Medicinal and Pharmaceutical Chemistry Commons^™

Ho-1 Upregulation Attenuates Adipocyte Dysfunction, Obesity, And Isoprostane Levels In Mice Fed High Fructose Diets, Zeid Khitan, Mohit Harsh, Komal Sodhi, Joseph Shapiro, Nader Abraham

Nader G. Abraham

Background. Fructose metabolism is an unregulated metabolic pathway and excessive fructose consumption is known to activate ROS.HO-1 is a potent antioxidant gene that plays a key role in decreasing ROS and isoprostanes.We examinedwhether the fructosemediated increase in adipocyte dysfunction involves an increase in isoprostanes and that pharmacological induction ofHO-1would decrease both isoprostane levels and adipogenesis. Methods and Results. We examined the effect of fructose, on adipogenesis in human MSCs in the presence and absence of CoPP, an inducer of HO-1. Fructose increased adipogenesis and the number of large lipid droplets while decreasing the number of small lipid droplets (𝑃 < 0.05). …

Grp78/Bip Is Involved In Ouabain-Induced Endocytosis Of The Na/K-Atpase In Llc-Pk1 Cells, Riad Kesiry, Jiang Liu

Jiang Liu

We have demonstrated that ouabain causes dose- and time-dependent decreases both in 86Rb+ uptake and plasmalemmal Na/K-ATPase content of LLC-PK1 cells, which is related to ouabain-induced endocytosis of plasmalemmal Na/K-ATPase in LLC-PK1 cells through a clathrin-dependent mechanism. GRP78/BiP is a resident protein of the endoplasmic reticulum (ER) and acts as a molecular chaperone. Recently, several studies have shown that GRP78/BiP is also expressed on the cell surface and forms heterogeneous, high molecular weight complexes with other proteins. To identify the proteins that are possibly involved in ouabain-induced endocytosis of the Na/K-ATPase in LLC-PK1 cells, we separated and identified endosomal proteins …

Ouabain And Insulin Induce Sodium Pump Endocytosis In Renal Epithelium, Shalini Gupta, Yanling Yan, Deepak Malhotra, Jiang Liu, Zijian Xie, Sonia Najjar, Joseph Shapiro

Jiang Liu

Cardiotonic steroids signaling through the basolateral sodium pump (Na/K-ATPase) have been shown to alter renal salt handling in intact animals. Because the relationship between renal salt handling and blood pressure is a key determinant of hypertension, and patients with insulin resistance are frequently hypertensive, we chose to examine whether there might be competition for resources necessary for receptor-mediated endocytosis. In LLC-PK1 cells, the Na/K-ATPase-α1 and carcinoembryonic antigen cell adhesion molecule 1, a plasma membrane protein that promotes receptor-mediated endocytosis, colocalized in the plasma membranes and translocated to the intracellular region in response to ouabain. Either ouabain or insulin alone caused …

Effects Of Cardiac Glycosides On Sodium Pump Expression And Function In Llc-Pk1 And Mdck Cells, Jiang Liu, Sankaridrug Periyasamy, William Gunning, Olga Fedorova, Alexi Bagrov, Deepak Malhorta, Zijian Xie, Joseph Shapiro

Zijian Xie

Background: The decreases in proximal tubule sodium reabsorption seen with chronic renal failure and volume expansion have been ascribed to circulating digitalis-like substances (DLS). However, the circulating concentrations of DLS do not acutely inhibit the sodium pump to a degree consistent with the observed changes in proximal tubule sodium reabsorption.

Methods: We examined how cell lines that simulated proximal (LLC-PK1) and distal tubule (MDCK) cells responded to acute (30 min) and long-term (up to 12 hours) Na⁺,K⁺-ATPase inhibition with DLS.

Results: In LLC-PK1, but not MDCK cells, low concentrations of ouabain decreased ⁸⁶Rb uptake profoundly …

Ouabain Induces Endocytosis Of Plasmalemmal Na/K-Atpase In Llc-Pk1 Cells By A Clathrin-Dependent Mechanism, Jiang Liu, Riad Kesiry, Sankaridrug Periyasamy, Deppak Malhorta, Zijian Xie, Joseph Shapiro

Zijian Xie

Background: We have demonstrated that ouabain causes dose- and time-dependent decreases in ⁸⁶Rb uptake in porcine proximal tubular (LLC-PK1) cells. The present study addresses the molecular mechanisms involved in this process.

Methods: Studies were performed with cultured LLC-PK1 and Src family kinase deficient (SYF) cells.

Results: We found that 50 nmol/L ouabain applied to the basal, but not apical, aspect for 12 hours caused decreases in the plasmalemmal Na/K-ATPase. This loss of plasmalemmal Na/K-ATPase reverses completely within 12 to 24 hours after removal of ouabain. Ouabain also increased the Na/K-ATPase content in both early and late endosomes, activated phosphatidylinositol …

Cardiac Glycoside Downregulates Nhe3 Activity And Expression In Llc-Pk1 Cells, Shadi Oweis, Liang Wu, Pawel Kiela, Deepak Malhotra, Fayez Ghishan, Zijian Xie, Joseph Shapiro, Jiang Liu

Zijian Xie

Ouabain, a cardiotonic steroid and a specific inhibitor of the Na+-K+-ATPase, has been shown to significantly inhibit transcellular Na+ transport without altering the intracellular Na+ concentration ([Na+]i) in the epithelial cells derived from the renal proximal tubules. We therefore studied whether ouabain affects the activity and expression of Na_/H_ exchanger isoform 3 (NHE3) representing the major route of apical Na_ reabsorption in LLC-PK1 cells. Chronic basolateral, but not apical, exposure to low-concentration ouabain (50 and 100 nM) did not change [Na+]i but significantly reduced NHE3 activity, NHE3 protein, and mRNA wxpression. Inhibition of c-Src or phosphoinositide 3-kinase (PI3K) with PP2 …

Ouabain-Induced Endocytosis Of The Plasmalemmal Na/K-Atpase In Llc-Pk1 Cells Requires Caveolin-1, Jiang Liu, Man Liang, Lijun Liu, Deepak Malhotra, Zijian Xie, Joseph Shapiro

Zijian Xie

Background: We have demonstrated that ouabain causes dose- and time-dependent decreases in 86Rb uptake in pig renal proximal tubule cell line (LLC-PK1) cells; and ouabain induces endocytosis of plasmalemmal Na/K-ATPase in LLC-PK1 cells in a clathrin-dependent pathway. Our data also suggest a role of endocytosis in both ouabain-induced signal transduction and proximal tubule sodium handling. The present study addresses the molecular mechanisms involved in this process. Methods: Studies were performed with cultured LLC-PK1 and a stable-expressed caveolin-1 knockdown LLC-PK1 cell line by SiRNA method. Results: In wild-type LLC-PK1 cells, depletion of cholesterol by methyl -cyclodextrin reduced ouabain-induced accumulation of Na/K-ATPase …

Ouabain And Insulin Induce Sodium Pump Endocytosis In Renal Epithelium, Shalini Gupta, Yanling Yan, Deepak Malhotra, Jiang Liu, Zijian Xie, Sonia Najjar, Joseph Shapiro

Zijian Xie

Cardiotonic steroids signaling through the basolateral sodium pump (Na/K-ATPase) have been shown to alter renal salt handling in intact animals. Because the relationship between renal salt handling and blood pressure is a key determinant of hypertension, and patients with insulin resistance are frequently hypertensive, we chose to examine whether there might be competition for resources necessary for receptor-mediated endocytosis. In LLC-PK1 cells, the Na/K-ATPase-α1 and carcinoembryonic antigen cell adhesion molecule 1, a plasma membrane protein that promotes receptor-mediated endocytosis, colocalized in the plasma membranes and translocated to the intracellular region in response to ouabain. Either ouabain or insulin alone caused …

Ouabain-Stimulated Trafficking Regulation Of The Na/K-Atpase And Nhe3 In Renal Proximal Tubule Cells, Yanling Yan, Steven Haller, Anna Shapiro, Nathan Malhotra, Jiang Tian, Zijian Xie, Deepak Malhotra, Joseph Shapiro, Jiang Liu

Zijian Xie

We have demonstrated that ouabain regulates protein trafficking of the Na/K-ATPase a1 subunit and NHE3 (Na/H exchanger, isoform 3) via ouabain-activated Na/K-ATPase signaling in porcine LLC-PK1 cells. To investigate whether this mechanism is species-specific, ouabain-induced regulation of the a1 subunit and NHE3 as well as transcellular 22Na? transport were compared in three renal proximal tubular cell lines (human HK-2, porcine LLC-PK1, and AAC-19 originated from LLC-PK1 in which the pig a1 was replaced by ouabain-resistant rat a1). Ouabain-induced inhibition of transcellular 22Na? transport is due to an ouabain-induced redistribution of the a1 subunit and NHE3. In LLC-PK1 cells, ouabain also …

Salt Loading Induces Redistribution Of The Plasmalemmal Na/K-Atpase In Proximal Tubule Cells, Sankaridrug Periyasamy, Jiang Liu, Feras Tanta, Besher Kabak, Brent Wakefield, Deepak Malhorta, David Kennedy, Alaa Nodoor, Olga Fedorova, William Gunning, Zijian Xie, Alexi Bagrov, Joseph Shapiro

Zijian Xie

Background: We have reported that digitalis-like substances (cardiotonic steroids), including marinobufagenin (MBG), induce endocytosis of the plasmalemmal Na/K-ATPase in LLCPK1 cells. The current report addresses the potential relevance of plasmalemmal Na/K-ATPase redistribution to in vivo salt handling. Methods: Male Sprague-Dawley rats were given 1 week of a high salt (4.0% NaCl) or normal salt (0.4% NaCl) diet. Urinary sodium excretion, as well as MBG excretion, was monitored, and proximal tubules were isolated using a Percoll gradient method. Tubular 86Rb uptake, Na/K-ATPase enzymatic activity, and Na/K-ATPase a1 subunit density were determined. Results: The high salt diet increased urinary sodium (17.8 ± …

Ouabain Induces Endocytosis Of Plasmalemmal Na/K-Atpase In Llc-Pk1 Cells By A Clathrin-Dependent Mechanism, Jiang Liu, Riad Kesiry, Sankaridrug Periyasamy, Deppak Malhorta, Zijian Xie, Joseph Shapiro

Jiang Liu

Background: We have demonstrated that ouabain causes dose- and time-dependent decreases in ⁸⁶Rb uptake in porcine proximal tubular (LLC-PK1) cells. The present study addresses the molecular mechanisms involved in this process.

Methods: Studies were performed with cultured LLC-PK1 and Src family kinase deficient (SYF) cells.

Results: We found that 50 nmol/L ouabain applied to the basal, but not apical, aspect for 12 hours caused decreases in the plasmalemmal Na/K-ATPase. This loss of plasmalemmal Na/K-ATPase reverses completely within 12 to 24 hours after removal of ouabain. Ouabain also increased the Na/K-ATPase content in both early and late endosomes, activated phosphatidylinositol …

Ouabain-Induced Endocytosis And Signal Transduction Of The Na/K-Atpase, Jiang Liu

Jiang Liu

Na/K-ATPase can function as a signal transducer as well as an energy transducing ion pump. Cardiac glycosides (including ouabain and marinobufagin, MBG) are a new class of steroid hormones. Ouabain-activated signaling pathways lead to the induction of some early response proto-oncogenes, activation of transcription factors, and cardiac hypertrophy. Low concentration of ouabain also induced endocytosis of the Na/K-ATPase and compartmentalization of some signaling molecules (e.g. c-Src, EGFR, and p42/44 MAPKs) into clathrin-coated pits, early and late endosomes. Ouabain-induced endocytosis of the Na/K-ATPase depends on the activation of Src kinase, clathrin-coated pits formation, and caveolin-1 (the major component of caveolae). Moreover, …

Effects Of Cardiac Glycosides On Sodium Pump Expression And Function In Llc-Pk1 And Mdck Cells, Jiang Liu, Sankaridrug Periyasamy, William Gunning, Olga Fedorova, Alexi Bagrov, Deepak Malhorta, Zijian Xie, Joseph Shapiro

Jiang Liu

Background: The decreases in proximal tubule sodium reabsorption seen with chronic renal failure and volume expansion have been ascribed to circulating digitalis-like substances (DLS). However, the circulating concentrations of DLS do not acutely inhibit the sodium pump to a degree consistent with the observed changes in proximal tubule sodium reabsorption.

Methods: We examined how cell lines that simulated proximal (LLC-PK1) and distal tubule (MDCK) cells responded to acute (30 min) and long-term (up to 12 hours) Na⁺,K⁺-ATPase inhibition with DLS.

Results: In LLC-PK1, but not MDCK cells, low concentrations of ouabain decreased ⁸⁶Rb uptake profoundly …

Cardiac Glycoside Downregulates Nhe3 Activity And Expression In Llc-Pk1 Cells, Shadi Oweis, Liang Wu, Pawel Kiela, Deepak Malhotra, Fayez Ghishan, Zijian Xie, Joseph Shapiro, Jiang Liu

Jiang Liu

Ouabain, a cardiotonic steroid and a specific inhibitor of the Na+-K+-ATPase, has been shown to significantly inhibit transcellular Na+ transport without altering the intracellular Na+ concentration ([Na+]i) in the epithelial cells derived from the renal proximal tubules. We therefore studied whether ouabain affects the activity and expression of Na_/H_ exchanger isoform 3 (NHE3) representing the major route of apical Na_ reabsorption in LLC-PK1 cells. Chronic basolateral, but not apical, exposure to low-concentration ouabain (50 and 100 nM) did not change [Na+]i but significantly reduced NHE3 activity, NHE3 protein, and mRNA wxpression. Inhibition of c-Src or phosphoinositide 3-kinase (PI3K) with PP2 …

Ouabain-Induced Endocytosis Of The Plasmalemmal Na/K-Atpase In Llc-Pk1 Cells Requires Caveolin-1, Jiang Liu, Man Liang, Lijun Liu, Deepak Malhotra, Zijian Xie, Joseph Shapiro

Jiang Liu

Background: We have demonstrated that ouabain causes dose- and time-dependent decreases in 86Rb uptake in pig renal proximal tubule cell line (LLC-PK1) cells; and ouabain induces endocytosis of plasmalemmal Na/K-ATPase in LLC-PK1 cells in a clathrin-dependent pathway. Our data also suggest a role of endocytosis in both ouabain-induced signal transduction and proximal tubule sodium handling. The present study addresses the molecular mechanisms involved in this process. Methods: Studies were performed with cultured LLC-PK1 and a stable-expressed caveolin-1 knockdown LLC-PK1 cell line by SiRNA method. Results: In wild-type LLC-PK1 cells, depletion of cholesterol by methyl -cyclodextrin reduced ouabain-induced accumulation of Na/K-ATPase …

Ouabain-Stimulated Trafficking Regulation Of The Na/K-Atpase And Nhe3 In Renal Proximal Tubule Cells, Yanling Yan, Steven Haller, Anna Shapiro, Nathan Malhotra, Jiang Tian, Zijian Xie, Deepak Malhotra, Joseph Shapiro, Jiang Liu

Jiang Liu

We have demonstrated that ouabain regulates protein trafficking of the Na/K-ATPase a1 subunit and NHE3 (Na/H exchanger, isoform 3) via ouabain-activated Na/K-ATPase signaling in porcine LLC-PK1 cells. To investigate whether this mechanism is species-specific, ouabain-induced regulation of the a1 subunit and NHE3 as well as transcellular 22Na? transport were compared in three renal proximal tubular cell lines (human HK-2, porcine LLC-PK1, and AAC-19 originated from LLC-PK1 in which the pig a1 was replaced by ouabain-resistant rat a1). Ouabain-induced inhibition of transcellular 22Na? transport is due to an ouabain-induced redistribution of the a1 subunit and NHE3. In LLC-PK1 cells, ouabain also …

Salt Loading Induces Redistribution Of The Plasmalemmal Na/K-Atpase In Proximal Tubule Cells, Sankaridrug Periyasamy, Jiang Liu, Feras Tanta, Besher Kabak, Brent Wakefield, Deepak Malhorta, David Kennedy, Alaa Nodoor, Olga Fedorova, William Gunning, Zijian Xie, Alexi Bagrov, Joseph Shapiro

Jiang Liu

Background: We have reported that digitalis-like substances (cardiotonic steroids), including marinobufagenin (MBG), induce endocytosis of the plasmalemmal Na/K-ATPase in LLCPK1 cells. The current report addresses the potential relevance of plasmalemmal Na/K-ATPase redistribution to in vivo salt handling. Methods: Male Sprague-Dawley rats were given 1 week of a high salt (4.0% NaCl) or normal salt (0.4% NaCl) diet. Urinary sodium excretion, as well as MBG excretion, was monitored, and proximal tubules were isolated using a Percoll gradient method. Tubular 86Rb uptake, Na/K-ATPase enzymatic activity, and Na/K-ATPase a1 subunit density were determined. Results: The high salt diet increased urinary sodium (17.8 ± …

Acid-Suppressing Agents And Risk For Clostridium Difficile Infection In Pediatric Patients, Katelyn Brown, Chad Knoderer, Kristen Nichols, Ashley Crumby

Chad A. Knoderer

Background. Acid-suppressing agents have been associated with increased Clostridium difficile infection (CDI) in adults. The objective of this study was to evaluate the association of acid-suppressing therapy with the development of CDI in the pediatric population. Methods. This was a retrospective case-control study. Children aged 1 through 17 years with a positive C difficile polymerase chain reaction (PCR) result obtained between June 1, 2008, and June 1, 2012, were randomly matched to a control population selected from patients with negative PCR. Results. A total of 458 children were included. No difference was observed in acid-suppressive therapy prior to PCR in …

Teratogen Use In Women Of Childbearing Potential: An Intervention Study, Karie Morical-Kline, Alison Walton, Tracey Guildenbecher

Alison M. Walton

Purpose: The purpose of this study was to quantify the number of women of childbearing potential who are prescribed angiotensin-converting enzyme inhibitor (ACE inhibitor), angiotensin receptor blocker (ARB), or HMG-coenzyme A reductase inhibitor (statin) and to determine the number of documented teratogenic risk discussions (risk documentation) before and after educational interventions. Methods: The institutional review board–approved retrospective chart review included female patients ages 15 to 45 years who were prescribed an ACE inhibitor, ARB, or statin between January 1, 2007, and March 1, 2009. Exclusion criteria were tubal ligation and hysterectomy. A survey determined physician knowledge of teratogenic risks and …

Y-Site Compatibility Of Vancomycin And Piperacillin/ Tazobactam At Commonly Utilized Pediatric Concentrations, Kristen Nichols, Michael Demarco, Michael Vertin, Chad Knoderer

Chad A. Knoderer

Background: Vancomycin and piperacillin/tazobactam are common empiric antibiotics in hospitalized pediatric patients. Studies evaluating intravenous (IV) compatibility at various concentrations show inconsistent results. Objective: The objective of this study was to determine the Y-site compatibility of vancomycin 10 mg/mL and piperacillin/tazobactam 112.5 mg/mL. Methods: Vancomycin (10 g vial) was reconstituted using sterile water for injection (SWFI) and diluted with 5% dextrose in water (D5W) to a final concentration of 10 mg/mL in an evacuated IV bag. Piperacillin/tazobactam (40.5 g vial) was reconstituted and diluted with SWFI to a final concentration of 112.5 mg/mL (100 mg/mL piperacillin) in an evacuated IV …

Linezolid-Associated Thrombocytopenia In Children With Renal Impairment, Sara Jones, Kristen Nichols, Heather Deyoung, Elaine Cox, Chad Knoderer

Chad A. Knoderer

Impaired renal function has been associated with an increased risk of thrombocytopenia in adults receiving linezolid. Findings from this retrospective cohort demonstrate an association between thrombocytopenia and lower creatinine clearance in children receiving linezolid.

Linezolid-Associated Thrombocytopenia In Children With Renal Impairment, Sara Jones, Kristen Nichols, Heather Deyoung, Elaine Cox, Chad Knoderer

Kristen R. Nichols

Impaired renal function has been associated with an increased risk of thrombocytopenia in adults receiving linezolid. Findings from this retrospective cohort demonstrate an association between thrombocytopenia and lower creatinine clearance in children receiving linezolid.

Y-Site Compatibility Of Vancomycin And Piperacillin/ Tazobactam At Commonly Utilized Pediatric Concentrations, Kristen Nichols, Michael Demarco, Michael Vertin, Chad Knoderer

Kristen R. Nichols

Background: Vancomycin and piperacillin/tazobactam are common empiric antibiotics in hospitalized pediatric patients. Studies evaluating intravenous (IV) compatibility at various concentrations show inconsistent results. Objective: The objective of this study was to determine the Y-site compatibility of vancomycin 10 mg/mL and piperacillin/tazobactam 112.5 mg/mL. Methods: Vancomycin (10 g vial) was reconstituted using sterile water for injection (SWFI) and diluted with 5% dextrose in water (D5W) to a final concentration of 10 mg/mL in an evacuated IV bag. Piperacillin/tazobactam (40.5 g vial) was reconstituted and diluted with SWFI to a final concentration of 112.5 mg/mL (100 mg/mL piperacillin) in an evacuated IV …