Papers by Thomas Pulinilkunnil
Cell Death Discovery
Triple-negative breast cancers (TNBCs) are characterized by poor survival, prognosis, and gradual... more Triple-negative breast cancers (TNBCs) are characterized by poor survival, prognosis, and gradual resistance to cytotoxic chemotherapeutics, like doxorubicin (DOX). The clinical utility of DOX is limited by its cardiotoxic and chemoresistant effects that manifest over time. To induce chemoresistance, TNBC rewires oncogenic gene expression and cell signaling pathways. Recent studies have demonstrated that reprogramming of branched-chain amino acids (BCAAs) metabolism facilitates tumor growth and survival. Branched-chain ketoacid dehydrogenase kinase (BCKDK), a regulatory kinase of the rate-limiting enzyme of the BCAA catabolic pathway, is reported to activate RAS/RAF/MEK/ERK signaling to promote tumor cell proliferation. However, it remains unexplored if BCKDK action remodels TNBC proliferation and survival per se and influences susceptibility to DOX-induced genotoxic stress. TNBC cells treated with DOX exhibited reduced BCKDK expression and intracellular BCKAs. Genetic and pharmacol...
Drug Metabolism Letters
Purpose: Previous studies have shown catabolism of adenosine 5’-triphosphate (ATP) in systemic bl... more Purpose: Previous studies have shown catabolism of adenosine 5’-triphosphate (ATP) in systemic blood is a potential surrogate biomarker for cardiovascular toxicity. We compared the acute toxicity of high doses of doxorubicin (DOX) and isoproterenol (ISO) on hemodynamics and ATP catabolism in systemic circulation. Methods: Sprague Dawley (SD) rats (n = 8 – 11) were each given either a single dose of 30 mg/kg ISO, or twice-daily dose of 10 mg/kg of DOX or normal saline (control) for 4 doses by subcutaneous injection. Blood samples were collected up to 6 hours for measuring concentrations of ATP and its catabolites. Hemodynmics was recorded continuously. Difference was considered significant at p < 0.05 (ANOVA). Results and Discussion: Mortality was 1/8, 5/11 and 0/11 for the DOX, ISO and control groups, respectively. Systolic blood pressure was significantly lower in the DOX and ISO treated rats than in the control measured at the last recorded time (76 ± 9 for DOX vs 42 ± 8 for IS...
Branched-chain α-keto acids (BCKAs) are downstream catabolites of branched-chain amino acids (BCA... more Branched-chain α-keto acids (BCKAs) are downstream catabolites of branched-chain amino acids (BCAAs). Mitochondrial oxidation of BCKAs is catalyzed by branched-chain ketoacid dehydrogenase (BCKDH), an enzyme sensitive to inhibitory phosphorylation by BCKD kinase (BCKDK). Emerging studies show that defective BCAA catabolism and elevated BCKAs levels correlate with glucose intolerance and cardiac dysfunction. However, if/how BCKDH and BCKDK exert control on the availability and flux of intramyocellular BCKAs and if BCKA reprograms nutrient metabolism by influencing insulin action remains unexplored. We observed altered BCAA catabolizing enzyme expression in the murine heart and skeletal muscle during physiological fasting and diet-induced obesity and after ex vivo exposure of C2C12 cells to increasing concentration of saturated fatty acid, palmitate. BCKAs per se impaired insulin-induced AKT phosphorylation and AKT activity in skeletal myotubes and cardiomyocytes. In skeletal muscle c...
Nutrients
Deregulation of lipid metabolism and insulin function in muscle and adipose tissue are hallmarks ... more Deregulation of lipid metabolism and insulin function in muscle and adipose tissue are hallmarks of systemic insulin resistance, which can progress to type 2 diabetes. While previous studies suggested that milk proteins influence systemic glucose homeostasis and insulin function, it remains unclear whether bioactive peptides generated from whey alter lipid metabolism and its accumulation in muscle and adipose tissue. Therefore, we incubated murine 3T3-L1 preadipocytes and C2C12 myotubes with a whey peptide mixture produced through pepsin-pancreatin digestion, mimicking peptides generated in the gut from whey protein hydrolysis, and examined its effect on indicators of lipid metabolism and insulin sensitivity. Whey peptides, particularly those derived from bovine serum albumin (BSA), promoted 3T3-L1 adipocyte differentiation and triacylglycerol (TG) accumulation in accordance with peroxisome proliferator-activated receptor γ (PPARγ) upregulation. Whey/BSA peptides also increased lipo...
Frontiers in Cardiovascular Medicine
Journal of Molecular and Cellular Cardiology
Although cancer cells use heparanase for tumor metastasis, favourable effects of heparanase have ... more Although cancer cells use heparanase for tumor metastasis, favourable effects of heparanase have been reported in the management of Alzheimer's disease and diabetes. Indeed, we previously established a protective function for heparanase in the acutely diabetic heart, where it conferred cardiomyocyte resistance to oxidative stress and apoptosis by provoking changes in gene expression. In this study, we tested if overexpression of heparanase can protect the heart against chemically induced or ischemia/reperfusion (I/R) injury. Transcriptomic analysis of Hep-tg hearts reveal that 240 genes related to the stress response, immune response, cell death, and development were altered in a pro-survival direction encompassing genes promoting the unfolded protein response (UPR) and autophagy, as well as those protecting against oxidative stress. The observed UPR activation was adaptive and not apoptotic, was mediated by activation of ATF6α, and when combined with mTOR inhibition, induced autophagy. Subjecting wild type (WT) mice to increasing concentrations of the ER stress inducer thapsigargin evoked a transition from adaptive to apoptotic UPR, an effect that was attenuated in Hep-tg mouse hearts. Consistent with these observations, when exposed to I/R, the infarct size and markers of apoptosis were significantly lower in the Hep-tg heart compared to WT. Finally, UPR and autophagy inhibitors reduced the protective effects of heparanase overexpression during I/R. Our data suggest that the mechanisms that underlie the role of heparanase in promoting cell survival could be uniquely beneficial to the heart by providing protection against cellular stresses, and could be useful for exploitation as a therapeutic target for the treatment of heart disease.
Journal of lipid research, Jan 2, 2018
Autotaxin (ATX) is an adipokine that generates the bioactive lipid, lysophosphatidic acid (LPA). ... more Autotaxin (ATX) is an adipokine that generates the bioactive lipid, lysophosphatidic acid (LPA). ATX-LPA signaling has been implicated in diet-induced obesity and systemic insulin resistance. However, it remains unclear whether the ATX-LPA pathway influences insulin function and energy metabolism in target tissues, particularly skeletal muscle, the major site of insulin-stimulated glucose disposal. The objective of this study was to test whether the ATX-LPA pathway impacts tissue insulin signaling and mitochondrial metabolism in skeletal muscle during obesity. Male mice with heterozygous ATX deficiency (ATX+/-) were protected from obesity, systemic insulin resistance, and cardiomyocyte dysfunction following high-fat high-sucrose (HFHS) feeding. HFHS-fed ATX+/- mice also had improved insulin-stimulated AKT phosphorylation in white adipose tissue, liver, heart, and skeletal muscle. Preserved insulin-stimulated glucose transport in muscle from HFHS-fed ATX+/- mice was associated with i...
PloS one, 2017
Lysophosphatidic acid (LPA) receptor signaling has been implicated in cardiovascular and obesity-... more Lysophosphatidic acid (LPA) receptor signaling has been implicated in cardiovascular and obesity-related metabolic disease. However, the distribution and regulation of LPA receptors in the myocardium and adipose tissue remain unclear. This study aimed to characterize the mRNA expression of LPA receptors (LPA1-6) in the murine and human myocardium and adipose tissue, and its regulation in response to obesity. LPA receptor mRNA levels were determined by qPCR in i) heart ventricles, isolated cardiomyocytes, and perigonadal adipose tissue from chow or high fat-high sucrose (HFHS)-fed male C57BL/6 mice, ii) 3T3-L1 adipocytes and HL-1 cardiomyocytes under conditions mimicking gluco/lipotoxicity, and iii) human atrial and subcutaneous adipose tissue from non-obese, pre-obese, and obese cardiac surgery patients. LPA1-6 were expressed in myocardium and white adipose tissue from mice and humans, except for LPA3, which was undetectable in murine adipocytes and human adipose tissue. Obesity was...
Journal of Molecular and Cellular Cardiology
Doxorubicin (DOX)-induced cardiotoxicity has been a well-known phenomenon to clinicians and scien... more Doxorubicin (DOX)-induced cardiotoxicity has been a well-known phenomenon to clinicians and scientists for decades; however, molecular mechanisms underlying DOX cardiotoxicity are still being uncovered. Although the majority of prior research have implicated nuclear and mitochondrial events to be an important etiological aspects of DOX cardiomyopathy, recent discoveries in autophagy have highlighted the renewed interest in the role of lysosome in DOX cardiomyopathy. Indeed, dysregulation of lysosomal autophagy is observed in pre-clinical models of DOX cardiotoxicity. In this review, we provide a comprehensive overview on mechanisms describing regulation of the autophagy pathway by DOX and its influence on cardiotoxic outcomes. We have put specific emphasis on experimental models, dosing and treatment duration with DOX, and methods to monitor autophagy, all of which contribute to inconsistencies observed in the literature. We have clarified processes by which DOX dysregulates macroautophagy in the heart by primarily focusing on the contribution of LC3, p62, Beclin, mTOR and AMPK pathways. We have also highlighted the impact of DOX on mitochondrial reactive oxygen species (ROS) and its contribution to the process of mitophagy. We have presented mechanisms by which DOX compromises lysosomal acidification, integrity and chaperone-mediated autophagy through its effect on lysosome-associated and resident proteins such as LAMP, vATPase, Hsp90, Hsc70 and cathepsins. Furthermore, we have discussed novel pathways in DOX cardiotoxicity, the most prominent being DOX-induced loss of TFEB, a member of the MITF family of transcription factors, which governs lysosomal biogenesis and function. This review summarizes that in the myocardium, DOX dysregulates autophagy by impairing transcriptional factors regulating lysosomal function, thereby, precipitating proteotoxicity, mitochondrial dysfunction and cell death, thus rendering the heart susceptible to cardiomyopathic failure.
Molecular Cancer Research
The microphthalmia family (MITF, TFEB, TFE3, and TFEC) of transcription factors is emerging as gl... more The microphthalmia family (MITF, TFEB, TFE3, and TFEC) of transcription factors is emerging as global regulators of cancer cell survival and energy metabolism, both through the promotion of lysosomal genes as well as newly characterized targets, such as oxidative metabolism and the oxidative stress response. In addition, MiT/TFE factors can regulate lysosomal signaling, which includes the mTORC1 and Wnt/b-catenin pathways, which are both substantial contributors to oncogenic signaling. This review describes recent discoveries in MiT/TFE research and how they impact multiple cancer subtypes. Furthermore, the literature relating to TFE-fusion proteins in cancers and the potential mechanisms through which these genomic rearrangements promote tumorigenesis is reviewed. Likewise, the emerging function of the Folliculin (FLCN) tumor suppressor in negatively regulating the MiT/TFE family and how loss of this pathway promotes cancer is examined. Recent reports are also presented that relate to the role of MiT/TFE-driven lysosomal biogenesis in sustaining cancer cell metabolism and signaling in nutrient-limiting conditions. Finally, a discussion is provided on the future directions and unanswered questions in the field. In summary, the research surrounding the MiT/TFE family indicates that these transcription factors are promising therapeutic targets and biomarkers for cancers that thrive in stressful niches. Mol Cancer Res; 15(12); 1637-43. Ó2017 AACR.
Journal of Pharmacology and Experimental Therapeutics
Dieldrin is a legacy organochlorine pesticide that is persistent in the environment, despite bein... more Dieldrin is a legacy organochlorine pesticide that is persistent in the environment, despite being discontinued from use in North America since the 1970s. Some epidemiologic studies suggest that exposure to dieldrin is associated with increased risks of neurodegenerative disease and breast cancer by inducing inflammatory responses in tissues as well as oxidative stress. However, the direct effects of organochlorine pesticides on the heart have not been adequately addressed to date given that these chemicals are detectable in human serum and are environmentally persistent; thus, individuals may show latent adverse effects in the cardiovascular system due to long-term, low-dose exposure over time. Our objective was to determine whether low-level exposure to dieldrin at an environmentally relevant dose results in aberrant molecular signaling in the vertebrate heart. Using transcriptomic profiling and immunoblotting, we determined the global gene and targeted protein expression response to dieldrin treatment and show that dieldrin affects gene networks in the heart that are associated with processes related to cardiovascular disease, specifically cardiac arrest and ventricular fibrillation. We report that genes regulating inflammatory responses, a significant risk factor for cardiovascular disease, are upregulated by dieldrin whereas transcripts related to lysosomal function are significantly downregulated. To verify these findings, proteins in these pathways were examined with immunoblotting, and our results demonstrate that dieldrin constitutively activates Akt/mTOR signaling and downregulates lysosomal genes, participating in autophagy. Our data demonstrate that dieldrin induces genes associated with cardiovascular dysfunction and compromised lysosomal physiology, thereby identifying a novel mechanism for pesticideinduced cardiotoxicity.
Endocrinology, 2017
Autotaxin (ATX) is an adipokine that generates the bioactive lipid, lysophosphatidic acid. Despit... more Autotaxin (ATX) is an adipokine that generates the bioactive lipid, lysophosphatidic acid. Despite recent studies implicating adipose-derived ATX in metabolic disorders including obesity and insulin resistance, the nutritional and hormonal regulation of ATX in adipocytes remains unclear. The current study examined the regulation of ATX in adipocytes by glucose and insulin and the role of ATX in adipocyte metabolism. Induction of insulin resistance in adipocytes with high glucose and insulin concentrations increased ATX secretion, whereas coincubation with the insulin sensitizer, rosiglitazone, prevented this response. Moreover, glucose independently increased ATX messenger RNA (mRNA), protein, and activity in a time- and concentration-dependent manner. Glucose also acutely upregulated secreted ATX activity in subcutaneous adipose tissue explants. Insulin elicited a biphasic response. Acute insulin stimulation increased ATX activity in a PI3Kinase-dependent and mTORC1-independent man...
Frontiers in cardiovascular medicine, 2017
The characteristics of circulating inflammatory cells (leukocytes) in patients undergoing heart s... more The characteristics of circulating inflammatory cells (leukocytes) in patients undergoing heart surgery remains poorly understood. Recently, neutrophil-to-lymphocyte ratio (NLR) and specific monocyte subsets (based on CD14/CD16 expression) have been suggested as markers of inflammation and predictors of outcomes. The present study aims to characterize the influence cardiac surgery with cardiopulmonary bypass has on specific circulating leukocytes. All enrolled patients had blood samples taken pre- (0 days), early post- (5 days), and late post- (90 days) surgery. Complete blood counts were performed and whole leukocyte isolations were obtained from blood samples and analyzed with flow cytometry. Fluorophore-linked antibodies (CD45, CD11b, CD14, and CD16) were added to the blood cell isolations and later assessed by flow cytometry. Seventeen patients were enrolled and samples obtained at 0, 5, and 90 days. We demonstrated a significant increase in NLR (2.2-fold; p = 0.0028) and CD16 m...
Circulation, Nov 22, 2011
Circulation, Nov 22, 2011
Despite the association between myocardial steatosis and cardiac dysfunction it remains unclear w... more Despite the association between myocardial steatosis and cardiac dysfunction it remains unclear whether increased cardiac lipid accumulation in the absence of increased fatty acid uptake or genetic...
Circulation, Nov 22, 2011
Anthracyclines such as, doxorubicin (DOX) are an effective class of antineoplastic agents. Despit... more Anthracyclines such as, doxorubicin (DOX) are an effective class of antineoplastic agents. Despite its efficacy in the treatment of a variety of cancers including breast cancer, the clinical use of...
American Journal of Physiology - Endocrinology And Metabolism, 2015
Adipose triglyceride lipase (ATGL) is the rate-limiting enzyme mediating triacylglycerol hydrolys... more Adipose triglyceride lipase (ATGL) is the rate-limiting enzyme mediating triacylglycerol hydrolysis in virtually all cells, including adipocytes and skeletal myocytes, and hence, plays a critical role in mobilizing fatty acids. Global ATGL deficiency promotes skeletal myopathy and exercise intolerance in mice and humans, and yet the tissue-specific contributions to these phenotypes remain unknown. The goal of this study was to determine the relative contribution of ATGL-mediated triacylglycerol hydrolysis in adipocytes vs. skeletal myocytes to acute exercise performance. To achieve this goal, we generated murine models with adipocyte- and skeletal myocyte-specific targeted deletion of ATGL. We then subjected untrained mice to acute peak and submaximal exercise interventions and assessed exercise performance and energy substrate metabolism. Impaired ATGL-mediated lipolysis within adipocytes reduced peak and submaximal exercise performance, reduced peripheral energy substrate availabi...
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Papers by Thomas Pulinilkunnil