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1.
JCI Insight ; 8(5)2023 03 08.
Artículo en Inglés | MEDLINE | ID: mdl-36749637

RESUMEN

Healthy expansion of adipose tissue is critical for the maintenance of metabolic health, providing an optimized reservoir for energy storage in the form of triacylglycerol-rich lipoproteins. Dysfunctional adipocytes that are unable to efficiently store lipid can result in lipodystrophy and contribute to nonalcoholic fatty liver disease (NAFLD) and metabolic syndrome. Leucine-rich repeat containing protein 8a/SWELL1 functionally encodes the volume-regulated anion channel complex in adipocytes, is induced in early obesity, and is required for normal adipocyte expansion during high-fat feeding. Adipose-specific SWELL1 ablation (Adipo KO) leads to insulin resistance and hyperglycemia during caloric excess, both of which are associated with NAFLD. Here, we show that Adipo-KO mice exhibited impaired adipose depot expansion and excess lipolysis when raised on a variety of high-fat diets, resulting in increased diacylglycerides and hepatic steatosis, thereby driving liver injury. Liver lipidomic analysis revealed increases in oleic acid-containing hepatic triacylglycerides and injurious hepatic diacylglyceride species, with reductions in hepatocyte-protective phospholipids and antiinflammatory free fatty acids. Aged Adipo-KO mice developed hepatic steatosis on a regular chow diet, and Adipo-KO male mice developed spontaneous, aggressive hepatocellular carcinomas (HCCs). These data highlight the importance of adipocyte SWELL1 for healthy adipocyte expansion to protect against NAFLD and HCC in the setting of overnutrition and with aging.


Asunto(s)
Carcinoma Hepatocelular , Neoplasias Hepáticas , Enfermedad del Hígado Graso no Alcohólico , Animales , Masculino , Ratones , Dieta Alta en Grasa , Glucosa/metabolismo , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Obesidad/metabolismo
2.
Nat Commun ; 13(1): 784, 2022 02 10.
Artículo en Inglés | MEDLINE | ID: mdl-35145074

RESUMEN

Type 2 diabetes is associated with insulin resistance, impaired pancreatic ß-cell insulin secretion, and nonalcoholic fatty liver disease. Tissue-specific SWELL1 ablation impairs insulin signaling in adipose, skeletal muscle, and endothelium, and impairs ß-cell insulin secretion and glycemic control. Here, we show that ICl,SWELL and SWELL1 protein are reduced in adipose and ß-cells in murine and human diabetes. Combining cryo-electron microscopy, molecular docking, medicinal chemistry, and functional studies, we define a structure activity relationship to rationally-design active derivatives of a SWELL1 channel inhibitor (DCPIB/SN-401), that bind the SWELL1 hexameric complex, restore SWELL1 protein, plasma membrane trafficking, signaling, glycemic control and islet insulin secretion via SWELL1-dependent mechanisms. In vivo, SN-401 restores glycemic control, reduces hepatic steatosis/injury, improves insulin-sensitivity and insulin secretion in murine diabetes. These findings demonstrate that SWELL1 channel modulators improve SWELL1-dependent systemic metabolism in Type 2 diabetes, representing a first-in-class therapeutic approach for diabetes and nonalcoholic fatty liver disease.


Asunto(s)
Diabetes Mellitus Tipo 2/metabolismo , Control Glucémico/métodos , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Tejido Adiposo/metabolismo , Animales , Microscopía por Crioelectrón , Diabetes Mellitus Experimental/metabolismo , Glucosa/metabolismo , Insulina/metabolismo , Resistencia a la Insulina , Secreción de Insulina , Células Secretoras de Insulina/metabolismo , Hígado/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Simulación del Acoplamiento Molecular , Transducción de Señal , Transcriptoma
3.
Elife ; 102021 02 25.
Artículo en Inglés | MEDLINE | ID: mdl-33629656

RESUMEN

The endothelium responds to numerous chemical and mechanical factors in regulating vascular tone, blood pressure, and blood flow. The endothelial volume-regulated anion channel (VRAC) has been proposed to be mechanosensitive and thereby sense fluid flow and hydrostatic pressure to regulate vascular function. Here, we show that the leucine-rich repeat-containing protein 8a, LRRC8A (SWELL1), is required for VRAC in human umbilical vein endothelial cells (HUVECs). Endothelial LRRC8A regulates AKT-endothelial nitric oxide synthase (eNOS) signaling under basal, stretch, and shear-flow stimulation, forms a GRB2-Cav1-eNOS signaling complex, and is required for endothelial cell alignment to laminar shear flow. Endothelium-restricted Lrrc8a KO mice develop hypertension in response to chronic angiotensin-II infusion and exhibit impaired retinal blood flow with both diffuse and focal blood vessel narrowing in the setting of type 2 diabetes (T2D). These data demonstrate that LRRC8A regulates AKT-eNOS in endothelium and is required for maintaining vascular function, particularly in the setting of T2D.


Asunto(s)
Endotelio/fisiología , Proteínas de la Membrana/genética , Óxido Nítrico Sintasa de Tipo III/genética , Proteínas Proto-Oncogénicas c-akt/genética , Animales , Femenino , Masculino , Proteínas de la Membrana/metabolismo , Ratones , Óxido Nítrico Sintasa de Tipo III/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Transducción de Señal
4.
Elife ; 92020 09 15.
Artículo en Inglés | MEDLINE | ID: mdl-32930093

RESUMEN

Maintenance of skeletal muscle is beneficial in obesity and Type 2 diabetes. Mechanical stimulation can regulate skeletal muscle differentiation, growth and metabolism; however, the molecular mechanosensor remains unknown. Here, we show that SWELL1 (Lrrc8a) functionally encodes a swell-activated anion channel that regulates PI3K-AKT, ERK1/2, mTOR signaling, muscle differentiation, myoblast fusion, cellular oxygen consumption, and glycolysis in skeletal muscle cells. LRRC8A over-expression in Lrrc8a KO myotubes boosts PI3K-AKT-mTOR signaling to supra-normal levels and fully rescues myotube formation. Skeletal muscle-targeted Lrrc8a KO mice have smaller myofibers, generate less force ex vivo, and exhibit reduced exercise endurance, associated with increased adiposity under basal conditions, and glucose intolerance and insulin resistance when raised on a high-fat diet, compared to wild-type (WT) mice. These results reveal that the LRRC8 complex regulates insulin-PI3K-AKT-mTOR signaling in skeletal muscle to influence skeletal muscle differentiation in vitro and skeletal myofiber size, muscle function, adiposity and systemic metabolism in vivo.


Skeletal muscles ­ the force-generating tissue attached to bones ­ must maintain their mass and health for the body to work properly. It is therefore necessary to understand how an organism can regulate the way skeletal muscles form, grow and heal. A multitude of factors can control how muscles form, including mechanical signals. The molecules that can sense these mechanical stimuli, however, remain unknown. Mechanoresponsive ion channels provide possible candidates for these molecular sensors. These proteins are studded through the cell membranes, where they can respond to mechanical changes by opening and allowing the flow of ions in and out of a cell, or by changing interactions with other proteins. The SWELL1 protein is a component of an ion channel known as VRAC, which potentially responds to mechanical stimuli. This channel is associated with many biological processes such as cells multiplying, migrating, growing and dying, but it is still unclear how. Here, Kumar et al. first tested whether SWELL1 controls how skeletal muscle precursors mature into their differentiated and functional form. These experiments showed that SWELL1 regulates this differentiation process under the influence of the hormone insulin, as well as mechanical signals such as cell stretching. In addition, this work revealed that SWELL1 relies on an adaptor molecule called GRB2 to relay these signals in the cell. Next, Kumar et al. genetically engineered mice lacking SWELL1 only in skeletal muscle. These animals had smaller muscle cells, as well as muscles that were weaker and less enduring. When raised on a high-calorie diet, the mutant mice also got more obese and developed resistance to insulin, which is an important step driving obesity-induced diabetes. Together, these findings show that SWELL1 helps to regulate the formation and function of muscle cells, and highlights how an ion channel participates in these processes. Healthy muscles are key for overall wellbeing, as they also protect against obesity and obesity-related conditions such as type 2 diabetes or nonalcoholic fatty liver disease. This suggests that targeting SWELL1 could prove advantageous in a clinical setting.


Asunto(s)
Adiposidad/genética , Glucosa/metabolismo , Proteínas de la Membrana/genética , Ratones/fisiología , Músculo Esquelético/fisiología , Transducción de Señal/genética , Animales , Tamaño de la Célula , Femenino , Masculino , Proteínas de la Membrana/metabolismo , Ratones/genética , Células Musculares
5.
Biochemistry ; 48(36): 8559-67, 2009 Sep 15.
Artículo en Inglés | MEDLINE | ID: mdl-19681593

RESUMEN

The coiled-coil domain of cartilage oligomeric matrix protein (COMPcc) assembles into a homopentamer that naturally recognizes the small molecule 1,25-dihydroxyvitamin D(3) (vit D). To identify the residues critical for the structure, stability, oligomerization, and binding to vit D as well as two other small molecules, all-trans-retinol (ATR) and curcumin (CCM), here we perform an alanine scanning mutagenesis study. Ten residues lining the hydrophobic pocket of COMPcc were mutated into alanine; of the mutated residues, the N-terminal aliphatic residues L37, L44, V47, and L51 are responsible for maintaining the structure and function. Furthermore, two polar residues, T40 and Q54, within the N-terminal region when converted into alanine improve the alpha-helical structure, stability, and self-assembly behavior. Helical stability, oligomerization, and binding appear to be linked in a manner in which mutations that abolish helical structure and assembly bind poorly to vit D, ATR, and CCM. These results provide not only insight into COMPcc and its functional role but also useful guidelines for the design of stable, pentameric coiled-coils capable of selectively storing and delivering various small molecules.


Asunto(s)
Proteínas de la Matriz Extracelular/química , Proteínas de la Matriz Extracelular/metabolismo , Glicoproteínas/química , Glicoproteínas/metabolismo , Leucina , Fragmentos de Péptidos/química , Fragmentos de Péptidos/metabolismo , Estabilidad Proteica , Valina , Alanina/genética , Secuencia de Aminoácidos , Animales , Proteína de la Matriz Oligomérica del Cartílago , Curcumina/metabolismo , Proteínas de la Matriz Extracelular/genética , Glicoproteínas/genética , Humanos , Leucina/genética , Proteínas Matrilinas , Datos de Secuencia Molecular , Fragmentos de Péptidos/genética , Unión Proteica/genética , Estructura Terciaria de Proteína/genética , Ratas , Valina/genética , Vitamina A/genética , Vitamina A/metabolismo , Vitamina D/genética , Vitamina D/metabolismo
6.
Adipocyte ; 8(1): 223-228, 2019 12.
Artículo en Inglés | MEDLINE | ID: mdl-31112068

RESUMEN

Obesity is becoming a global epidemic, predisposing to Type 2 diabetes, cardiovascular disease, fatty liver disease, pulmonary disease, osteoarthritis and cancer. Therefore, understanding the biology of adipocyte expansion in response to overnutrition is critical to devising strategies to treat obesity, and the associated burden of morbidity and mortality. Through exploratory patch-clamp experiments in freshly isolated primary murine and human adipocytes, we recently determined that SWELL1/LRRC8a, a leucine-rich repeat containing transmembrane protein, functionally encoded an ion channel signalling complex (the volume-regulated anion channel, or VRAC) on the adipocyte plasma membrane. The SWELL1-/LRRC8 channel complex activates in response to increases in adipocyte volume and in the context of obesity. SWELL1 is also required for insulin-PI3K-AKT2 signalling to regulate adipocyte growth and systemic glycaemia. This commentary delves further into our working models for the molecular mechanisms of adipocyte SWELL1-mediated VRAC activation, proposed signal transduction mechanisms, and putative impact on adipocyte hypertrophy during caloric excess.


Asunto(s)
Adipocitos/metabolismo , Gotas Lipídicas/metabolismo , Proteínas de la Membrana/metabolismo , Transducción de Señal , Animales , Glucosa/metabolismo , Homeostasis , Humanos , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo
7.
Nat Commun ; 9(1): 367, 2018 01 25.
Artículo en Inglés | MEDLINE | ID: mdl-29371604

RESUMEN

Insulin secretion is initiated by activation of voltage-gated Ca2+ channels (VGCC) to trigger Ca2+-mediated insulin vesicle fusion with the ß-cell plasma membrane. The firing of VGCC requires ß-cell membrane depolarization, which is regulated by a balance of depolarizing and hyperpolarizing ionic currents. Here, we show that SWELL1 mediates a swell-activated, depolarizing chloride current (ICl,SWELL) in both murine and human ß-cells. Hypotonic and glucose-stimulated ß-cell swelling activates SWELL1-mediated ICl,SWELL and this contributes to membrane depolarization and activation of VGCC-dependent intracellular calcium signaling. SWELL1 depletion in MIN6 cells and islets significantly impairs glucose-stimulated insulin secretion. Tamoxifen-inducible ß-cell-targeted Swell1 KO mice have normal fasting serum glucose and insulin levels but impaired glucose-stimulated insulin secretion and glucose tolerance; and this is further exacerbated in mild obesity. Our results reveal that ß-cell SWELL1 modulates insulin secretion and systemic glycaemia by linking glucose-mediated ß-cell swelling to membrane depolarization and activation of VGCC-triggered calcium signaling.


Asunto(s)
Glucemia/metabolismo , Glucosa/metabolismo , Células Secretoras de Insulina/metabolismo , Proteínas de la Membrana/metabolismo , Animales , Sistemas CRISPR-Cas , Calcio/metabolismo , Canales de Calcio/metabolismo , Línea Celular Tumoral , Femenino , Glucosa/farmacología , Humanos , Insulina/metabolismo , Secreción de Insulina , Células Secretoras de Insulina/efectos de los fármacos , Islotes Pancreáticos/efectos de los fármacos , Islotes Pancreáticos/metabolismo , Masculino , Proteínas de la Membrana/genética , Ratones Noqueados , Ratones Transgénicos
8.
Channels (Austin) ; 11(6): 673-677, 2017 Nov 02.
Artículo en Inglés | MEDLINE | ID: mdl-28873008

RESUMEN

Obesity is associated with a loss of insulin-sensitivity and systemic dysglycemia, resulting in Type 2 diabetes, however the molecular mechanisms underlying this association are unclear. Through adipocyte patch-clamp studies, we recently showed that SWELL1 is required for the Volume-Regulated Anion Current (VRAC) in adipocytes and that SWELL1-mediated VRAC is activated by both mechanical and pathophysiological adipocyte expansion. We also demonstrated that adipocyte SWELL1 is required for maintaining insulin signaling and glucose homeostasis, particularly in the setting of obesity. Here we show that SWELL1 protein expression is induced in subcutaneous fat, visceral fat and liver in the setting of obesity. Long- term AAV/rec2-shRNA mediated SWELL1 knock-down in both fat and liver are associated with increased weight gain, increased adiposity and exacerbated insulin resistance in mice raised on a high-fat diet. These data further support the notion that SWELL1 induction occurs in insulin- sensitive tissues (liver and adipose) in the setting of over-nutrition and contributes to improved systemic glycemia by supporting enhanced insulin-sensitivity.


Asunto(s)
Adipocitos/metabolismo , Insulinas/metabolismo , Hígado/metabolismo , Proteínas de la Membrana/biosíntesis , Obesidad/metabolismo , Animales , Aniones/metabolismo , Dependovirus/metabolismo , Dieta Alta en Grasa/efectos adversos , Glucosa/metabolismo , Homeostasis , Resistencia a la Insulina , Ratones Endogámicos C57BL , ARN Interferente Pequeño/metabolismo , Transducción de Señal
9.
Nat Cell Biol ; 19(5): 504-517, 2017 05.
Artículo en Inglés | MEDLINE | ID: mdl-28436964

RESUMEN

Adipocytes undergo considerable volumetric expansion in the setting of obesity. It has been proposed that such marked increases in adipocyte size may be sensed via adipocyte-autonomous mechanisms to mediate size-dependent intracellular signalling. Here, we show that SWELL1 (LRRC8a), a member of the Leucine-Rich Repeat Containing protein family, is an essential component of a volume-sensitive ion channel (VRAC) in adipocytes. We find that SWELL1-mediated VRAC is augmented in hypertrophic murine and human adipocytes in the setting of obesity. SWELL1 regulates adipocyte insulin-PI3K-AKT2-GLUT4 signalling, glucose uptake and lipid content via SWELL1 C-terminal leucine-rich repeat domain interactions with GRB2/Cav1. Silencing GRB2 in SWELL1 KO adipocytes rescues insulin-pAKT2 signalling. In vivo, shRNA-mediated SWELL1 knockdown and adipose-targeted SWELL1 knockout reduce adiposity and adipocyte size in obese mice while impairing systemic glycaemia and insulin sensitivity. These studies identify SWELL1 as a cell-autonomous sensor of adipocyte size that regulates adipocyte growth, insulin sensitivity and glucose tolerance.


Asunto(s)
Adipocitos/metabolismo , Tamaño de la Célula , Metabolismo Energético , Glucosa/metabolismo , Insulina/metabolismo , Proteínas de la Membrana/metabolismo , Obesidad/metabolismo , Transducción de Señal , Adipocitos/patología , Adiposidad , Animales , Células Cultivadas , Canales de Cloruro/metabolismo , Modelos Animales de Enfermedad , Proteína Forkhead Box O1/genética , Proteína Forkhead Box O1/metabolismo , Proteína Adaptadora GRB2/genética , Proteína Adaptadora GRB2/metabolismo , Transportador de Glucosa de Tipo 4/genética , Transportador de Glucosa de Tipo 4/metabolismo , Glucógeno Sintasa Quinasa 3 beta/genética , Glucógeno Sintasa Quinasa 3 beta/metabolismo , Homeostasis , Humanos , Resistencia a la Insulina , Activación del Canal Iónico , Masculino , Potenciales de la Membrana , Proteínas de la Membrana/genética , Ratones Endogámicos C57BL , Obesidad/genética , Obesidad/patología , Fosfatidilinositol 3-Quinasa/metabolismo , Fosforilación , Proteínas Proto-Oncogénicas c-akt/metabolismo , Interferencia de ARN , Factores de Tiempo , Transfección
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