Chronic stress targets mitochondrial respiratory efficiency in the skeletal muscle of C57BL/6 mice.

Episodes of chronic stress can result in psychic disorders like post-traumatic stress disorder, but also promote the development of metabolic syndrome and type 2 diabetes. We hypothesize that muscle, as main regulator of whole-body energy expenditure, is a central target of acute and adaptive molecular effects of stress in this context. Here, we investigate the immediate effect of a stress period on energy metabolism in Musculus gastrocnemius in our established C57BL/6 chronic variable stress (Cvs) mouse model. Cvs decreased lean body mass despite increased energy intake, reduced circadian energy expenditure (EE), and substrate utilization. Cvs altered the proteome of metabolic components but not of the oxidative phosphorylation system (OXPHOS), or other mitochondrial structural components. Functionally, Cvs impaired the electron transport chain (ETC) capacity of complex I and complex II, and reduces respiratory capacity of the ETC from complex I to ATP synthase. Complex I-OXPHOS correlated to diurnal EE and complex II-maximal uncoupled respiration correlated to diurnal and reduced nocturnal EE. Bioenergetics assessment revealed higher optimal thermodynamic efficiencies (ƞ-opt) of mitochondria via complex II after Cvs. Interestingly, transcriptome and methylome were unaffected by Cvs, thus excluding major contributions to supposed metabolic adaptation processes. In summary, the preclinical Cvs model shows that metabolic pressure by Cvs is initially compensated by adaptation of mitochondria function associated with high thermodynamic efficiency and decreased EE to manage the energy balance. This counter-regulation of mitochondrial complex II may be the driving force to longitudinal metabolic changes of muscle physiological adaptation as the basis of stress memory.

Adaptation of Oxidative Phosphorylation Machinery Compensates for Hepatic Lipotoxicity in Early Stages of MAFLD.

Alterations in mitochondrial function are an important control variable in the progression of metabolic dysfunction-associated fatty liver disease (MAFLD), while also noted by increased de novo lipogenesis (DNL) and hepatic insulin resistance. We hypothesized that the organization and function of a mitochondrial electron transport chain (ETC) in this pathologic condition is a consequence of shifted substrate availability. We addressed this question using a transgenic mouse model with increased hepatic insulin resistance and DNL due to constitutively active human SREBP-1c. The abundance of ETC complex subunits and components of key metabolic pathways are regulated in the liver of these animals. Further omics approaches combined with functional assays in isolated liver mitochondria and primary hepatocytes revealed that the SREBP-1c-forced fatty liver induced a substrate limitation for oxidative phosphorylation, inducing enhanced complex II activity. The observed increased expression of mitochondrial genes may have indicated a counteraction. In conclusion, a shift of available substrates directed toward activated DNL results in increased electron flows, mainly through complex II, to compensate for the increased energy demand of the cell. The reorganization of key compounds in energy metabolism observed in the SREBP-1c animal model might explain the initial increase in mitochondrial function observed in the early stages of human MAFLD.

Physiological Disturbance in Fatty Liver Energy Metabolism Converges on IGFBP2 Abundance and Regulation in Mice and Men.

Fatty liver occurs from simple steatosis with accumulated hepatic lipids and hepatic insulin resistance to severe steatohepatitis, with aggravated lipid accumulation and systemic insulin resistance, but this progression is still poorly understood. Analyses of hepatic gene expression patterns from alb-SREBP-1c mice with moderate, or aP2-SREBP-1c mice with aggravated, hepatic lipid accumulation revealed IGFBP2 as key nodal molecule differing between moderate and aggravated fatty liver. Reduced IGFBP2 expression in aggravated fatty liver was paralleled with promoter hypermethylation, reduced hepatic IGFBP2 secretion and IGFBP2 circulating in plasma. Physiologically, the decrease of IGFBP2 was accompanied with reduced fatty acid oxidation and increased de novo lipogenesis potentially mediated by IGF1 in primary hepatocytes. Furthermore, methyltransferase and sirtuin activities were enhanced. In humans, IGFBP2 serum concentration was lower in obese men with non-alcoholic fatty liver disease (NAFLD) and steatohepatitis (NASH) compared to non-obese controls, and liver fat reduction by weight-loss intervention correlated with an increase of IGFBP2 serum levels. In conclusion, hepatic IGFBP2 abundance correlates to its circulating level and is related to hepatic energy metabolism and de novo lipogenesis. This designates IGFBP2 as non-invasive biomarker for fatty liver disease progression and might further provide an additional variable for risk prediction for pathogenesis of fatty liver in diabetes subtype clusters.

Adipokinome Signatures in Obese Mouse Models Reflect Adipose Tissue Health and Are Associated with Serum Lipid Composition.

Adipocyte and hepatic lipid metabolism govern whole-body metabolic homeostasis, whereas a disbalance of de novo lipogenesis (DNL) in fat and liver might lead to obesity, with severe co-morbidities. Nevertheless, some obese people are metabolically healthy, but the "protective" mechanisms are not yet known in detail. Especially, the adipocyte-derived molecular mediators that indicate adipose functionality are poorly understood. We studied transgenic mice (alb-SREBP-1c) with a "healthy" obese phenotype, and obob mice with hyperphagia-induced "sick" obesity to analyze the impact of the tissue-specific DNL on the secreted proteins, i.e., the adipokinome, of the primary adipose cells by label-free proteomics. Compared to the control mice, adipose DNL is reduced in both obese mouse models. In contrast, the hepatic DNL is reduced in obob but elevated in alb-SREBP-1c mice. To investigate the relationship between lipid metabolism and adipokinomes, we formulated the "liver-to-adipose-tissue DNL" ratio. Knowledge-based analyses of these results revealed adipocyte functionality with proteins, which was involved in tissue remodeling or metabolism in the alb-SREBP-1c mice and in the control mice, but mainly in fibrosis in the obob mice. The adipokinome in "healthy" obesity is similar to that in a normal condition, but it differs from that in "sick" obesity, whereas the serum lipid patterns reflect the "liver-to-adipose-tissue DNL" ratio and are associated with the adipokinome signature.

CDH13 abundance interferes with adipocyte differentiation and is a novel biomarker for adipose tissue health.

BACKGROUND: CDH13, an atypical member of the cadherin superfamily, has been identified in adipocyte secretomes of lean mouse models. CDH13 abundance differs in mouse models according to their susceptibility to develop metabolic disorders, but the role of CDH13 in adipose tissue is unknown. METHODS: Secreted CDH13 protein levels and mRNA levels in visceral adipose tissue were determined in lean and obese mouse models. In vitro studies were performed in 3T3-L1 adipocytes to determine the role of CDH13 in adipocyte differentiation. The pathophysiological impact of visceral adipose tissue CDH13 mRNA and circulating CDH13 levels were determined in humans (normal-weight men n = 37, obese men n = 109 including n = 51 type 2 diabetes patients) and in obese patients (n = 14) pre- and post-metabolic surgery. RESULTS: This study shows that in visceral adipose tissue CDH13 protein secretion and mRNA levels were decreased in obese mouse models. Mechanistically, CDH13 affects lipid metabolism during adipogenesis but not in mature adipocytes. CDH13 knockdown during adipogenesis reduced fatty acid uptake and lipid content in developing adipocytes. Furthermore, CDH13 depletion during adipogenesis lowered the induction of PPARγ and C/EBPα expression. These observations are of pathophysiological impact since visceral adipose tissue CDH13 mRNA and circulating CDH13 levels were decreased in obese men compared to normal-weight controls. Weight loss induced by bariatric surgery restored circulating CDH13 to levels found in normal-weight controls. CONCLUSIONS: CDH13 levels in adipose tissue and the circulation are affected by obesity in mouse models and humans and are restored by weight loss in humans. CDH13 interferes with the differentiation potential of adipocytes and therefore is a marker for plasticity of fat tissue that might reflect the health status of adipose tissue.

Soluble CD14 inhibits contractile function and insulin action in primary adult rat cardiomyocytes.

Epicardial adipose tissue (EAT) from patients with type 2 diabetes (T2D) is characterized by monocyte infiltrations and displays an elevated release of the monocyte marker soluble cluster of differentiation 14 (sCD14) versus EAT from patients without T2D. We propose that an increased abundance of sCD14 in EAT from patients with T2D may impair the function and insulin sensitivity of the adjacent cardiomyocytes. To examine this, primary adult rat cardiomyocytes were incubated with increasing concentrations of sCD14 in the presence and absence of the co-receptor lipopolysaccharide (LPS), and analyzed for effects on determinants of contractile function, activation of inflammation signalling and insulin action. Exposing cardiomyocytes to sCD14 increased the phosphorylation of the stress kinases p38 and extracellular-signal regulated kinase (ERK). In contrast, insulin-mediated phosphorylation of Akt on Thr308 and Ser473 was inhibited. Furthermore, sCD14 impaired sarcomere shortening and cytosolic Ca2+-fluxes. All responses were concentration-dependent and became significant at 1ng/ml sCD14. LPS, either alone or in complex with sCD14, did not affect contractile function or the activation of stress kinases and insulin signalling pathways. Similar data on protein phosphorylation were obtained when exposing human umbilical vein endothelial cells to sCD14. Finally, pharmacological inhibition of p38 reversed the detrimental effects of sCD14 on contractile function, but not on sCD14-induced insulin resistance. Collectively, these data show that sCD14 impairs the function and insulin sensitivity of cardiomyocytes, suggesting that an enhanced sCD14 release from EAT in patients with T2D may contribute to the pathogenesis of diabetes-related cardiometabolic complications.

Secretory products from epicardial adipose tissue from patients with type 2 diabetes impair mitochondrial ß-oxidation in cardiomyocytes via activation of the cardiac renin-angiotensin system and induction of miR-208a.

Secretory products from epicardial adipose tissue (EAT) from patients with type 2 diabetes (T2D) impair cardiomyocyte function. These changes associate with alterations in miRNA expression, including the induction of miR-208a. Recent studies suggest that activation of the cardiac-specific renin-angiotensin system (RAS) may affect cardiac energy metabolism via induction of miR-208a. This study investigated whether cardiomyocyte dysfunction induced by conditioned media (CM) from EAT-T2D involves activation of the RAS/miR-208a pathway. Therefore, primary adult rat cardiomyocytes were incubated with CM generated from EAT biopsies from patients with T2D and without T2D (ND). Exposing cardiomyocytes to CM-EAT-T2D reduced sarcomere shortening and increased miR-208a expression versus cells exposed to CM-EAT-ND or control medium. The angiotensin II receptor type 1 (AGTR1) antagonist losartan reversed these effects. Accordingly, incubation with angiotensin II (Ang II) reduced sarcomere shortening, and lowered palmitate-induced mitochondrial respiration and carnitine palmitoyltransferase 1c (CPT1c) expression in cardiomyocytes. Locked-nucleic-acid-mediated inhibition of miR-208a function reversed the detrimental effects induced by Ang II. Interestingly, Ang II levels in CM-EAT-T2D were increased by 2.6-fold after culture with cardiomyocytes. The paracrine activation of the cardiac-specific RAS by CM-EAT-T2D was corroborated by increases in the expression of AGTR1 and renin, as well as a reduction in angiotensin-converting enzyme 2 levels. Collectively, these data show that secretory products from EAT-T2D impair cardiomyocyte contractile function and mitochondrial ß-oxidation via activation of the cardiac-specific RAS system and induction of miR-208a, and suggest that alterations in the secretory profile of EAT may contribute to the development of diabetes-related heart disease.

Chronic stress alters hepatic metabolism and thermodynamic respiratory efficiency affecting epigenetics in C57BL/6 mice.

Chronic stress episodes increase metabolic disease risk even after recovery. We propose that persistent stress detrimentally impacts hepatic metabolic reprogramming, particularly mitochondrial function. In male C57BL/6 mice chronic variable stress (Cvs) reduced energy expenditure (EE) and body mass despite increased energy intake versus controls. This coincided with decreased glucose metabolism and increased lipid ß-oxidation, correlating with EE. After Cvs, mitochondrial function revealed increased thermodynamic efficiency (ƞ-opt) of complex CI, positively correlating with blood glucose and NEFA and inversely with EE. After Cvs recovery, the metabolic flexibility of hepatocytes was lost. Reduced CI-driving NAD+/NADH ratio, and diminished methylation-related one-carbon cycle components hinted at epigenetic regulation. Although initial DNA methylation differences were minimal after Cvs, they diverged during the recovery phase. Here, the altered enrichment of mitochondrial DNA methylation and linked transcriptional networks were observed. In conclusion, Cvs rapidly initiates the reprogramming of hepatic energy metabolism, supported by lasting epigenetic modifications.

Long-term adjustment of hepatic lipid metabolism after chronic stress and the role of FGF21.

Chronic stress leads to post-traumatic stress disorder (PTSD) and metabolic disorders including fatty liver. We hypothesized that stress-induced molecular mechanisms alter energy metabolism, thereby promoting hepatic lipid accumulation even after a stress-free recovery period. In this context, we investigated fibroblast growth factor-21 (FGF21) as protective for energy and glucose homeostasis. FGF21 knockout mice (B6.129S6(SJL)-Fgf21tm1.2Djm; FGF21KO) and control mice (C57BL6; WT) were subjected to chronic variable stress. Mice were examined directly after acute intervention (Cvs) and long-term after 3 months of recovery (3mCvs). In WT, Cvs reduced insulin sensitivity and hepatic lipid accumulation, whilst fatty acid uptake increased. FGF21KO mice responded to Cvs with improved glucose tolerance, insulin resistance but liver triglycerides and plasma lipids were unaltered. Hepatic gene expression was specifically altered by genotype and stress e.g. by PPARa and SREBP-1 regulated genes. The stress-induced alteration of hepatic metabolism persisted after stress recovery. In hepatocytes at 3mCvs, differential gene regulation and secreted proteins indicated a genotype specific progression of liver dysfunction. Overall, at 3mCvs FGF21 was involved in maintaining mitochondrial activity, attenuating de novo lipogenesis, increased fatty acid uptake and histone acetyltransferase activity. Glucocorticoid release and binding to the FGF21 promoter may contribute to prolonged FGF21 release and protection against hepatic lipid accumulation. In conclusion, we showed that stress favors fatty liver disease and FGF21 protected against hepatic lipid accumulation after previous chronic stress loading by i) restored physiological function, ii) modulated gene expression via DNA-modifying enzymes, and iii) maintained energy metabolism.

Rhein, a novel Histone Deacetylase (HDAC) inhibitor with antifibrotic potency in human myocardial fibrosis.

Although fibrosis depicts a reparative mechanism, maladaptation of the heart due to excessive production of extracellular matrix accelerates cardiac dysfunction. The anthraquinone Rhein was examined for its anti-fibrotic potency to mitigate cardiac fibroblast-to-myofibroblast transition (FMT). Primary human ventricular cardiac fibroblasts were subjected to hypoxia and characterized with proteomics, transcriptomics and cell functional techniques. Knowledge based analyses of the omics data revealed a modulation of fibrosis-associated pathways and cell cycle due to Rhein administration during hypoxia, whereas p53 and p21 were identified as upstream regulators involved in the manifestation of cardiac fibroblast phenotypes. Mechanistically, Rhein acts inhibitory on HDAC classes I/II as enzymatic inhibitor. Rhein-mediated cellular effects were linked to the histone deacetylase (HDAC)-dependent protein stabilization of p53 under normoxic but not hypoxic conditions. Functionally, Rhein inhibited collagen contraction, indicating anti-fibrotic property in cardiac remodeling. This was accompanied by increased abundance of SMAD7, but not SMAD2/3, and consistently SMAD-specific E3 ubiquitin ligase SMURF2. In conclusion, this study identifies Rhein as a novel potent direct HDAC inhibitor that may contribute to the treatment of cardiac fibrosis as anti-fibrotic agent. As readily available drug with approved safety, Rhein constitutes a promising potential therapeutic approach in the supplemental and protective intervention of cardiac fibrosis.

Fatty Liver Due to Increased de novo Lipogenesis: Alterations in the Hepatic Peroxisomal Proteome.

In non-alcoholic fatty liver disease (NAFLD) caused by ectopic lipid accumulation, lipotoxicity is a crucial molecular risk factor. Mechanisms to eliminate lipid overflow can prevent the liver from functional complications. This may involve increased secretion of lipids or metabolic adaptation to ß-oxidation in lipid-degrading organelles such as mitochondria and peroxisomes. In addition to dietary factors, increased plasma fatty acid levels may be due to increased triglyceride synthesis, lipolysis, as well as de novo lipid synthesis (DNL) in the liver. In the present study, we investigated the impact of fatty liver caused by elevated DNL, in a transgenic mouse model with liver-specific overexpression of human sterol regulatory element-binding protein-1c (alb-SREBP-1c), on hepatic gene expression, on plasma lipids and especially on the proteome of peroxisomes by omics analyses, and we interpreted the results with knowledge-based analyses. In summary, the increased hepatic DNL is accompanied by marginal gene expression changes but massive changes in peroxisomal proteome. Furthermore, plasma phosphatidylcholine (PC) as well as lysoPC species were altered. Based on these observations, it can be speculated that the plasticity of organelles and their functionality may be directly affected by lipid overflow.

The adipokine sFRP4 induces insulin resistance and lipogenesis in the liver.

Secreted frizzled-related protein (sFRP) 4 is an adipokine with increased expression in white adipose tissue from obese subjects with type 2 diabetes and non-alcoholic fatty liver disease (NAFLD). Yet, it is unknown whether sFRP4 action contributes to the development of these pathologies. Here, we determined whether sFRP4 expression in visceral fat associates with NAFLD and whether it directly interferes with insulin action and lipid and glucose metabolism in primary hepatocytes and myotubes. The association of sFRP4 with clinical measures was investigated in obese men with or without type 2 diabetes and with or without biopsy-proven NAFLD. To determine the impact of sFRP4 on metabolic parameters, primary human myotubes (hSkMC), or primary hepatocytes from metabolic healthy C57Bl6 and from systemic insulin-resistant mice, i.e. aP2-SREBP-1c, were used. Gene expression of sFRP4 in visceral fat from obese men associated with insulin sensitivity, triglycerides and NAFLD. In C57Bl6 hepatocytes, sFRP4 disturbed insulin action. Specifically, sFRP4 decreased the abundance of IRS1 and FoxO1 together with impaired insulin-mediated activation of Akt-signalling and glycogen synthesis and a reduced suppression of gluconeogenesis by insulin. Moreover, sFRP4 enhanced insulin-stimulated hepatic de novo lipogenesis (DNL). In hSkMC, sFRP4 induced glycolysis rather than inhibiting insulin signalling. Finally, in hepatocytes from aP2-SREBP-1c mice, sFRP4 potentiates existing insulin resistance. Collectively, we show that sFRP4 interferes with hepatocyte insulin action. Physiologically, sFRP4 promotes DNL in hepatocytes and glycolysis in myotubes. These sFRP4-mediated responses may result in a vicious cycle, in which enhanced rates of DNL and glycolysis aggravate hepatic lipid accumulation and insulin resistance.

Alteration of Liver Peroxisomal and Mitochondrial Functionality in the NZO Mouse Model of Metabolic Syndrome.

PURPOSE: Metabolic syndrome (MetS) consists of five risk factors: elevated blood pressure and fasting glucose, visceral obesity, dyslipidemia, and hypercholesterinemia. The physiological impact of lipid metabolism indicated as visceral obesity and hepatic lipid accumulation on MetS is still under debate. One major cause of disturbed lipid metabolism might be dysfunction of cellular organelles controlling energy homeostasis, i.e., mitochondria and peroxisomes. EXPERIMENTAL DESIGN: The New Zealand Obese (NZO) mouse model exhibits a polygenic syndrome of obesity, insulin resistance, triglyceridemia, and hypercholesterolemia that resembles human metabolic syndrome. We applied a multi-omics approach combining lipidomics with liver transcriptomics and top-down MS based organelle proteomics (2D-DIGE) of highly enriched mitochondria and peroxisomes in male mice, to investigate molecular mechanisms related to the impact of lipid metabolism in the pathophysiology of the metabolic syndrome. CONCLUSIONS AND CLINICAL RELEVANCE: Proteome analyses of liver organelles indicate differences in fatty acid and cholesterol metabolism, mainly influenced by PG-C1α/PPARα and other nuclear receptor mediated pathways. These results are in accordance with altered serum lipid profiles and elevated organelle functionality. These data emphasize that metabolic syndrome is accompanied with increased mitochondria and peroxisomal activity to cope with dyslipidemia and hypercholesterinemia driven hepatic lipid overflow in developing a fatty liver.

Mechanisms of Insulin Resistance in Primary and Secondary Nonalcoholic Fatty Liver.

Nonalcoholic fatty liver disease is associated with hepatic insulin resistance and may result primarily from increased hepatic de novo lipogenesis (PRIM) or secondarily from adipose tissue lipolysis (SEC). We studied mice with hepatocyte- or adipocyte-specific SREBP-1c overexpression as models of PRIM and SEC. PRIM mice featured increased lipogenic gene expression in the liver and adipose tissue. Their selective, liver-specific insulin resistance was associated with increased C18:1-diacylglycerol content and protein kinase Cε translocation. SEC mice had decreased lipogenesis mediated by hepatic cholesterol responsive element-binding protein and featured portal/lobular inflammation along with total, whole-body insulin resistance. Hepatic mitochondrial respiration transiently increased and declined with aging along with higher muscle reactive oxygen species production. In conclusion, hepatic insulin resistance originates from lipotoxicity but not from lower mitochondrial capacity, which can even transiently adapt to increased peripheral lipolysis. Peripheral insulin resistance is prevented during increased hepatic lipogenesis only if adipose tissue lipid storage capacity is preserved.

Reduced expression of chemerin in visceral adipose tissue associates with hepatic steatosis in patients with obesity.

OBJECTIVE: This study aimed to evaluate whether circulating levels and/or visceral adipose tissue (VAT) expression of recently described adipokines associate with histopathological severity of nonalcoholic fatty liver disease (NAFLD), independent of obesity and insulin resistance. METHODS: Serum levels of adiponectin, omentin, chemerin, monocyte chemoattractant protein-1, and secreted frizzled-related protein 4 were measured using enzyme-linked immunosorbent assay in 81 patients with obesity and NAFLD and 18 lean control subjects. Expression in VAT was measured using real-time PCR and histopathological grading was scored using the NAFLD activity score (NAS). RESULTS: When NAFLD patients were subdivided into groups with simple steatosis, borderline nonalcoholic steatohepatitis (NASH), and NASH, adiponectin serum levels and omentin expression were lower in NASH versus simple steatosis patients. Serum adiponectin was generally lower with higher histopathological grading. Chemerin VAT expression was negatively associated with NAS (r = -0.331, P = 0.022) and steatosis score (r = -0.335, P = 0.020), independent of age, BMI, and HOMA-IR. In addition, adjusting for chemerin VAT expression in a multivariate model explained part of the association between NAS and HOMA-IR. CONCLUSIONS: These findings suggest that lower VAT expression of chemerin in patients with obesity may be involved in the pathophysiology of hepatic steatosis, potentially by modulating the link between insulin resistance and NAFLD.

Effect of Sfrp5 on cytokine release and insulin action in primary human adipocytes and skeletal muscle cells.

Secreted frizzled-related protein 5 (Sfrp5) is an adipokine with anti-inflammatory and insulin-sensitizing properties in mice. However, the mechanism of Sfrp5 action, especially in humans, is largely unknown. Therefore, cytokine release and insulin signaling were analyzed to investigate the impact of Sfrp5 on inflammation and insulin signaling in primary human adipocytes and skeletal muscle cells (hSkMC). Sfrp5 neither affected interleukin (IL)-6, monocyte chemoattractant protein-1 (MCP-1) and adiponectin release from human adipocytes, nor IL-6 and IL-8 release from hSkMC. In tumor necrosis factor (TNF) α-treated adipocytes, Sfrp5 reduced IL-6 release by 49% (p<0.05), but did not affect MCP-1 and adiponectin release. In MCP-1-treated hSkMC, Sfrp5 did not affect cytokine secretion. In untreated adipocytes, Sfrp5 decreased the insulin-mediated phosphorylation of Akt-Ser473, Akt-Thr308, GSK3α-Ser21 and PRAS40-Thr246 by 34% (p<0.01), 31% (p<0.05), 37% (p<0.05) and 34% (p<0.01), respectively, and the stimulation of glucose uptake by 25% (p<0.05). Incubation with TNFα increased the phosphorylation of JNK and NFκB, and impaired insulin signaling. When Sfrp5 and TNFα were combined, there was no additional effect on insulin signaling and JNK phosphorylation, but phosphorylation of NFκB was reversed to basal levels. Sfrp5 had no effect on insulin signaling in untreated or in MCP-1 treated hSkMC. Thus, Sfrp5 lowered IL-6 release and NFκB phosphorylation in cytokine-treated human adipocytes, but not under normal conditions, and decreased insulin signaling in untreated human adipocytes. Sfrp5 did not act on hSkMC. Therefore, the cellular actions of Sfrp5 seem to depend on the type of tissue as well as its inflammatory and metabolic state.