Frank Sellke | Brown University (original) (raw)

Papers by Frank Sellke

Journal of nature and science, 2016

Calpains are a 15-member class of calcium activated nonlysosomal neutral proteases which are invo... more Calpains are a 15-member class of calcium activated nonlysosomal neutral proteases which are involved in a broad range of cellular function. Calpains are usually localized to the cytosol and within mitochondria. Calpastatin is an endogenous protein that specifically binds to and inhibits calpain. Overactivation of calpain has been implicated in a number of disease processes of the brain, eyes, heart, lungs, pancreas, kidneys, vascular system and skeletal muscle. Therefore, calpain may serve as a potential therapeutic target for a wide variety of disease processes. This review briefly outlines the current literature regarding the involvement of calpain overactivation in the pathogenesis of almost every organ in the body.

Surgery

Background-Moderate alcohol consumption is cardioprotective but the mechanism of action remains u... more Background-Moderate alcohol consumption is cardioprotective but the mechanism of action remains unclear. Nuclear Factor kappa-B (NFκB) regulates the expression of genes involved in inflammation, stress and apoptosis. We used a swine model of diet-induced metabolic syndrome to investigate the effects of red wine and vodka on NFκB signaling and cytokine activity in chronically ischemic myocardium. Methods-Yorkshire swine were given a high-fat diet for four weeks; an ameroid constrictor was then placed on the left circumflex artery. The high-fat diet was continued and the swine were divided into three groups for 7 weeks: hypercholesterolemic diet alone (Control, n=8), hypercholesterolemic diet with vodka (Vodka, n=8), and hypercholesterolemic diet with wine (Wine, n=8). Ischemic myocardium was analyzed by Western blot and cytokine array. Results-Administration of alcohol was associated with decreased expression of IKKα, IKKβ and p-IκBα in the ischemic myocardium compared to the control group. Alcohol administration demonstrated an increase in NFκB in the ischemic myocardium. Both wine and vodka demonstrated a significant decrease in leptin, IL-1α, IL-13, IL-15 and IFN-γ. Vodka demonstrated a significant decrease in phosphorylated BCL-2 and caspase-9. Conclusions-In ischemic myocardium, alcohol modulates the NFκB pathway, which may contribute to the adaptive response of tissues to the stress of ischemia. Furthermore, both wine and vodka decreased multiple pro-inflammatory cytokines. This study provides a mechanism by which alcohol may be cardioprotective in ischemic myocardium.

The Annals of thoracic surgery, Jan 18, 2017

We hypothesized that upregulation of inducible cyclooxygenase 2 (COX-2) contributes to altered co... more We hypothesized that upregulation of inducible cyclooxygenase 2 (COX-2) contributes to altered coronary arteriolar reactivity early after cardioplegic arrest and cardiopulmonary bypass (CP/CPB) in patients with diabetes mellitus who are undergoing cardiac surgery. The right atrial tissue samples of nondiabetes (ND), controlled diabetes (CDM), and uncontrolled diabetes (UDM) patients undergoing cardiac surgery were harvested before and after CP/CPB. Coronary arterioles (80 to 150 μm) were dissected from the harvested atrial tissue samples, cannulated, and pressurized. The changes in diameter were measured with video microscopy. The protein expression and localization of COX-1 and COX-2 were assayed by Western blot and immunohistochemistry. In the diabetes arterioles, bradykinin-induced relaxation response was inhibited by the selective COX-2 inhibitor NS398 at baseline (p < 0.05). This effect was more pronounced in UDM arterioles than CDM (p < 0.05). After CP/CPB, bradykinin-in...

Cardiovascular Research, 2017

There are conflicting reports on the role of reactive oxygen species (ROS) i.e. beneficial vs. ha... more There are conflicting reports on the role of reactive oxygen species (ROS) i.e. beneficial vs. harmful, in vascular endothelium. Here, we aim to examine whether duration of exposure to ROS and/or subcellular ROS levels are responsible for the apparently paradoxical effects of oxidants on endothelium.

Surgery, 2016

Introduction-Emerging data suggests a link between calpain activation and the enhanced inflammato... more Introduction-Emerging data suggests a link between calpain activation and the enhanced inflammatory response of the cardiovascular system. We hypothesize that calpain activation associates with altered inflammatory protein expression in correlation with the pro-inflammatory profile of the myocardium. Our pig hypercholesterolemic model with chronic myocardial ischemia will be treated with calpain inhibitors (CI) to establish their potentials to improve cardiac function. Methods-Yorkshire swine, fed a high cholesterol diet (HC) for four weeks, underwent placement of an ameroid constrictor on the left circumflex artery. Three weeks later animals received either: no drug, high cholesterol control group (CON; n= 8); a low dose CI (0.12 mg/kg; LCI, n= 9); or a high dose CI (0.25 mg/kg; HCI, n= 8). The HC and CI were continued for five weeks, after which the pig was euthanized. The left ventricular myocardial tissue (ischemic and non-ischemic) was harvested and analyzed for inflammatory proteins expression. Data was statistically analyzed via the Kruskal-Wallis and Dunn's post hoc test. Results-CI treatment coincides with increased expression of IKB-α and decreased expression of macrophages, NFkB, IL-1 and TNF-α in the ischemic myocardial tissue as compared to the control group. NFkB array revealed decreased expression of IRF5, JNK1/2, JNK2, CD18, NFkB p65, c-Rel, Sharpin, TNF R1, TNF R2 and DR5 in the ischemic myocardium of the HCI treated group compared to the control. Conclusions-Calpain activation in metabolic syndrome is a potential contributor to cardiac dysfunction in metabolic disorders with ischemic background. We suggest calpain inhibition down regulates NFkB signaling in the vessel walls which might be useful for improving myocardial blood flow in ischemic conditions.

Journal of the American Heart Association, Jan 24, 2015

Diabetes is associated with coronary arteriolar endothelial dysfunction. We investigated the role... more Diabetes is associated with coronary arteriolar endothelial dysfunction. We investigated the role of the small/intermediate (SKC a/IKC a) conductance of calcium-activated potassium channels in diabetes-related endothelial dysfunction. Coronary arterioles (80 to 150 μm in diameter) were dissected from discarded right atrial tissues of diabetic (glycosylated hemoglobin = 9.6±0.25) and nondiabetic patients (glycosylated hemoglobin 5.4±0.12) during coronary artery bypass graft surgery (n=8/group). In-vitro relaxation response of precontracted arterioles was examined in the presence of the selective SKC a/IKC a activator NS309 and other vasodilatory agents. The channel density and membrane potential of diabetic and nondiabetic endothelial cells was measured by using the whole cell patch-clamp technique. The protein expression and distribution of the SKC a/IKC a in the human myocardium and coronary arterioles was examined by Western blotting and immunohistochemistry. Our results indicate ...

Cardiovascular Risk Factors, 2012

The Annals of thoracic surgery, 2014

Chronic hyperglycemia has been associated with increased oxidative stress in skeletal muscle and ... more Chronic hyperglycemia has been associated with increased oxidative stress in skeletal muscle and sympathetic nerve dysfunction. We investigated the effect of chronic hyperglycemia on the myocardium of patients with uncontrolled diabetes (UD) compared with patients with well-controlled diabetes (CD) and patients without diabetes (ND) after cardioplegic cardiopulmonary bypass (CP/CPB) with acute intraoperative glycemic control. Atrial tissue and serum were collected from 47 patients (ND=18 with glycated hemoglobin [HbA1c] of 5.8±0.2; CD=8 with HbA1c of 6.1±0.1; with UD=21 with HbA1c=9.6±0.5) before and after CP/CPB for immunoblotting, protein oxidation assays, immunohistochemical evaluation, and microarray analysis. The uncontrolled group had increased total protein oxidation (p<0.05) and decreased levels of antioxidative enzyme manganese superoxide dismutase (MnSOD) (p<0.05) after CP/CPB compared with the controlled group. Collagen staining revealed increased fibrosis in patien...

The Journal of Thoracic and Cardiovascular Surgery, 2014

Autophagy is a cellular process by which damaged components are removed. Although autophagy can r... more Autophagy is a cellular process by which damaged components are removed. Although autophagy can result in cell death, when optimally regulated, it might be cardioprotective. Resveratrol is a naturally occurring polyphenol also believed to be cardioprotective. Using a clinically relevant swine model of metabolic syndrome, we investigated the effects of resveratrol on autophagy in the chronically ischemic myocardium. Methods: Yorkshire swine were fed a regular diet (n ¼ 7), a high cholesterol diet (n ¼ 7), or a high cholesterol diet with supplemental resveratrol (n ¼ 6). After 4 weeks, an ameroid constrictor was surgically placed on the left circumflex artery to induce chronic myocardial ischemia. The diets were continued another 7 weeks, and then the ischemic and nonischemic myocardium were harvested for protein analysis. Results: In the ischemic myocardium, a high cholesterol diet partly attenuated the autophagy, as determined by an increase in phosphorylated mammalian target of rapamycin (p-mTOR) and a decrease in p70 S6 kinase (P70S6K), lysosome-associated membrane protein (LAMP)-2, and autophagy-related gene 12-5 conjugate (ATG 12-5; P < .05). The addition of resveratrol blunted many of these changes, because the p-mTOR, P70S6K, and LAMP-2 levels were not significantly altered from those of the pigs fed a regular diet. Other autophagy markers were increased with a high cholesterol diet, including light chain 3A-II and beclin 1 (P < .05). In the nonischemic myocardium, beclin 1 was decreased in the high cholesterol-fed pigs (P <.05); otherwise no significant changes in protein expression were noted among the 3 groups. Conclusions: In the chronically ischemic myocardium, resveratrol partly reversed the effects of a high cholesterol diet on autophagy. This might be a mechanism by which resveratrol exerts its cardioprotective effects.

The Journal of Thoracic and Cardiovascular Surgery, 2010

Frontiers in Bioscience, 2009

Cardiovascular disease remains the leading cause of mortality in the industrialized world. Despit... more Cardiovascular disease remains the leading cause of mortality in the industrialized world. Despite advances in pharmacotherapy and catheter based interventions, coronary artery bypass grafting remains an essential therapeutic modality. The majority of coronary artery bypass operations, as well as other cardiac surgical procedures require the use of ischemic cardioplegic arrest and cardiopulmonary bypass, both of which result in iatrogenic injury to the vasculature and microcirculation. This injury can manifest as impaired vasorelaxation or vasoconstriction, depending upon the organ system involved, resulting in impaired tissue perfusion and the development of edema. Key to this dysfunction are changes in the following: nitric oxide signaling secondary to changes in eNOS and iNOS expression and activity, cyclooxygenase function with increases in proinflammatory COX-2 activity, alterations in Protein Kinase C and Mitogen Activated Protein Kinase signaling, and an increase in Vascular Endothelial Growth Factor expression increasing vascular permeability and dilatation. This review discusses our current understanding of cardioplegia and cardiopulmonary bypass induced changes in the vasculature, and therapeutic interventions aimed at modulating the altered signaling pathways.

Circulation, 2013

D iabetes mellitus is associated with increased risk of ischemic heart disease and increased morb... more D iabetes mellitus is associated with increased risk of ischemic heart disease and increased morbidity and mortality after open heart operations involving cardioplegia (CP) and cardiopulmonary bypass (CPB). 1,2 Previous studies have demonstrated that diabetes mellitus is associated with increased myocardial oxidative stress, attenuation of cell survival pathways, and induction of apoptosis in animal models. 3-6 Recent clinical trials have also shown that glycemic control significantly reduces microvascular complications, such as retinopathy, nephropathy, and peripheral arterial disease. 7,8 In addition, perioperative glucose control in some studies is associated with improved outcomes after coronary artery bypass graft surgery. 9 Recently, we found that poorly controlled diabetes mellitus is associated with microvascular dysfunction and worsens the recovery of arteriolar endothelial and smooth muscle function after CP/CPB. 10,11 However, whether diabetes mellitus, especially uncontrolled type 2 diabetes mellitus (UDM), affects apoptosis and apoptosisrelated signaling after CP/CPB has not been investigated. The goal of this research is to investigate the effect of diabetes mellitus on cardioplegic arrest and reperfusion (CP/Rep)-induced changes in myocardial apoptosis and apoptosis-related signaling. Specifically, this study was designed to directly test the effect of diabetes mellitus and CP/Rep on the development of apoptosis and apoptosis-related protein expression/localization in human atrial tissues harvested from patients with coronary artery bypass graft surgery. Methods Human Subjects and Tissue Harvesting Samples of right atrial appendage were harvested from patients undergoing coronary artery bypass graft surgery before and after exposure of the heart to blood cardioplegia and short-term reperfusion under Background-We investigated the effects of cardioplegic arrest and reperfusion (CP/Rep) on myocardial apoptosis and key apoptotic mediators, such as apoptosis-inducing factor, caspase 3, caspase 8, caspase 9, poly(adenosine diphosphateribose) polymerase, B-cell lymphoma 2 (Bcl-2) family proteins, and protein kinase C (PKC), in uncontrolled type 2 diabetic, controlled type 2 diabetic, and nondiabetic patients. Methods and Results-Right atrial tissue was harvested pre-and post-CP/Rep from uncontrolled type 2 diabetic patients (hemoglobin A1c=9.6±0.25), controlled type 2 diabetic patients (hemoglobin A1c=6.5±0.15), and nondiabetic patients (hemoglobin A1c=5.4±0.12) undergoing coronary artery bypass grafting (n=8/group). Terminal deoxynucleotidyl transferase dUTP nick-end labeling staining was used for the identification of apoptotic cells. Total and modified apoptosis-inducing factor, Bcl-2 family proteins, phospho-PKC-α, phospho-PKC-β1, and poly(adenosine diphosphate-ribose) polymerase were quantified by immunoblotting or immunohistochemistry. At baseline, the number of apoptotic cells and expression of total apoptosis-inducing factor, Bcl-2, Bak, and Bax in the pre-CP/Rep atrial tissue from uncontrolled type 2 diabetic patients were significantly increased compared with those of nondiabetic or controlled type 2 diabetic patients (P<0.05). After CP/Rep, the amount of apoptotic cells, apoptosis-inducing factor, phospho-Bad, phospho-PKC-α, phospho-PKC-β1, and cleaved poly(adenosine diphosphate-ribose) polymerase in post-CP/Rep atrial tissue were increased in all 3 groups compared with pre-CP/Rep. These increases after CP/Rep were more pronounced in the uncontrolled type 2 diabetic group. In addition, there were significant increases in the expression of cleaved caspase 8 and caspase 9 in the basal and post-CP/Rep atrium of uncontrolled type 2 diabetic group compared with nondiabetic or controlled type 2 diabetic group. Conclusions-Uncontrolled diabetes mellitus is associated with increases in myocardial apoptosis and expression of key apoptosis mediators at baseline and in the setting of CP/Rep. (Circulation. 2013;128[suppl 1]:S144-S151.

Circulation, 2010

Background— We investigated the efficacy of novel thrombin fragment TP508 on ischemia-reperfusion... more Background— We investigated the efficacy of novel thrombin fragment TP508 on ischemia-reperfusion injury using a porcine model of type 1 diabetes mellitus. Methods and Results— Alloxan-induced diabetic male Yucatan swine underwent 60 minutes of mid-left anterior descending coronary artery occlusion, followed by 120 minutes of reperfusion. Fifty minutes into ischemia, animals received either placebo (DM; n=8) or TP508 as a bolus of 1 mg/kg followed by infusion at 2.5 mg/kg per hour (DMT; n=8). Hemodynamic parameters and myocardial function were monitored. Monastryl blue/triphenyl tetrazolium chloride staining was used to assess sizes of the areas at risk and infarction. Coronary microvascular reactivity was measured and expression of cell survival and proapoptotic proteins quantified. Preoperative serum glucose values were similar between groups (309±57 mg/dL in DM versus 318±67 mg/dL in DMT; P =0.92). Infarct size was smaller in the TP508-treated group (5.3±1.9% in DMT versus 19.4±5...

Circulation, 2011

Background— Cardioplegia and cardiopulmonary bypass (CP/CPB) subjects myocardium to complex injur... more Background— Cardioplegia and cardiopulmonary bypass (CP/CPB) subjects myocardium to complex injurious stimuli that can result in cardiomyocyte and vascular contractile abnormalities. Rottlerin, originally identified as a delta-protein kinase C inhibitor, has a number of known additional effects that may be beneficial in the setting of CP/CPB. We tested the hypothesis that rottlerin mitigates deleterious effects associated with CP/CPB. Methods and Results— Langendorff-perfused isolated rat hearts were subjected to 2 hours intermittent cold (10°C) CP (St Thomas II) followed by 30 minutes normothermic reperfusion. CP was delivered every 30 minutes for 1 minute. Hearts were treated with rottlerin 1 μmol/L (CP+R) (n=7) or without rottlerin (CP) (n=9), and the BK Ca++ channel inhibitor paxilline 100 nmol/L was supplied in the CP. Hearts constantly perfused with KHB served as controls (n=6). Baseline parameters of cardiac function were similar between groups. CP resulted in reduced cardiac...

Cardiovascular Research, 2006

Objective: Cyclooxygenase-2 (COX-2) is induced by hypoxic stimuli and is also involved in the pro... more Objective: Cyclooxygenase-2 (COX-2) is induced by hypoxic stimuli and is also involved in the process of angiogenesis. We previously demonstrated that vascular endothelial growth factor (VEGF) is one of the principal factors produced by hypoxic myocytes and is responsible for the induction of COX-2 expression in endothelial cells. Yet the signaling pathways by which VEGF modulates COX-2 gene expression are still less well defined. We therefore examined the regulation of VEGF-induced COX-2 expression by the mitogen-activated protein kinase (MAPK) family in endothelial cells. Methods and results: Human umbilical vascular endothelial cells (HUVECs) were incubated with U0126 (ERK1/2 inhibitor, 10 AM), SB203580 (p38 inhibitor, 20 AM), and SP600125 (JNK inhibitor, 20 AM), as well as the COX-2 selective inhibitor, NS398, for 1 h before treating with VEGF (20 ng/ml). COX-2 expression induced by VEGF at both mRNA and protein levels was significantly inhibited by selective p38 and JNK inhibitors but not by the ERK1/2 inhibitor. The phosphorylation of p38 and JNK kinases was observed as early as 5 min in HUVECs after VEGF stimulation. Furthermore, the biological significance of the COX-2 gene in endothelial cells was examined by over-expressing or knocking down COX-2 gene expression. 3 H-Thymidine incorporation and Matrigel techniques were used to determine cell proliferation and vascular structure formation. VEGF-induced cell proliferation was significantly reduced when HUVECs were either pre-treated with NS398 (21.52 T 3.6%) or transfected with COX-2 siRNA (34.12 T 5.81%). In contrast, in HUVECs with over-expression of COX-2, VEGF-induced cell proliferation was increased 42.56 T 7.69%. Moreover, the formation of vascular structure assayed by Matrigel demonstrated that VEGF-induced vascular structure formation was accelerated in COX-2 over-expressing cells but attenuated in COX-2 siRNA-transfected cells. Conclusion: COX-2 plays an important role in VEGF-induced angiogenesis via p38 and JNK kinase activation pathways. These findings suggest that the cardioprotective role of COX-2 may be, at least in part, through its angiogenic activity.

Arteriosclerosis, Thrombosis, and Vascular Biology, 2010