Mitochondrial electron transport chain, ceramide, and coenzyme Q are linked in a pathway that drives insulin resistance in skeletal muscle.

Insulin resistance (IR) is a complex metabolic disorder that underlies several human diseases, including type 2 diabetes and cardiovascular disease. Despite extensive research, the precise mechanisms underlying IR development remain poorly understood. Previously we showed that deficiency of coenzyme Q (CoQ) is necessary and sufficient for IR in adipocytes and skeletal muscle (Fazakerley et al., 2018). Here, we provide new insights into the mechanistic connections between cellular alterations associated with IR, including increased ceramides, CoQ deficiency, mitochondrial dysfunction, and oxidative stress. We demonstrate that elevated levels of ceramide in the mitochondria of skeletal muscle cells result in CoQ depletion and loss of mitochondrial respiratory chain components, leading to mitochondrial dysfunction and IR. Further, decreasing mitochondrial ceramide levels in vitro and in animal models (mice, C57BL/6J) (under chow and high-fat diet) increased CoQ levels and was protective against IR. CoQ supplementation also rescued ceramide-associated IR. Examination of the mitochondrial proteome from human muscle biopsies revealed a strong correlation between the respirasome system and mitochondrial ceramide as key determinants of insulin sensitivity. Our findings highlight the mitochondrial ceramide-CoQ-respiratory chain nexus as a potential foundation of an IR pathway that may also play a critical role in other conditions associated with ceramide accumulation and mitochondrial dysfunction, such as heart failure, cancer, and aging. These insights may have important clinical implications for the development of novel therapeutic strategies for the treatment of IR and related metabolic disorders.

Deoxysphingolipids: Atypical Skeletal Muscle Lipids Related to Insulin Resistance in Humans That Decrease Insulin Sensitivity In Vitro.

Sphingolipids are thought to promote skeletal muscle insulin resistance. Deoxysphingolipids (dSLs) are atypical sphingolipids that are increased in the plasma of individuals with type 2 diabetes and cause ß-cell dysfunction in vitro. However, their role in human skeletal muscle is unknown. We found that dSL species are significantly elevated in muscle of individuals with obesity and type 2 diabetes compared with athletes and lean individuals and are inversely related to insulin sensitivity. Furthermore, we observed a significant reduction in muscle dSL content in individuals with obesity who completed a combined weight loss and exercise intervention. Increased dSL content in primary human myotubes caused a decrease in insulin sensitivity associated with increased inflammation, decreased AMPK phosphorylation, and altered insulin signaling. Our findings reveal a central role for dSL in human muscle insulin resistance and suggest dSLs as therapeutic targets for the treatment and prevention of type 2 diabetes. ARTICLE HIGHLIGHTS: Deoxysphingolipids (dSLs) are atypical sphingolipids elevated in the plasma of individuals with type 2 diabetes, and their role in muscle insulin resistance has not been investigated. We evaluated dSL in vivo in skeletal muscle from cross-sectional and longitudinal insulin-sensitizing intervention studies and in vitro in myotubes manipulated to synthesize higher dSLs. dSLs were increased in the muscle of people with insulin resistance, inversely correlated to insulin sensitivity, and significantly decreased after an insulin-sensitizing intervention; increased intracellular dSL concentrations cause myotubes to become more insulin resistant. Reduction of muscle dSL levels is a potential novel therapeutic target to prevent/treat skeletal muscle insulin resistance.

Deoxysphingolipids - atypical skeletal muscle lipids related to insulin resistance in humans that decrease insulin sensitivity in vitro.

Sphingolipids are thought to promote skeletal muscle insulin resistance. 1-Deoxysphingolipids (dSL) are atypical sphingolipids that are increased in plasma of individuals with type 2 diabetes and cause ß-cell dysfunction in vitro. However, their role in human skeletal muscle in unknown. We found that dSL species are significantly elevated in muscle of individuals with obesity and type 2 diabetes compared to athletes and lean individuals and are inversely related to insulin sensitivity. Furthermore, we observed a significant reduction in muscle dSL content in individuals with obesity who completed a combined weight loss and exercise intervention. Increased dSL content in primary human myotubes caused a decrease in insulin sensitivity associated with increased inflammation, decreased AMP-activated kinase (AMPK) phosphorylation, and altered insulin signaling. Our findings reveal a central role for dSL in human muscle insulin resistance and suggest dSL as therapeutic targets for the treatment and prevention of type 2 diabetes.

Mitochondrial electron transport chain, ceramide and Coenzyme Q are linked in a pathway that drives insulin resistance in skeletal muscle.

Insulin resistance (IR) is a complex metabolic disorder that underlies several human diseases, including type 2 diabetes and cardiovascular disease. Despite extensive research, the precise mechanisms underlying IR development remain poorly understood. Here, we provide new insights into the mechanistic connections between cellular alterations associated with IR, including increased ceramides, deficiency of coenzyme Q (CoQ), mitochondrial dysfunction, and oxidative stress. We demonstrate that elevated levels of ceramide in the mitochondria of skeletal muscle cells results in CoQ depletion and loss of mitochondrial respiratory chain components, leading to mitochondrial dysfunction and IR. Further, decreasing mitochondrial ceramide levels in vitro and in animal models (under chow and high fat diet) increased CoQ levels and was protective against IR. CoQ supplementation also rescued ceramide-associated IR. Examination of the mitochondrial proteome from human muscle biopsies revealed a strong correlation between the respirasome system and mitochondrial ceramide as key determinants of insulin sensitivity. Our findings highlight the mitochondrial Ceramide-CoQ-respiratory chain nexus as a potential foundation of an IR pathway that may also play a critical role in other conditions associated with ceramide accumulation and mitochondrial dysfunction, such as heart failure, cancer, and aging. These insights may have important clinical implications for the development of novel therapeutic strategies for the treatment of IR and related metabolic disorders.

Quantifying the inflammatory secretome of human intermuscular adipose tissue.

Adipose tissue secretes an abundance of lipid and protein mediators, and this secretome is depot-specific, with local and systemic effects on metabolic regulation. Intermuscular adipose tissue (IMAT) accumulates within the skeletal muscle compartment in obesity, and is associated with insulin resistance and metabolic disease. While the human IMAT secretome decreases insulin sensitivity in vitro, its composition is entirely unknown. The current study was conducted to investigate the composition of the human IMAT secretome, compared to that of the subcutaneous (SAT) and visceral adipose tissue (VAT) depots. IMAT, SAT, and VAT explants from individuals with obesity were used to generate conditioned media. Proteomics analysis of conditioned media was performed using multiplex proximity extension assays, and eicosanoid analysis using liquid chromatography-tandem mass spectrometry. Compared to SAT and/or VAT, IMAT secreted significantly more cytokines (IL2, IL5, IL10, IL13, IL27, FGF23, IFNγ and CSF1) and chemokines (MCP1, IL8, CCL11, CCL20, CCL25 and CCL27). Adipokines hepatocyte growth factor and resistin were secreted significantly more by IMAT than SAT or VAT. IMAT secreted significantly more eicosanoids (PGE2, TXB2 , 5-HETE, and 12-HETE) compared to SAT and/or VAT. In the context of obesity, IMAT is a distinct adipose tissue with a highly immunogenic and inflammatory secretome, and given its proximity to skeletal muscle, may be critical to glucose regulation and insulin resistance.

Serum dihydroceramides correlate with insulin sensitivity in humans and decrease insulin sensitivity in vitro.

Serum ceramides, especially C16:0 and C18:0 species, are linked to CVD risk and insulin resistance, but details of this association are not well understood. We performed this study to quantify a broad range of serum sphingolipids in individuals spanning the physiologic range of insulin sensitivity and to determine if dihydroceramides cause insulin resistance in vitro. As expected, we found that serum triglycerides were significantly greater in individuals with obesity and T2D compared with athletes and lean individuals. Serum ceramides were not significantly different within groups but, using all ceramide data relative to insulin sensitivity as a continuous variable, we observed significant inverse relationships between C18:0, C20:0, and C22:0 species and insulin sensitivity. Interestingly, we found that total serum dihydroceramides and individual species were significantly greater in individuals with obesity and T2D compared with athletes and lean individuals, with C18:0 species showing the strongest inverse relationship to insulin sensitivity. Finally, we administered a physiological mix of dihydroceramides to primary myotubes and found decreased insulin sensitivity in vitro without changing the overall intracellular sphingolipid content, suggesting a direct effect on insulin resistance. These data extend what is known regarding serum sphingolipids and insulin resistance and show the importance of serum dihydroceramides to predict and promote insulin resistance in humans.

Exploring Visceral and Subcutaneous Adipose Tissue Secretomes in Human Obesity: Implications for Metabolic Disease.

Adipose tissue secretions are depot-specific and vary based on anatomical location. Considerable attention has been focused on visceral (VAT) and subcutaneous (SAT) adipose tissue with regard to metabolic disease, yet our knowledge of the secretome from these depots is incomplete. We conducted a comprehensive analysis of VAT and SAT secretomes in the context of metabolic function. Conditioned media generated using SAT and VAT explants from individuals with obesity were analyzed using proteomics, mass spectrometry, and multiplex assays. Conditioned media were administered in vitro to rat hepatocytes and myotubes to assess the functional impact of adipose tissue signaling on insulin responsiveness. VAT secreted more cytokines (IL-12p70, IL-13, TNF-α, IL-6, and IL-8), adipokines (matrix metalloproteinase-1, PAI-1), and prostanoids (TBX2, PGE2) compared with SAT. Secretome proteomics revealed differences in immune/inflammatory response and extracellular matrix components. In vitro, VAT-conditioned media decreased hepatocyte and myotube insulin sensitivity, hepatocyte glucose handling, and increased basal activation of inflammatory signaling in myotubes compared with SAT. Depot-specific differences in adipose tissue secretome composition alter paracrine and endocrine signaling. The unique secretome of VAT has distinct and negative impact on hepatocyte and muscle insulin action.

Utilization of Mid-Thigh Magnetic Resonance Imaging to Predict Lean Body Mass and Knee Extensor Strength in Obese Adults.

Purpose: To train and test a machine learning model to automatically measure mid-thigh muscle cross-sectional area (CSA) to provide rapid estimation of appendicular lean mass (ALM) and predict knee extensor torque of obese adults. Methods: Obese adults [body mass index (BMI) = 30-40 kg/m2, age = 30-50 years] were enrolled for this study. Participants received full-body dual-energy X-ray absorptiometry (DXA), mid-thigh MRI, and completed knee extensor and flexor torque assessments via isokinetic dynamometer. Manual segmentation of mid-thigh CSA was completed for all MRI scans. A convolutional neural network (CNN) was created based on the manual segmentation to develop automated quantification of mid-thigh CSA. Relationships were established between the automated CNN values to the manual CSA segmentation, ALM via DXA, knee extensor, and flexor torque. Results: A total of 47 obese patients were enrolled in this study. Agreement between the CNN-automated measures and manual segmentation of mid-thigh CSA was high (>0.90). Automated measures of mid-thigh CSA were strongly related to the leg lean mass (r = 0.86, p < 0.001) and ALM (r = 0.87, p < 0.001). Additionally, mid-thigh CSA was strongly related to knee extensor strength (r = 0.76, p < 0.001) and moderately related to knee flexor strength (r = 0.48, p = 0.002). Conclusion: CNN-measured mid-thigh CSA was accurate compared to the manual segmented values from the mid-thigh. These values were strongly predictive of clinical measures of ALM and knee extensor torque. Mid-thigh MRI may be utilized to accurately estimate clinical measures of lean mass and function in obese adults.