Biochimica et Biophysica Acta (BBA) - Molecular and Cell Biology of Lipids
Glucolipotoxicity diminishes cardiomyocyte TFEB and inhibits lysosomal autophagy during obesity and diabetes
Introduction
A subset of obese and diabetic patients suffers from cardiac muscle specific contractile dysfunction termed as cardiomyopathy [1], [2], [3], [4], [5]. Etiology in the progression of cardiomyopathy in rodent models of obesity and diabetes includes diastolic dysfunction [6], [7], myocyte hypertrophy [8], [9], [10], interstitial fibrosis [8], [11], [12], early-onset metabolic maladaptation [9], [10], [13], [14], and progressive lipid accumulation [9], [10], all of which precede heart failure. The underlying cause for metabolic inflexibility in cardiomyopathy is the lack of insulin or insulin action resulting in hyperglycemia and fatty acid overutilization, both of which lead to a condition known as “glucolipotoxicity” [1], [15], [16], [17]. Numerous studies have demonstrated that glucolipotoxic effects in the cardiomyocytes, originate or terminate primarily in the mitochondria and the endoplasmic reticulum (ER) [18], [19], [20], [21], [22].
A glucolipotoxic milieu in the cardiomyocyte impairs protein quality control, inducing ER stress and activating protein degradation pathway [23], [24]. To counter the damaging effects of glucolipotoxicity, proteasomal degradation is acutely activated to clear the cellular proteotoxic load [25]. Indeed, ubiquitin mRNA levels, caspase-3 and ATP-dependent proteasomal degradation are augmented acutely following streptozotocin (STZ)-induced type-1 diabetes [25]. However, sustained glucolipotoxicity exacerbates ER stress, saturating and impairing proteasomal protein degradation, causing toxic accumulation of misfolded proteins [26], [27]. In agreement with this theory, failing hearts from chronically obese humans with type-2 diabetes display significant accumulation of non-degraded proteins [28], suggesting that impaired proteasomal degradation in late stages of obesity and diabetes promotes a maladaptive buildup of cytotoxic proteins causing and/or exacerbating cardiomyopathy. Recent experimental evidence suggests that if the proteasome is impaired then damaged proteins must be degraded by the lysosomal machinery via autophagy to maintain cellular homeostasis [29], [30], [31]. Interestingly, islet dysfunction in ob/ob mice is exacerbated following treatment with lysosomal function inhibitors [32], suggesting that disruption in lysosomal function accelerates cell death. However, the underlying mechanisms by which glucolipotoxicity affects inter- and intra-lysosomal signaling, metabolism and function remain to be examined. Furthermore, whether clinically observed proteotoxicity and cardiomyopathy in obese and diabetic hearts involves negative targeting of lysosomes by glucolipotoxic substrates remains to be investigated.
Autophagy degrades short- and long-lived proteins in the lysosome [33], [34], [35] either via macroautophagy [36], [37], [38] or via chaperone-mediated autophagy (CMA) [39], [40]. Macroautophagy in the ER-cytosol interface requires lipidation of microtubule-associated protein 1 light chain B subtype 3 (LC3B-I) to form LC3B-II [37], [41] resulting in autophagosome formation, maturation and fusion with the lysosome to degrade proteins. The macroautophagy process utilizes polyubiquitin cargo-receptor, p62/SQSTM1 to engage in partial-selection of bulk load of intracellular protein and organelle content [23], [24] and therefore, changes in LC3B and p62/SQSTM1 signify changes in macroautophagy. Lysosomal CMA is a process by which cytosolic proteins targeted for degradation are delivered to lysosomal membrane protein-type 2A (LAMP-2A), which internalizes the protein cargo for lysosomal degradation [39], [40]. Notably, humans and mice with loss of function of LAMP-2 exhibit impaired autophagosome clearance, lysosomal dysfunction, and cardiomyopathy, suggesting that lysosomal autophagy is critical for cardiac function [42], [43], [44]. Expression of numerous lysosomal proteins responsible for autophagic processes are under the direct control of transcription factor EB (TFEB), a transcriptional regulator of lysosome autophagy and biogenesis [45], [46]. TFEB-action not only generates autophagosomes, but also accelerates their delivery and clearance by lysosomes via increases in lysosomal biogenesis [46]. It is plausible that changes in lysosome function and biogenesis could significantly impact cellular function in the setting of obesity, insulin resistance and diabetes, however, limited studies have examined the impact of glucolipotoxicity on TFEB and its downstream functions.
In this study, we examined whether glucolipotoxicity negatively targets TFEB, a transcriptional regulator of lysosome function, to impair autophagy and thereby render cardiomyocytes susceptible to proteotoxicity and cell death. Utilizing a mouse model of diet-induced obesity, type-1 diabetes and ex-vivo model of glucolipotoxicity (rat cardiomyofibroblasts and neonatal rat cardiomyocyte), we demonstrated that (1) baseline macroautophagy is reciprocally regulated in obesity and type-1 diabetes, (2) an obese and diabetic environment in-vivo suppressed lysosome signaling proteins, inhibited autophagic flux and reduced lysosomal proteolysis, (3) lysosomal protein suppression following ex-vivo myocyte nutrient overload is FA specific since palmitate or glucose/palmitate but not oleate or high glucose alone, reduces lysosomal protein content, (4) in the obese and diabetic heart, TFEB is decreased and this effect is recapitulated not only ex-vivo myocytes exposed to glucolipotoxic milieu but also in human heart tissue from patients with Class-1 obesity. Collectively, our data highlights a novel mechanism by which glucolipotoxicity targets TFEB to inhibit lysosomal integrity and render cardiomyocytes susceptible to proteotoxicity, injury and failure.
Section snippets
Animal models
All protocols involving rodents were approved by the Dalhousie University, Institutional Animal Care and Use Committee.
Heart from mice with diet induced obesity exhibit cellular stress with concomitant upregulation of cell growth pathways
To examine the impact of obesity and diabetes on cardiac autophagy, we fed C57BL/6J mice HFHS diet (45% kcal fat) for 16 weeks. This type of diet and length of feeding is reported to cause mild to moderate glucolipotoxicity in multiple tissues including the heart [9]. Consistent with previous diet-induced obesity studies [9], our mice fed HFHS diet exhibited increases in body weight (Fig. 1A) and fed glucose levels (Fig. 1B). Metabolic inflexibility in HFHS mice was evident from systemic glucose
Discussion
Protein degradation is severely impaired in the failing hearts of obese humans [28] and this impairment worsens with diabetes leading to buildup of cytotoxic proteins and causing cardiomyopathy. Experimental evidence suggests that cytotoxic proteins that are unable to be degraded by proteasome must be degraded by lysosomal autophagy to maintain cellular homeostasis [29], [30], [31]. Therefore, it is plausible that changes in lysosome function and biogenesis could significantly impact cellular
Conflict of interest
The authors declare that there is no conflict of interest.
Author contribution
T.P., P.C.T. and J.J.B. designed the research; P.C.T. and J.J.B. performed the experiments; P.C.T. and T.P. analyzed and interpreted the data and wrote the paper; L.J.P. generated mRNA data; P.C.K. provided intellectual inputs and technical assistance; K.R.B., J.L. & A.H assisted with clinical sample collection and provided intellectual inputs to clinical study.
Guarantor statement
Dr. Thomas Pulinilkunnil is the guarantor of this work, had full access to all the data, and takes full responsibility for the integrity of data and the accuracy of data analysis.
Transparency document
Acknowledgements
This work was supported by the Natural Sciences and Engineering Research Council of Canada (RGPIN-2014-03687), the Canadian Diabetes Association (NOD_OG-3-15-5037-TP) and the New Brunswick Health Research Foundation grants to T.P.; a Dalhousie Medicine New Brunswick graduate studentship to P.T. The clinical arm of this work was funded by a Chesley Grant (OPOS Study) to A.H. & P.C.K. in Saint John, NB, and by CIHR “REACH” grant to J.L. in Halifax, NS. We thank Dr. Gerard Karsenty for providing
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2021, Ageing Research ReviewsCitation Excerpt :On the other hand, calcineurin (CaN) and protein phosphatase 2A (PP2A) can activate TFEB activity by triggering its dephosphorylation and nuclear translocation (Fig. 3) (Medina et al., 2015). Glucolipotoxicity has been revealed to suppress TFEB in cardiomyocytes and inhibits lysosomal autophagy during obesity and diabetes (Trivedi et al., 2016). ZKSCAN3 acts as a key transcriptional repressor of autophagy flux, particularly, in the nutrient abundant state (Chauhan et al., 2013).