Adipose tissue place of origin and obesity influence sphingolipid signaling pathway in the adipocytes differentiated from ADMSCs isolated from morbidly obese women.

Adipose derived mesenchymal stem cells (ADMSCs) are a component of adipose tissue that in recent years has gained on importance. The progenitor cells serve as an essentially unlimited source of new adipocytes and therefore are considered to be an important determinant of the tissue's physiology. In this paper we investigated mature adipocytes differentiated from ADMSCs obtained from subcutaneous/visceral fat of patients with different metabolic status (lean, obese without and with metabolic syndrome). We focused our interests on the sphingolipid signaling pathway, i.e.a signal transduction system indispensable for cells functioning, but also implicated in the development of medical conditions associated with obesity. We observed that the cells derived from visceral tissue had significantly greater levels of almost all the examined sphingolipids (especially Cer, dhCer, SM). Moreover, obesity and metabolic syndrome present in donor patients was associated with an increased level of sphingosine kinase (SPHK) and the product of its reaction sphingosine-1-phosphate (S1P). Moreover, the condition appeared to display a tissue specific pattern. Namely, the adipocytes of subcutaneous provenance had an increased activation of ceramide de novo synthesis pathway when the donors of ADMSCs had metabolic syndrome. The above translated into greater accumulation of ceramide in the cells. To our knowledge this is the first study that demonstrated altered sphingolipid profile in the mature adipocytes differentiated from ADMSCs with respect to the stem cells tissue of origin and the donor patient metabolic status.

Alteration of zeta potential and cell viability in rat-derived L6 skeletal muscle cells and H9c2 cardiomyocytes: A study with submicron polystyrene particles.

BACKGROUND: Microand nanoplastics pollution can cause substantial damage to ecosystems. Since scientists have focused mainly on their impact on aquatic environments, less attention has been paid to the accumulation of polymer particles in terrestrial organisms. OBJECTIVES: We checked if submicron (<5 mm) polystyrene (PS) particles, which can accumulate in living organisms, lead to changes in the physicochemical properties of mammalian cell membranes. MATERIAL AND METHODS: The influence of submicron PS particles on the properties of rat-derived L6 myocytes and H9c2 cardiomyocytes was analyzed. Non-functionalized and amine-functionalized PS particles of 100 nm and 200 nm in diameter were used. The MTT assay was performed to evaluate the viability of the polymers-treated cells. The effect of short (6 h) and prolonged (48 h) incubation with different concentrations of PS particles on the cell's zeta (ζ) potential was examined with the electrophoretic light scattering technique (ELS). Polystyrene particles' physicochemical characteristics (size and stability) were performed using dynamic light scattering (DLS) and electrophoretic light scattering methods. RESULTS: The results show that submicron PS particles affect cell viability and cause changes in the physiochemical parameters of rat cell membranes. Differences were observed depending on the origin of the cells. We observed doseand time-dependent alterations in the studied parameters after submicron PS particle incubation in L6 myotubes and H9c2 cardiomyocytes. CONCLUSIONS: The size and modification of PS particle surfaces determine the extent to which they affect the analyzed properties of rat cardiomyocytes and myocytes membranes.

Efficacy of adipose-derived mesenchymal stem cell therapy in the treatment of chronic micro- and macrovascular complications of diabetes.

Diabetes mellitus is a highly prevalent disease characterized by hyperglycaemia that damages the vascular system, leading to micro- (retinopathy, neuropathy, nephropathy) and macrovascular diseases (cardiovascular disease). There are also secondary complications of diabetes (cardiomyopathy, erectile dysfunction or diabetic foot ulcers). Stem cell-based therapies have become a promising tool targeting diabetes symptoms and its chronic complications. Among all stem cells, adipose-derived mesenchymal stem cells (ADMSCs) are of great importance because of their abundance, non-invasive isolation and no ethical limitations. Characteristics that make ADMSCs good candidates for cell-based therapy are their wide immunomodulatory properties and paracrine activities through the secretion of an array of growth factors, chemokines, cytokines, angiogenic factors and anti-apoptotic molecules. Besides, after transplantation, ADMSCs show great ex vivo expansion capacity and differentiation to other cell types, including insulin-producing cells, cardiomyocytes, chondrocytes, hepatocyte-like cells, neurons, endothelial cells, photoreceptor-like cells, or astrocytes. Preclinical studies have shown that ADMSC-based therapy effectively improved visual acuity, ameliorated polyneuropathy and foot ulceration, arrested the development and progression of diabetic kidney disease, or alleviated the diabetes-induced cardiomyocyte hypertrophy. However, despite the positive results obtained in animal models, there are still several challenges that need to be overcome before the results of preclinical studies can be translated into clinical applications. To date, there are several clinical trials or ongoing trials using ADMSCs in the treatment of diabetic complications, most of them in the treatment of diabetic foot ulcers. This narrative review summarizes the most recent outcomes on the usage of ADMSCs in the treatment of long-term complications of diabetes in both animal models and clinical trials.

Multivessel Coronary Artery Disease Complicated by Diabetes Mellitus Has a Relatively Small Effect on Endothelial and Lipoprotein Lipases Expression in the Human Atrial Myocardium and Coronary Perivascular Adipose Tissue.

Endothelial (EL) and lipoprotein (LPL) lipases are enzymes involved in lipoproteins metabolism and formation of atherosclerosis, a pathological feature of coronary artery disease (CAD). This paper examines the role of the lipases in the right atrial appendage (RAA) and coronary perivascular adipose tissue (PVAT) of patients with CAD alone or with accompanying diabetes. Additionally, correlation analysis for plasma concentration of the lipases, apolipoproteins (ApoA-ApoJ) and blood lipids (Chol, HDL-C, LDL-C, TAG) was performed. We observed that CAD had little effect on the lipases gene/protein levels in the RAA, while their transcript content was elevated in the PVAT of diabetic CAD patients. Interestingly, the RAA was characterized by higher expression of EL/LPL (EL: +1-fold for mRNA, +5-fold for protein; LPL: +2.8-fold for mRNA, +12-fold for protein) compared to PVAT. Furthermore, ApoA1 plasma concentration was decreased, whereas ApoC1 and ApoH were increased in the patients with CAD and/or diabetes. The concentrations of ApoC3 and ApoD were strongly positively correlated with TAG content in the blood, and the same was true for ApoB with respect to LDL-C and total cholesterol. Although plasma concentrations of EL/LPL were elevated in the patients with diabetes, CAD alone had little effect on blood, myocardial and perivascular fat expression of the lipases.

RabGAP AS160/TBC1D4 deficiency increases long-chain fatty acid transport but has little additional effect on obesity and metabolic syndrome in ADMSCs-derived adipocytes of morbidly obese women.

The Akt substrate of 160 kDa (AS160), also known as TBC1 domain family member 4 (TBC1D4), represents a crucial regulator of insulin-stimulated glucose uptake in skeletal muscle and adipose tissue. Recent evidence suggests that AS160/TBC1D4 may also control the cellular entry of long-chain fatty acids (LCFAs), resulting in changes to the lipid profile of muscles and fat cells in lean subjects. However, there are virtually no data on AS160/TBC1D4 expression and its modulatory role in lipid metabolism in the adipocytes from morbidly obese individuals of different metabolic status. In this study, we evaluated the effect of the three main factors, i.e., AS160 silencing, obesity, and metabolic syndrome on lipid uptake and profile in fully differentiated adipocytes derived from mesenchymal stem cells (ADMSCs) of lean and obese (with/without metabolic syndrome) postmenopausal women. Additionally, we tested possible interactions between the explanatory variables. In general, obesity translated into a greater content of fatty acid transporters (especially CD36/SR-B2 and SLC27A4/FATP4) and boosted accumulation of all the examined lipid fractions, i.e., triacylglycerols (TAGs), diacylglycerols (DAGs), and free fatty acids (FFAs). The aforementioned were further enhanced by metabolic syndrome. Moreover, AS160 deficiency also increased the abundance of SLC27A4/FATP4 and CD36/SR-B2, especially on the cell surface of the adipocytes derived from ADMSCs of subcutaneous deposit. This was further accompanied by increased LCFA (palmitic acid) uptake. Despite the aforementioned, AS160 silencing seemed unable to significantly affect the phenotype of the adipocytes stemming from obese patients with respect to their cellular lipid profile as we observed virtually no changes in TAG, DAG, and FFA contents when compared to cells with the reference level of proteins. Nevertheless, knockdown of AS160 stimulated fatty acid oxidation, which may indicate that adaptive mechanisms counteract excessive lipid accumulation. At the same time, adipocytes of visceral origin were rather insensitive to the applied intervention.

Adipose-derived Mesenchymal Stem Cells Therapy as a new Treatment Option for Diabetes Mellitus.

The worldwide increase in the prevalence of diabetes mellitus has raised the demand for new therapeutic strategies targeting diabetic symptoms and its chronic complications. Among different treatment options for diabetes, adipose-derived mesenchymal stem cells (ADMSCs) therapy attract the most attention. The therapeutic effects of ADMSCs are based primarily on their paracrine release of immunomodulatory, anti-inflammatory, and trophic factors. Animal models of diabetes as well as human clinical trials have shown that ADMSCs can effectively facilitate endogenous ß cell regeneration, preserve residual ß cell mass, reduce islet graft rejection, regulate the immune system, and ultimately improve insulin sensitivity or ameliorate insulin resistance in peripheral tissues. Nevertheless, transplantation of mesenchymal stem cells is associated with certain risks; therefore recently much attention has been devoted to ADMSCs derivatives, such as exosomes or conditioned media, as therapeutic agents for the treatment of diabetes. Compared to ADMSCs, cell-free therapy has even better therapeutic potential. This narrative review summarizes recent outcomes and molecular mechanisms of ADMSCs action in the treatment for both type 1 DM and type 2 DM, as well as shows their feasibility, benefits, and current limitations.

The Phenotype of the Adipocytes Derived from Subcutaneous and Visceral ADMSCs Is Altered When They Originate from Morbidly Obese Women: Is There a Memory Effect?

Adipose tissue is an abundant source of mesenchymal stem cells (ADMSCs). Evidence has suggested that depot-specific ADMSCs (obtained from subcutaneous or visceral adipose tissue-subADMSCs or visADMSCs, respectively) account for differential responses of each depot to metabolic challenges. However, little is known about the phenotype and changes in metabolism of the adipocytes derived from ADMSCs of obese individuals. Therefore, we investigated the phenotypic and metabolic characteristics, particularly the lipid profile, of fully differentiated adipocytes derived from ADMSCs of lean and obese (with/without metabolic syndrome) postmenopausal women. We observed a depot-specific pattern, with more pronounced changes present in the adipocytes obtained from subADMSCs. Namely, chronic oversupply of fatty acids (present in morbid obesity) triggered an increase in CD36/SR-B2 and FATP4 protein content (total and cell surface), which translated to an increased LCFA influx (3H-palmitate uptake). This was associated with the accumulation of TAG and DAG in these cells. Furthermore, we observed that the adipocytes of visADMSCs origin were larger and showed smaller granularity than their counterparts of subADMSCs descent. Although ADMSCs were cultured in vitro, in a fatty acids-deprived environment, obesity significantly influenced the functionality of the progenitor adipocytes, suggesting the existence of a memory effect.

The influence of the pH on the incorporation of caffeic acid into biomimetic membranes and cancer cells.

Caffeic acid (CA) is a phenolic compound synthesized by all plant species. It constitutes the main hydroxycinnamic acid found in human diet and presents a variety of beneficial effects including anticancer activity. Current data suggests essential role of the interplay between anticancer drugs and the cell membrane. Given this, biophysical interactions between CA and cancer cells or biomimetic membranes were investigated. Glioblastoma cell line U118MG and colorectal adenocarcinoma cell line DLD-1, as well as lipid bilayers and liposomes, were used as in vitro models. Electrophoretic light scattering was used to assess the effect of CA on the surface charge of cancer cells and liposomal membranes. Electrochemical impedance spectroscopy was chosen to evaluate CA-dependent modulatory effect on the electrical capacitance and electrical resistance of the bilayers. Our results suggest that CA fulfills physicochemical criteria determining drug-like properties of chemical compounds, and may serve as a potential cytostatic agent in cancer treatment.

Grade-dependent changes in sphingolipid metabolism in clear cell renal cell carcinoma.

There is a host of evidence for the role of bioactive sphingolipids in cancer biology, and dysregulated sphingolipid metabolism was observed in many malignant tumors. The aim of the present study was to provide more detailed data on sphingolipid metabolism in different stages of clear cell renal cell carcinoma (ccRCC). Samples of the tumor and noncancerous fragments of the same kidney were collected from patients who underwent a radical nephrectomy. The subjects were stratified according to the degree of malignancy of the tumor (n = 14 for G2, 12 for G3, and 9 for G4). The content of bioactive sphingolipids/glycosphingolipids was measured with an HPLC and HPTLC method, and the mRNA and protein expression of sphingolipid transporters and metabolizing enzymes was evaluated using real-time polymerase chain reaction (PCR) and Western blot, respectively. Compared to healthy kidney tissue, ccRCC was characterized by accumulation of sphingosine, sphingosine-1-phosphate (S1P), ceramide, dihydrosphingosine, and dihydroceramide. However, in the case of the latter two, the accumulation was limited to higher malignancy grades. In addition, compared to the healthy tissue, the content of gangliosides in the tumor was increased at the expense of globosides. We also found profound grade-dependent changes in the mRNA level of S1P-metabolizing enzymes, and spinster homolog 2. In general, their expression was much higher in G2 tumors compared to higher malignancy grades. We conclude that ccRCC is characterized by profound and multilevel alterations in sphingolipid metabolism, which to a large extent are grade-dependent. We hypothesize that dysregulation of sphingolipid metabolism contributes to the progression of ccRCC.

Using adipose-derived mesenchymal stem cells to fight the metabolic complications of obesity: Where do we stand?

Obesity is a critical risk factor for the development of metabolic diseases, and its prevalence is increasing worldwide. Stem cell-based therapies have become a promising tool for therapeutic intervention. Among them are adipose-derived mesenchymal stem cells (ADMSCs), secreting numerous bioactive molecules, like growth factors, cytokines, and chemokines. Their unique features, including immunosuppressive and immunomodulatory properties, make them an ideal candidates for clinical applications. Numerous experimental studies have shown that ADMSCs can improve pancreatic islet cell viability and function, ameliorate hyperglycemia, improve insulin sensitivity, restore liver function, counteract dyslipidemia, lower pro-inflammatory cytokines, and reduce oxidative stress in the animal models. These results prompted scientists to use ADMSCs clinically. However, up to date, there have been few clinical studies or ongoing trails using ADMSCs to treat metabolic disorders such as type 2 diabetes mellitus (T2DM) or liver cirrhosis. Most human studies have implemented autologous ADMSCs with minimal risk of cellular rejection. Because the functionality of ADMSCs is significantly reduced in subjects with obesity and/or metabolic syndrome, their efficacy is questioned. ADMSCs transplantation may offer a potential therapeutic approach for the treatment of metabolic complications of obesity, but randomized controlled trials are required to establish their safety and efficacy in humans prior to routine clinical use.

Does TBC1D4 (AS160) or TBC1D1 Deficiency Affect the Expression of Fatty Acid Handling Proteins in the Adipocytes Differentiated from Human Adipose-Derived Mesenchymal Stem Cells (ADMSCs) Obtained from Subcutaneous and Visceral Fat Depots?

TBC1D4 (AS160) and TBC1D1 are Rab GTPase-activating proteins that play a key role in the regulation of glucose and possibly the transport of long chain fatty acids (LCFAs) into muscle and fat cells. Knockdown (KD) of TBC1D4 increased CD36/SR-B2 and FABPpm protein expressions in L6 myotubes, whereas in murine cardiomyocytes, TBC1D4 deficiency led to a redistribution of CD36/SR-B2 to the sarcolemma. In our study, we investigated the previously unexplored role of both Rab-GAPs in LCFAs uptake in human adipocytes differentiated from the ADMSCs of subcutaneous and visceral adipose tissue origin. To this end we performed a single- and double-knockdown of the proteins (TBC1D1 and TBC1D4). Herein, we provide evidence that AS160 mediates fatty acid entry into the adipocytes derived from ADMSCs. TBC1D4 KD resulted in quite a few alterations to the cellular phenotype, the most obvious of which was the shift of the CD36/SR-B2 transport protein to the plasma membrane. The above translated into an increased uptake of saturated long-chain fatty acid. Interestingly, we observed a tissue-specific pattern, with more pronounced changes present in the adipocytes derived from subADMSCs. Altogether, our data show that in human adipocytes, TBC1D4, but not TBC1D1, deficiency increases LCFAs transport via CD36/SR-B2 translocation.

Treadmill Running Changes Endothelial Lipase Expression: Insights from Gene and Protein Analysis in Various Striated Muscle Tissues and Serum.

Endothelial lipase (EL) is an enzyme capable of HDL phospholipids hydrolysis. Its action leads to a reduction in the serum high-density lipoprotein concentration, and thus, it exerts a pro-atherogenic effect. This study examines the impact of a single bout exercise on the gene and protein expression of the EL in skeletal muscles composed of different fiber types (the soleus-mainly type I, the red gastrocnemius-mostly IIA, and the white gastrocnemius-predominantly IIX fibers), as well as the diaphragm, and the heart. Wistar rats were subjected to a treadmill run: (1) t = 30 [min], V = 18 [m/min]; (2) t = 30 [min], V = 28 [m/min]; (3) t = 120 [min], V = 18 [m/min] (designated: M30, F30, and M120, respectively). We established EL expression in the total muscle homogenates in sedentary animals. Resting values could be ordered with the decreasing EL protein expression as follows: endothelium of left ventricle > diaphragm > red gastrocnemius > right ventricle > soleus > white gastrocnemius. Furthermore, we observed that even a single bout of exercise was capable of inducing changes in the mRNA and protein level of EL, with a clearer pattern observed for the former. After 30 min of running at either exercise intensity, the expression of EL transcript in all the cardiovascular components of muscles tested, except the soleus, was reduced in comparison to the respective sedentary control. The protein content of EL varied with the intensity and/or duration of the run in the studied whole tissue homogenates. The observed differences between EL expression in vascular beds of muscles may indicate the muscle-specific role of the lipase.

Does supplementation with leucine-enriched protein alone and in combination with fish-oil-derived n-3 PUFA affect muscle mass, strength, physical performance, and muscle protein synthesis in well-nourished older adults? A randomized, double-blind, placebo-controlled trial.

BACKGROUND: Leucine-enriched protein (LEU-PRO) and long-chain (LC) n-3 (ω-3) PUFAs have each been proposed to improve muscle mass and function in older adults, whereas their combination may be more effective than either alone. OBJECTIVE: The impact of LEU-PRO supplementation alone and combined with LC n-3 PUFAs on appendicular lean mass, strength, physical performance and myofibrillar protein synthesis (MyoPS) was investigated in older adults at risk of sarcopenia. METHODS: This 24-wk, 3-arm parallel, randomized, double-blind, placebo-controlled trial was conducted in 107 men and women aged ≥65 y with low muscle mass and/or strength. Twice daily, participants consumed a supplement containing either LEU-PRO (3 g leucine, 10 g protein; n = 38), LEU-PRO plus LC n-3 PUFAs (0.8 g EPA, 1.1 g DHA; LEU-PRO+n-3; n = 38), or an isoenergetic control (CON; n = 31). Appendicular lean mass, handgrip strength, leg strength, physical performance, and circulating metabolic and renal function markers were measured pre-, mid-, and postintervention. Integrated rates of MyoPS were assessed in a subcohort (n = 28). RESULTS: Neither LEU-PRO nor LEU-PRO+n-3 supplementation affected appendicular lean mass, handgrip strength, knee extension strength, physical performance or MyoPS. However, isometric knee flexion peak torque (treatment effect: -7.1 Nm; 95% CI: -12.5, -1.8 Nm; P < 0.01) was lower postsupplementation in LEU-PRO+n-3 compared with CON. Serum triacylglycerol and total adiponectin concentrations were lower, and HOMA-IR was higher, in LEU-PRO+n-3 compared with CON postsupplementation (all P < 0.05). Estimated glomerular filtration rate was higher and cystatin c was lower in LEU-PRO and LEU-PRO+n-3 postsupplementation compared with CON (all P < 0.05). CONCLUSIONS: Contrary to our hypothesis, we did not observe a beneficial effect of LEU-PRO supplementation alone or combined with LC n-3 PUFA supplementation on appendicular lean mass, strength, physical performance or MyoPS in older adults at risk of sarcopenia. This trial was registered at clinicaltrials.gov as NCT03429491.

Pyrroloquinoline Quinone Modifies Lipid Profile, but Not Insulin Sensitivity, of Palmitic Acid-Treated L6 Myotubes.

Pyrroloquinoline quinone (PQQ) is a novel stimulator of mitochondrial biogenesis and cellular energy metabolism. This is the first study investigating regulatory mechanisms and metabolic responses underlying PQQ's action in palmitate-exposed L6 myotubes. Particularly, we assessed alterations in lipid content and composition, expression of metabolic enzymes, and changes in glucose transport. The experiments were conducted using muscle cells subjected to short (2 h) and prolonged (24 h) incubation with PQQ in a sequence of pre- and post-palmitic acid (PA) exposure. We demonstrated the opposite effects of 2 and 24 h treatments with PQQ on lipid content, i.e., a decline in the level of free fatty acids and triacylglycerols in response to short-time PQQ incubation as compared to increases in diacylglycerol and triacylglycerol levels observed after 24 h. We did not demonstrate a significant impact of PQQ on fatty acid transport. The analysis of metabolic enzyme expression showed that the vast majority of PQQ-dependent alterations cumulated in the PA/PQQ 24 h group, including elevated protein amount of peroxisome proliferator activated receptor γ co-activator 1α (PGC-1α), sirtuin-1 (SIRT1), phosphorylated 5'AMP-activated protein kinase (pAMPK), carnitine palmitoyltransferase I (CPT1), citrate synthase (CS), fatty acid synthase (FAS), and serine palmitoyltransferase, long chain base subunit 1 (SPT1). In conclusion, the results mentioned above indicate PQQ-dependent activation of both fatty acid oxidation and lipid synthesis in order to adapt cells to palmitic acid-rich medium, although PQQ did not attenuate insulin resistance in muscle cells.

Expression of zinc transporter 8 in thyroid tissues from patients with immune and non-immune thyroid diseases.

INTRODUCTION: Recent studies have revealed the presence of zinc and the expression of zinc transporter (ZnT) family members in most endocrine cell types. It was demonstrated that ZnT family plays an important role in the synthesis and secretion of many hormones. Moreover, recently ZnT8 was described as a newly islet autoantigen in type 1 diabetes. MATERIALS AND METHODS: We studied the expression of ZnT8 transporter in thyroid tissues from patients with immune and non-immune thyroid diseases. The study was performed in thyroid tissues after thyroidectomy from patients with thyroid non-toxic nodular goitre (NTNG; n = 17, mean age 15.8 ± 2.2 years) and cases with Graves' disease (n = 20, mean age 15.6 ± 2.8). In our study we investigated the expression of ZnT8 in human thyroid tissues from patients with immune and non-immune thyroid diseases using immunohistochemistry, Western Blot as well as immunofluorescence analyses. To the best of our knowledge, this is the first investigation which identified ZnT8 protein expression in human thyroid tissues, moreover, confirmed by three different laboratory techniques. Results and Conclusions Expression of ZnT8 transporter was identified by immunohistochemistry in the thyroid tissues from paediatric patients with Graves' disease (on +++) and non-toxic nodular goitre (on ++). ZnT8 transporter expression was found both in thyroid follicular cells (within the cytoplasm and cytoplasmic membrane in follicular cells) and C cells (membrane-cytoplasmic reaction) in fluorescence. Predominant expression of ZnT8 in band 41 kDa in immune than in non-immune thyroid disorders may suggest potential role of ZnT8 as a new thyroid autoanitgen but it requires further study on a larger cohort.

The Gene and Protein Expression of the Main Components of the Lipolytic System in Human Myocardium and Heart Perivascular Adipose Tissue. Effect of Coronary Atherosclerosis.

The aim of our study was to examine the regulation of triacylglycerols (TG) metabolism in myocardium and heart perivascular adipose tissue in coronary atherosclerosis. Adipose triglyceride lipase (ATGL) is the major TG-hydrolase. The enzyme is activated by a protein called comparative gene identification 58 (CGI-58) and inhibited by a protein called G0/G1 switch protein 2 (G0S2). Samples of the right atrial appendage and perivascular adipose tissue were obtained from two groups of patients: 1-with multivessel coronary artery disease qualified for coronary artery bypass grafting (CAD), 2-patients with no atherosclerosis qualified for a valve replacement (NCAD). The mRNA and protein analysis of ATGL, HSL, CGI-58, G0S2, FABP4, FAT/CD36, LPL, ß-HAD, CS, COX4/1, FAS, SREBP-1c, GPAT1, COX-2, 15-LO, and NFκß were determined by using real-time PCR and Western Blot. The level of lipids (i.e., TG, diacylglycerol (DG), and FFA) was examined by GLC. We demonstrated that in myocardium coronary atherosclerosis increases only the transcript level of G0S2 and FABP4. Most importantly, ATGL, ß-HAD, and COX4/1 protein expression was reduced and it was accompanied by over double the elevation in TG content in the CAD group. The fatty acid synthesis and their cellular uptake were stable in the myocardium of patients with CAD. Additionally, the expression of proteins contributing to inflammation was increased in the myocardium of patients with coronary stenosis. Finally, in the perivascular adipose tissue, the mRNA of G0S2 was elevated, whereas the protein content of FABP-4 was increased and for COX4/1 diminished. These data suggest that a reduction in ATGL protein expression leads to myocardial steatosis in patients with CAD.

Assessment of the Main Compounds of the Lipolytic System in Treadmill Running Rats: Different Response Patterns between the Right and Left Ventricle.

The aim of the present study was to investigate the time and intensity dependent effects of exercise on the heart components of the lipolytic complex. Wistar rats ran on a treadmill with the speed of 18 m/min for 30 min (M30) or 120 min (M120) or with the speed of 28 m/min for 30 min (F30). The mRNA and protein expressions of the compounds adipose triglyceride lipase (ATGL), comparative gene identification-58 (CGI-58), G0/G1 switch gene 2 (G0S2), hormone sensitive lipase (HSL) and peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α) were examined by real-time PCR and Western blot, respectively. Lipid content of free fatty acids (FFA), diacylglycerols (DG) and triacylglycerols (TG) were estimated by gas liquid chromatography. We observed virtually no changes in the left ventricle lipid contents and only minor fluctuations in its ATGL mRNA levels. This was in contrast with its right counterpart i.e., the content of TG and DG decreased in response to both increased duration and intensity of a run. This occurred in tandem with increased mRNA expression for ATGL, CGI-58 and decreased expression of G0S2. It is concluded that exercise affects behavior of the components of the lipolytic system and the lipid content in the heart ventricles. However, changes observed in the left ventricle did not mirror those in the right one.

Metabolic Syndrome is Associated with Ceramide Accumulation in Visceral Adipose Tissue of Women with Morbid Obesity.

OBJECTIVE: Accelerated transmembrane transport of long-chain fatty acids dependent on fatty acid transporters is responsible for lipid accumulation and, eventually, the development of metabolic syndrome. This study determined the content of lipids (ceramide [CER], diacylglycerol [DAG], triacylglycerol, and free fatty acid [FFA]) and the expression of fatty acid translocase (FAT/CD36) and plasma membrane fatty acid-binding protein in visceral adipose tissue (VAT) and subcutaneous adipose tissue of women with morbid obesity without metabolic syndrome (MetSx-) or with metabolic syndrome (MetSx+) and compared the results with those of lean controls without metabolic syndrome. METHODS: Lipid content and fatty acid composition in each lipid subclass were estimated by gas liquid chromatography. For total, plasma membrane, and mitochondrial expression of fatty acid transporters, subfractionation with subsequent Western blot technique was used. RESULTS: A greater content of triacylglycerol in VAT of participants with obesity (MetSx-) was found. However, only the MetSx+ subjects had increased content of CER in VAT in relation to subcutaneous adipose tissue in MetSx+ and lean individuals. This was accompanied by increased total and membrane expression of FAT/CD36 in VAT in MetSx+ subjects. Accordingly, mitochondrial expression of FAT/CD36 and plasma membrane fatty acid-binding protein was decreased in both groups of subjects with obesity. CONCLUSIONS: Metabolic syndrome is associated with the accumulation of CER in VAT, possibly related to increased FAT/CD36 protein expression.

Dose- and time-dependent alterations in lipid metabolism after pharmacological PGC-1α activation in L6 myotubes.

Pyrroloquinoline quinone (PQQ) acts as a powerful modulator of PGC-1α activation and therefore regulates multiple pathways involved in cellular energy homeostasis. In the present study, we assessed the effects of L6 myotubes incubation with 0.5, 1, and 3 µM PQQ solution for 2 and 24 hr with respect to the cells' lipid metabolism. We demonstrated that PQQ significantly elevates PGC-1α content in a dose- and time-dependent manner with the highest efficiency for 0.5 and 1 µM. The level of free fatty acids was diminished (24 hr: -66%), while an increase in triacylglycerol (TAG) amount was most pronounced after 0.5 µM (2 hr: +93%, 24 hr: +139%) treatment. Ceramide (CER) content was elevated after 2 hr incubation with 0.5 µM and after prolonged exposure to all PQQ concentrations. The cells treated with PQQ for 2 hr exhibited decreased sphinganine (SFA) and sphinganine-1-phosphate (SFA1P) level, while 24 hr incubation resulted in an elevated sphingosine (SFO) amount. In summary, PGC-1α activation promotes TAG and CER synthesis.

Cerulein-Induced Acute Pancreatitis Affects Sphingomyelin Signaling Pathway in Rats.

OBJECTIVES: Acute pancreatitis (AP) is a common and severe gastrointestinal inflammatory disease with poorly understood pathogenesis. We adopted cerulein-induced pancreatitis, a well-established rat model shearing similarities with human AP, to determine the disease background. Special interest was placed on sphingolipids, because their signaling pathways are involved in many pathological states including hepatic steatosis, heart infarction, or pancreatic origin type 1 diabetes. METHODS: Sphingolipid levels in the blood and pancreas were determined by the means of chromatography (thin-layer and high-performance liquid chromatography). RESULTS: We found that AP leads to activation of ceramide de novo synthesis pathway, as evidenced by a significant increment in sphinganine, that is, ceramide synthesis precursor, content (+3.8-fold). Surprisingly, despite the reported growth in sphinganine concentration, we observed a reduced (-38%) ceramide level in the pancreas of rats with AP. The results could be explained by subsequent hydrolysis of ceramide to other secondary messengers, that is, sphingosine (+4-fold) or sphingosine-1-phosphate (+3-fold). CONCLUSIONS: Because it is known that sphingosine-1-phosphate and some of its analogs could have a protective role against AP complications, our findings may contribute to elaboration of new therapeutic strategies in the management of this severe medical condition.