MSC-induced lncRNA AGAP2-AS1 promotes stemness and trastuzumab resistance through regulating CPT1 expression and fatty acid oxidation in breast cancer.

Trastuzumab resistance has been becoming a major obstacle for treatment of HER-2-positive breast cancer patients. Increasing evidence suggests that mesenchymal stem cells (MSCs) play critical roles during the formation of drug resistance, however, the underlying mechanism is not well known. In this study, mass spectrometry, RNA pulldown and RNA immunoprecipitation assays were performed to verify the direct interactions among AGAP2-AS1 and other associated targets, such as human antigen R (HuR), miR-15a-5p, and carnitine palmitoyl transferase 1 (CPT1). In vitro and in vivo experimental assays were done to clarify the functional role of AGAP2-AS1 in trastuzumab resistance, stemness, and fatty acid oxidation (FAO). The results showed that MSC co-culture induced trastuzumab resistance. AGAP2-AS1 was upregulated in MSC-cultured cells, and knockdown of AGAP2-AS1 reversed the MSC-mediated trastuzumab resistance. Furthermore, MSC culture-induced AGAP2-AS1 regulates stemness and trastuzumab resistance via activating FAO. Mechanistically, AGAP2-AS1 is associated with HuR, and the AGAP2-AS1-HuR complex could directly bind to the CPT1, increasing its expression via improving RNA stability. In addition, AGAP2-AS1 could serve as ceRNA via sponging miR-15a-5p and releasing CPT1 mRNA. Clinically, increased expression of serum AGAP2-AS1 predicts poor response to trastuzumab treatment in breast cancer patients. In conclusion, MSC culture-induced AGAP2-AS1 caused stemness and trastuzumab resistance via promoting CPT1 expression and inducing FAO. Our results provide new insight of the role of MSCs in trastuzumab resistance and AGAP2-AS1 could be promising predictive biomarker and therapeutic target for HER-2+ breast cancer patients.

Modulation of adipocyte size and fat pad weight via resveratrol releasing scaffolds implanted into the epididymal adipose tissue.

Lipid overload of the adipose tissue, which can be caused by overnutrition, underlies metabolic disease. We hypothesized that increasing the energy demand of adipose tissue is a promising strategy to combat excessive lipid accumulation. Resveratrol, a natural polyphenol, activates lipid catabolism in fat tissue; however, its clinical success is hindered by poor bioavailability. Here, we implanted resveratrol releasing poly(lactide-co-glycolide) scaffolds into epididymal fat to overcome its poor bioavailability with the goal of enhancing local lipid catabolism. In lean mice, resveratrol scaffolds decreased adipocyte size relative to scaffolds with no drug, a response that correlated with AMP kinase activation. Immunohistochemistry indicated that macrophages and multinucleated giant cells within the scaffold expressed carnitine palmitoyltransferase 1 (CPT1) at higher levels than other cells in the adipose tissue. Furthermore, resveratrol increased CPT1 levels in cultured macrophages. Taken together, we propose that resveratrol scaffolds decrease adipocyte size because resveratrol increases lipid utilization in scaffold-infiltrating immune cells, possibly through elevating CPT1 levels or activity. In a follow-up study, mice that received resveratrol scaffolds 28-day prior to a high-fat diet exhibited decreased weight gain, adipose tissue expansion, and adipocyte hypertrophy compared to mice with control scaffolds. Notably, this scaffold-based strategy required a single resveratrol administration compared to the daily regiment generally needed for oral administration. These results indicate that localized delivery of metabolism modulating agents to the adipose tissue may overcome issues with bioavailability and that the role of biomaterials should be further investigated in this therapeutic strategy for metabolic disease.

LRRK2 Regulates CPT1A to Promote ß-Oxidation in HepG2 Cells.

Leucine-rich repeat kinase 2 (LRRK2) is involved in lipid metabolism; however, the role of LRRK2 in lipid metabolism to affect non-alcoholic fatty liver disease (NAFLD) is still unclear. In the mouse model of NAFLD induced by a high-fat diet, we observed that LRRK2 was decreased in livers. In HepG2 cells, exposure to palmitic acid (PA) down-regulated LRRK2. Overexpression and knockdown of LRRK2 in HepG2 cells were performed to further investigate the roles of LRRK2 in lipid metabolism. Our results showed that ß-oxidation in HepG2 cells was promoted by LRRK2 overexpression, whereas LRRK2 knockdown inhibited ß-oxidation. The critical enzyme of ß-oxidation, carnitine palmitoyltransferase 1A (CPT1A), was positively regulated by LRRK2. Our data suggested that the regulation of CPT1A by LRRK2 may be via the activation of AMP-activated protein kinase (AMPK) and peroxisome proliferator-activated receptor α (PPARα). The overexpression of LRRK2 reduced the concentration of a pro-inflammatory cytokine, tumor necrosis factor α (TNFα), induced by PA. The increase in ß-oxidation may promote lipid catabolism to suppress inflammation induced by PA. These results indicated that LRRK2 participated in the regulation of ß-oxidation and suggested that the decreased LRRK2 may promote inflammation by suppressing ß-oxidation in the liver.

CPT1a gene expression reverses the inflammatory and anti-phagocytic effect of 7-ketocholesterol in RAW264.7 macrophages.

BACKGROUND: Macrophage are specialized cells that contributes to the removal of detrimental contents via phagocytosis. Lipid accumulation in macrophages, whether from phagocytosis of dying cells or from circulating oxidized low-density lipoproteins, alters macrophage biology and functionality. It is known that carnitine palmitoyl transferase 1-a (CPT1a) gene encodes an enzyme involved in fatty acid oxidation and, therefore, lipid content. However, the potential of CPT1a to activate macrophage phagocytic function have not been elucidated. METHODS: Using a murine macrophage cell line, RAW264.7, we determine if intracellular accumulation of 7-ketocholesterol (7-KC) modulates macrophage phagocytic function through CPT1a gene expression. In addition, the effects of CPT1a genetic modification on macrophage phenotype and phagocytosis has been studied. RESULTS: Our results revealed that CPT1a gene expression decreased by the accumulation of 7-KC at the higher dose of 7-KC. This was concomitant with an impair ability to phagocytize bioparticles and an inflammatory phenotype. GW3965 treatment, which have shown to facilitate the efflux of cholesterol, eliminated the intracellular lipid droplets of 7-KC-laden macrophages, increased the gene expression of CPT1a, diminished the gene expression of the inflammatory marker iNOS and restored macrophage phagocytosis. Furthermore, CPT1a Knockdown per se was detrimental for macrophage phagocytosis whereas transcriptional activation of CPT1a heightened the uptake of bioparticles. CONCLUSIONS: Altogether, our findings indicate that downregulation of CPT1a by lipid content modulates macrophage phagocytosis and inflammatory phenotype.

CPT1A Supports Castration-Resistant Prostate Cancer in Androgen-Deprived Conditions.

Prostate cancer (PCa) is the most common cancer in men, and the global burden of the disease is rising. The majority of PCa deaths are due to metastasis that are highly resistant to current hormonal treatments; this state is called castration-resistant prostate cancer (CRPC). In this study, we focused on the role of the lipid catabolism enzyme CPT1A in supporting CRPC growth in an androgen-dependent manner. We found that androgen withdrawal promoted the growth of CPT1A over-expressing (OE) tumors while it decreased the growth of CPT1A under-expressing (KD) tumors, increasing their sensitivity to enzalutamide. Mechanistically, we found that CPT1A-OE cells burned more lipid and showed increased histone acetylation changes that were partially reversed with a p300 specific inhibitor. Conversely, CPT1A-KD cells showed less histone acetylation when grown in androgen-deprived conditions. Our results suggest that CPT1A supports CRPC by supplying acetyl groups for histone acetylation, promoting growth and antiandrogen resistance.

Identifying off-target effects of etomoxir reveals that carnitine palmitoyltransferase I is essential for cancer cell proliferation independent of ß-oxidation.

It has been suggested that some cancer cells rely upon fatty acid oxidation (FAO) for energy. Here we show that when FAO was reduced approximately 90% by pharmacological inhibition of carnitine palmitoyltransferase I (CPT1) with low concentrations of etomoxir, the proliferation rate of various cancer cells was unaffected. Efforts to pharmacologically inhibit FAO more than 90% revealed that high concentrations of etomoxir (200 µM) have an off-target effect of inhibiting complex I of the electron transport chain. Surprisingly, however, when FAO was reduced further by genetic knockdown of CPT1, the proliferation rate of these same cells decreased nearly 2-fold and could not be restored by acetate or octanoic acid supplementation. Moreover, CPT1 knockdowns had altered mitochondrial morphology and impaired mitochondrial coupling, whereas cells in which CPT1 had been approximately 90% inhibited by etomoxir did not. Lipidomic profiling of mitochondria isolated from CPT1 knockdowns showed depleted concentrations of complex structural and signaling lipids. Additionally, expression of a catalytically dead CPT1 in CPT1 knockdowns did not restore mitochondrial coupling. Taken together, these results suggest that transport of at least some long-chain fatty acids into the mitochondria by CPT1 may be required for anabolic processes that support healthy mitochondrial function and cancer cell proliferation independent of FAO.

Hypothalamic Regulation of Liver and Muscle Nutrient Partitioning by Brain-Specific Carnitine Palmitoyltransferase 1C in Male Mice.

Carnitine palmitoyltransferase (CPT) 1C, a brain-specific protein localized in the endoplasmic reticulum of neurons, is expressed in almost all brain regions. Based on global knockout (KO) models, CPT1C has demonstrated relevance in hippocampus-dependent spatial learning and in hypothalamic regulation of energy balance. Specifically, it has been shown that CPT1C is protective against high-fat diet-induced obesity (DIO), and that CPT1C KO mice show reduced peripheral fatty acid oxidation (FAO) during both fasting and DIO. However, the mechanisms mediating CPT1C-dependent regulation of energy homeostasis remain unclear. Here, we focus on the mechanistic understanding of hypothalamic CPT1C on the regulation of fuel selection in liver and muscle of male mice during energy deprivation situations, such as fasting. In CPT1C-deficient mice, modulation of the main hypothalamic energy sensors (5' adenosine monophosphate-activated protein kinase, Sirtuin 1, and mammalian target of rapamycin) was impaired and plasma catecholamine levels were decreased. Consequently, CPT1C-deficient mice presented defective fasting-induced FAO in liver, leading to higher triacylglycerol accumulation and lower glycogen levels. Moreover, muscle pyruvate dehydrogenase activity was increased, which was indicative of glycolysis enhancement. The respiratory quotient did not decrease in CPT1C KO mice after 48 hours of fasting, confirming a defective switch on fuel substrate selection under hypoglycemia. Phenotype reversion studies identified the mediobasal hypothalamus (MBH) as the main area mediating CPT1C effects on fuel selection. Overall, our data demonstrate that CPT1C in the MBH is necessary for proper hypothalamic sensing of a negative energy balance and fuel partitioning in liver and muscle.

Speract, a sea urchin egg peptide that regulates sperm motility, also stimulates sperm mitochondrial metabolism.

Sea urchin sperm have only one mitochondrion, that in addition to being the main source of energy, may modulate intracellular Ca(2+) concentration ([Ca(2+)]i) to regulate their motility and possibly the acrosome reaction. Speract is a decapeptide from the outer jelly layer of the Strongylocentrotus purpuratus egg that upon binding to its receptor in the sperm, stimulates sperm motility, respiration and ion fluxes, among other physiological events. Altering the sea urchin sperm mitochondrial function with specific inhibitors of this organelle, increases [Ca(2+)]i in an external Ca(2+) concentration ([Ca(2+)]ext)-dependent manner (Ardón, et al., 2009. BBActa 1787: 15), suggesting that the mitochondrion is involved in sperm [Ca(2+)]i homeostasis. To further understand the interrelationship between the mitochondrion and the speract responses, we measured mitochondrial membrane potential (ΔΨ) and NADH levels. We found that the stimulation of sperm with speract depolarizes the mitochondrion and increases the levels of NADH. Surprisingly, these responses are independent of external Ca(2+) and are due to the increase in intracellular pH (pHi) induced by speract. Our findings indicate that speract, by regulating pHi, in addition to [Ca(2+)]i, may finely modulate mitochondrial metabolism to control motility and ensure that sperm reach the egg and fertilize it.

Role of peroxisome proliferator-activated receptor alpha in the expression of hepatic fatty acid oxidation-related genes in chickens.

Liver is the most important target organ for investigation of lipid metabolism in domestic fowls. However, little is known about the regulatory mechanism of fatty acid oxidation in chicken liver. In mammals, proliferator-activated receptor alpha (PPARα), a transcription factor, plays an essential role in the regulation of hepatic fatty acid oxidation. The aim of the present study was to investigate the regulatory mechanisms of PPARα-induced gene expression involved in hepatic fatty acid oxidation in chickens in vivo and in vitro. WY14643, a PPARα agonist, significantly increased the messenger RNA (mRNA) levels of carnitine palmitoyltransferase 1a (CPT1a) and acyl-coenzyme A oxidase (ACO), but not long-, middle- and short-chain acyl-coenzyme A dehydrogenase (LCAD, MCAD and SCAD, respectively), hydroxyacyl-coenzyme A dehydrogenase (HAD), and PPARα itself in chicken hepatoma cells. In contrast, WY14643 significantly increased the mRNA levels of CPT1a, ACO, MCAD, SCAD, HAD and PPARα in human hepatoma cells. The mRNA levels of CPT1a and ACO in the liver were significantly increased by 6 h of fasting in chickens, whereas the mRNA levels of LCAD, MCAD, SCAD and HAD were unchanged. These results suggest that, unlike in mammals, CPT1a and ACO might play an important role in PPARα-induced fatty acid oxidation in the liver of chickens.

Carnitine palmitoyltransferase 1C: From cognition to cancer.

Carnitine palmitoyltransferase 1 (CPT1) C was the last member of the CPT1 family of genes to be discovered. CPT1A and CPT1B were identified as the gate-keeper enzymes for the entry of long-chain fatty acids (as carnitine esters) into mitochondria and their further oxidation, and they show differences in their kinetics and tissue expression. Although CPT1C exhibits high sequence similarity to CPT1A and CPT1B, it is specifically expressed in neurons (a cell-type that does not use fatty acids as fuel to any major extent), it is localized in the endoplasmic reticulum of cells, and it has minimal CPT1 catalytic activity with l-carnitine and acyl-CoA esters. The lack of an easily measurable biological activity has hampered attempts to elucidate the cellular and physiological role of CPT1C but has not diminished the interest of the biomedical research community in this CPT1 isoform. The observations that CPT1C binds malonyl-CoA and long-chain acyl-CoA suggest that it is a sensor of lipid metabolism in neurons, where it appears to impact ceramide and triacylglycerol (TAG) metabolism. CPT1C global knock-out mice show a wide range of brain disorders, including impaired cognition and spatial learning, motor deficits, and a deregulation in food intake and energy homeostasis. The first disease-causing CPT1C mutation was recently described in humans, with Cpt1c being identified as the gene causing hereditary spastic paraplegia. The putative role of CPT1C in the regulation of complex-lipid metabolism is supported by the observation that it is highly expressed in certain virulent tumor cells, conferring them resistance to glucose- and oxygen-deprivation. Therefore, CPT1C may be a promising target in the treatment of cancer. Here we review the molecular, biochemical, and structural properties of CPT1C and discuss its potential roles in brain function, and cancer.

Important role of ventromedial hypothalamic carnitine palmitoyltransferase-1a in the control of food intake.

Carnitine palmitoyltransferase-1 (CPT-1) liver isoform, or CPT-1a, is implicated in CNS control of food intake. However, the exact brain nucleus site(s) in mediating this action of CPT-1a has not been identified. In this report, we assess the role of CPT-1a in hypothalamic ventromedial nucleus (VMN). We stereotaxically injected an adenoviral vector containing CPT-1a coding sequence into the VMN of rats to induce overexpression and activation of CPT-1a. The VMN-selective activation of CPT-1a induced an orexigenic effect, suggesting CPT-1a in the VMN is involved in the central control of feeding. Intracerebroventricular administration of etomoxir, a CPT-1 inhibitor, decreases food intake. Importantly, in the animals with VMN overexpression of a CPT-1a mutant that antagonizes the CPT-1 inhibition by etomoxir, the anorectic response to etomoxir was attenuated. This suggests that VMN is involved in mediating the anorectic effect of central inhibition of CPT-1a. In contrast, arcuate nucleus (Arc) overexpression of the mutant did not alter etomoxir-induced inhibition of food intake, suggesting that Arc CPT-1a does not play significant roles in this anorectic action. Furthermore, in the VMN, CPT-1a appears to act downstream of hypothalamic malonyl-CoA action of feeding. Finally, we show that in the VMN CPT-1 activity was altered in concert with fasting and refeeding states, supporting a physiological role of CPT-1a in mediating the control of feeding. All together, CPT-1a in the hypothalamic VMN appears to play an important role in central control of food intake. VMN-selective modulation of CPT-1a activity may therefore be a promising strategy in controlling food intake and maintaining normal body weight.

Metabolomic analysis reveals that carnitines are key regulatory metabolites in phase transition of the locusts.

Phenotypic plasticity occurs prevalently and plays a vital role in adaptive evolution. However, the underlying molecular mechanisms responsible for the expression of alternate phenotypes remain unknown. Here, a density-dependent phase polyphenism of Locusta migratoria was used as the study model to identify key signaling molecules regulating the expression of phenotypic plasticity. Metabolomic analysis, using high-performance liquid chromatography and gas chromatography-mass spectrometry, showed that solitarious and gregarious locusts have distinct metabolic profiles in hemolymph. A total of 319 metabolites, many of which are involved in lipid metabolism, differed significantly in concentration between the phases. In addition, the time course of changes in the metabolic profiles of locust hemolymph that accompany phase transition was analyzed. Carnitine and its acyl derivatives, which are involved in the lipid ß-oxidation process, were identified as key differential metabolites that display robust correlation with the time courses of phase transition. RNAi silencing of two key enzymes from the carnitine system, carnitine acetyltransferase and palmitoyltransferase, resulted in a behavioral transition from the gregarious to solitarious phase and the corresponding changes of metabolic profiles. In contrast, the injection of exogenous acetylcarnitine promoted the acquisition of gregarious behavior in solitarious locusts. These results suggest that carnitines mediate locust phase transition possibly through modulating lipid metabolism and influencing the nervous system of the locusts.

Malonyl-CoA mediates leptin hypothalamic control of feeding independent of inhibition of CPT-1a.

Hypothalamic fatty acid metabolism is involved in central nervous system controls of feeding and energy balance. Malonyl-CoA, an intermediate of fatty acid biosynthesis, is emerging as a significant player in these processes. Notably, hypothalamic malonyl-CoA has been implicated in leptin's feeding effect. Leptin treatment increases malonyl-CoA level in the hypothalamic arcuate nucleus (Arc), and this increase is required for leptin-induced decrease in food intake. However, the intracellular downstream mediators of malonyl-CoA's feeding effect have not been identified. A primary biochemical action of malonyl-CoA is the inhibition of the acyltransferase activity of carnitine palmitoyltransferase-1 (CPT-1). In the hypothalamus, the predominant isoform of CPT-1 that possesses the acyltransferase activity is CPT-1 liver type (CPT-1a). To address the role of CPT-1a in malonyl-CoA's anorectic action, we used a recombinant adenovirus expressing a mutant CPT-1a that is insensitive to malonyl-CoA inhibition. We show that Arc overexpression of the mutant CPT-1a blocked the malonyl-CoA-mediated inhibition of CPT-1 activity. However, the overexpression of this mutant did not affect the anorectic actions of leptin or central cerulenin for which an increase in Arc malonyl-CoA level is also required. Thus, CPT-1a does not appear to be involved in the malonyl-CoA's anorectic actions induced by leptin. Furthermore, long-chain fatty acyl-CoAs, substrates of CPT-1a, dissociate from malonyl-CoA's actions in the Arc under different feeding states. Together, our results suggest that Arc intracellular mechanisms of malonyl-CoA's anorectic actions induced by leptin are independent of CPT-1a. The data suggest that target(s), rather than CPT-1a, mediates malonyl-CoA action on feeding.

Hepatitis C virus protein and iron overload induce hepatic steatosis through the unfolded protein response in mice.

BACKGROUND/AIM: Hepatic iron overload and steatosis play critical roles in the progression of hepatitis C virus (HCV)-associated chronic liver disease. However, how these two pathophysiological features affect each other remains unknown. The aim of this study was to investigate how hepatic iron overload contributes to the development of hepatic steatosis in the presence of HCV proteins. METHODS: Male C57BL/6 transgenic mice expressing the HCV polyprotein and nontransgenic littermates were fed an excess-iron diet or a control diet. Mice in each group were assessed for the molecules responsible for fat accumulation in the liver. RESULTS: Hepatic iron levels were positively correlated with triglyceride concentrations in the liver for all mice. As compared with the livers of nontransgenic mice fed the control diet, the livers of transgenic mice fed the excess-iron diet showed a lower expression of carnitine palmitoyl transferase I, a higher expression of sterol-regulatory element-binding protein 1 and fatty acid synthetase and an activated unfolded protein response indicated by a higher expression of unspliced and spliced X-box DNA-binding protein 1 (XBP-1), phosphorylated eukaryotic initiation factor-2alpha (p-eIF2alpha), CCAAT/enhancer-binding protein homology protein (CHOP) and abundant autophagosomes concomitant with increased production of reactive oxygen species. Six-month treatment with the anti-oxidant N-acetyl cysteine dramatically reduced hepatic steatosis in transgenic mice fed the excess-iron diet through decreased expression of unspliced and spliced XBP-1, p-eIF2alpha, and CHOP. CONCLUSIONS: The iron-induced unfolded protein response appears to be one of the mechanisms responsible for fat accumulation in the liver in transgenic mice expressing the HCV polyprotein.

Control of lipid oxidation during exercise: role of energy state and mitochondrial factors.

Despite considerable progress during recent years our understanding of how lipid oxidation (LOx) is controlled during exercise remains incomplete. This review focuses on the role of mitochondria and energy state in the control of LOx. LOx increases in parallel with increased energy demand up to an exercise intensity of about 50-60% of VO(2max) after which the contribution of lipid decreases. The switch from lipid to carbohydrate (CHO) is of energetic advantage due to the increased ATP/O(2) yield. In the low-intensity domain (<50%VO(2max)) a moderate reduction in energy state will stimulate both LOx and CHO oxidation and relative fuel utilization is mainly controlled by substrate availability and the capacity of the metabolic pathways. In the high-intensity domain (>60%VO(2max)) there is a pronounced decrease in energy state, which will stimulate glycolysis in excess of the substrate requirements of the oxidative processes. This will lead to acidosis, reduced levels of free Coenzyme A (CoASH) and reduced levels of free carnitine. Acidosis and reduced carnitine may limit the carnitine-mediated transfer of long-chain fatty acids (LCFA) into mitochondria and may thus explain the observed reduction in LOx during high-intensity exercise. Another potential mechanism, suggested in this review, is that Acyl-CoA synthetase (ACS), an initial step in LCFA catabolism, functions as a regulator of LOx. ACS activity is suggested to be under control of CoASH and energy state. Furthermore, evidence exists that additional control points exist beyond mitochondrial FA influx. The nature and site of this control remain to be investigated.

Phosphorylation of rat liver mitochondrial carnitine palmitoyltransferase-I: effect on the kinetic properties of the enzyme.

Hepatic carnitine palmitoyltransferase-I (CPT-IL) isolated from mitochondrial outer membranes obtained in the presence of protein phosphatase inhibitors is readily recognized by phosphoamino acid antibodies. Mass spectrometric analysis of CPT-IL tryptic digests revealed the presence of three phosphopeptides including one with a protein kinase CKII (CKII) consensus site. Incubation of dephosphorylated outer membranes with protein kinases and [gamma-32P]ATP resulted in radiolabeling of CPT-I only by CKII. Using mass spectrometry, only one region of phosphorylation was detected in CPT-I isolated from CKII-treated mitochondria. The sequence of the peptide and position of phosphorylated amino acids have been determined unequivocally as FpSSPETDpSHRFGK (residues 740-752). Furthermore, incubation of dephosphorylated outer membranes with CKII and unlabeled ATP led to increased catalytic activity and rendered malonyl-CoA inhibition of CPT-I from competitive to uncompetitive. These observations identify a new mechanism for regulation of hepatic CPT-I by phosphorylation.

[Rhabdomyolysis in carnitine palmitoyltransferase II deficiency: developments in pathophysiology, diagnosis and therapy]. / Rhabdomyolyse bei Carnitin-Palmitoyltransferase-II-Mangel: aktuelle Entwicklungen in Pathophysiologie, Diagnostik und Therapie.

In hospitalized patients rhabdomyolysis is an important clinical entity, leading to myoglobinuria and acute renal failure in 8-25% of cases. When common causes of rhabdomyolysis, such as crush, trauma, infections, and drug abuse are excluded, inherited disorders of energy metabolism, in particular lipid metabolism, should be considered. Carnitine palmitoyltransferase (CPT) II deficiency is a common disorder of mitochondrial lipid oxidation. There are two distinct clinical forms: a severe and usually fatal infantile form and a benign classical muscular form. Usually, patients with CPT II deficiency present with episodic myoglobinuria, muscle cramps and weakness prompted by strenuous exercise or prolonged fasting. Liver and cardiac dysfunction are rarely seen and indicate severe disease. Most affected patients are males, although CPT II deficiency shows an autosomal recessive mode of inheritance. The human CPT II gene has been cloned, sequenced and localised to chromosome 1p32. Several mutations have been detected in the human gene which differ in the remaining enzyme activity and may explain the heterogeneity in the clinical picture of this disorder. Diagnosis is by muscle biopsy. Normally, light microscopy shows no pathological findings, and diagnosis must be established by biochemical and molecular methods. In our report on two typical cases we set out to promote knowledge of this disorder and discuss the diagnostic approach, which requires a specialised laboratory.

Selected metabolic alterations in the ischemic heart and their contributions to arrhythmogenesis.

Myocardial ischemia in vivo is associated with dramatic electrophysiologic alterations which occur within minutes of cessation of coronary flow and are rapidly reversible with reperfusion. This suggests that subtle and reversible biochemical and/or ionic alterations within or near the sarcolemma may contribute to the electrophysiologic derangements. Our studies have concentrated on 2 amphipathic metabolites, long-chain acylcarnitines and lysophosphatidylcholine (LPC) which have been shown to increase rapidly in ischemic tissue in vivo and to elicit electrophysiologic derangements in normoxic tissue in vitro. Incorporation of these amphiphiles into the sarcolemma at concentrations of 1 to 2 mol%, elicits profound electrophysiologic derangements analogous to those observed in ischemic myocardium in vivo. LPC is produced in endothelial cells and myocytes in response to thrombin. Thus, activation of the coagulation system during ischemia may result in extracellular production and accumulation of LPC. The pathophysiological effects of the accumulation of both amphiphiles are thought to be mediated by alterations in the biophysical properties of the sarcolemmal membrane, although there is a possibility of a direct effect on ion channels. Inhibition of carnitine acyltransferase I in the ischemic cat heart was found to prevent the increase in both long-chain acylcarnitines and LPC and to significantly reduce the incidence of malignant arrhythmias including ventricular tachycardia and fibrillation. This review focuses on the influence of these amphiphiles on cardiac ionic currents observed during early ischemia and presents data supporting the concept that accumulation of these amphiphiles within the sarcolemma contributes to changes in ionic conductances leading to electrophysiological derangements. The contribution and the accumulation of these amphiphiles to alterations in intracellular Ca2+ as related to changes in Na/K-ATPase activity and intracellular Na+ are examined. Other alterations occur during early myocardial ischemia in addition to the events reviewed here; however, the results of multiple studies over the past 2 decades indicate that accumulation of these amphiphiles contributes importantly to arrhythmogenesis and that development of specific inhibitors of carnitine acyltransferase I or phospholipase A2 may be a promising therapeutic strategy to attenuate the incidence of lethal arrhythmias associated with ischemic heart disease in man.

Body fat and the metabolic control of food intake.

The role of body fat in the control of food intake is considered from the point of view that the oxidation of metabolic fuels generates a signal that governs feeding behavior. According to this perspective, the storage and mobilization of fat affect food intake indirectly by altering fuel oxidation. Hyperphagia during the development of obesity is thus treated as an appropriate response to a primary metabolic defect that causes fuels to be stored rather than oxidized. Evidence is presented that changes in insulin level and the activity of carnitine palmitoyltransferase I modulate feeding by altering the partitioning of fatty acids. The possibility that dietary interactions, acting through these mechanisms, may cause overeating of high-fat diets is discussed. It is proposed that the signal for feeding originates in the liver when both fatty acids and glucose are unavailable for oxidation.