenic mouse model that allows for conditional elimination of p16Ink4a -dependent senescent cells by administration of AP20187. Angiotensin II treatment for 3 weeks induced ATTAC transgene expression in kidneys but not in lung, spleen and brain tissues. In the kidneys increased expression of ATM, p15 and p21 matched with angiotensin II induction of senescence-associated secretory phenotype genes MMP3, FGF2, IGFBP2, and tPA. Senescent cells in the kidneys were identified as endothelial cells by detection of GFP expressed from the ATTAC transgene and increased expression of angiopoietin 2 and von Willebrand F...
Journal of Clinical and Translational Science, 2021
IMPACT: This work may lead to new treatments for crystalline nephropathies. OBJECTIVES/GOALS: Thi... more IMPACT: This work may lead to new treatments for crystalline nephropathies. OBJECTIVES/GOALS: This study investigated obeticholic acid (OCALIVA ®) as a potential treatment for 2,8-dihydroxyadenine (2,8-DHA) nephropathy using a mouse model. The treatment was investigated in both sexes at two timepoints. METHODS/STUDY POPULATION: Male and female C57BL/6J mice (12 weeks of age) were fed chow (Research Diets D19120401i) or chow admixed with adenine (0.2% w/w) ad lib for either 3.5 or 7 weeks. Mice were treated with either vehicle (corn oil) or obeticholic acid (10 mg/kg BW) by gavage 5 days per week. Each of the 16 combinations of sex/diet/timepoint/treatment groups had an n = 6 (96 mice in total). Food and body weights were measured twice per week, and 24-hour urines were collected prior to euthanasia. Serum and organs were collected and processed for biochemical and histopathological analyses. RESULTS/ANTICIPATED RESULTS: At both the 3.5-week and 7-week timepoints, dietary adenine rob...
Journal of the American Society of Nephrology, 1998
Administration of pharmacologic doses of glucocorticoid in vivo increases renal proximal tubule a... more Administration of pharmacologic doses of glucocorticoid in vivo increases renal proximal tubule apical membrane Na/H exchange and decreases Na/PO4 cotransport activity (1). Current data suggest that the NHE-3 and NaPi-2 proteins mediate significant fractions of proximal tubule apical membrane Na/H exchange and Na/PO4 cotransport, respectively. This study examines whether glucocorticoid excess or deficiency affects NHE-3 and NaPi-2 protein abundance and the intrarenal distribution of these transporters. Protein abundance of NHE-3 and NaPi-2 in control rats was compared to rats rendered glucocorticoid-deficient by bilateral adrenalectomy, and to rats receiving pharmacologic doses of dexamethasone using immunoblots and immunohistochemistry. Adrenalectomy had modest effects on NHE-3 protein abundance, but dexamethasone administration to either adrenalectomized or sham-operated rats significantly increased NHE-3 protein abundance in both the proximal tubule and thick ascending limb, but ...
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Papers by Moshe Levi