Correction to: Licochalcone A induces apoptosis through endoplasmic reticulum stress via a phospholipase Cγ1-, Ca2+-, and reactive oxygen species-dependent pathway in HepG2 human hepatocellular carcinoma cells.

The original version of this article contained mistakes in figures. The western blot data for pro-caspase-3 and cleaved caspase-3 (Fig. 1d), ß-actin (Fig. 1d), PLCγ1 (Fig. 5d), and eIF2α (Fig. 7d) are incorrect. The corrected Figs. 1d, 5d, and 7d are shown below. The corrections do not influence either the validity of the published data or the conclusion described in the article.

Licochalcone A induces apoptosis through endoplasmic reticulum stress via a phospholipase Cγ1-, Ca(2+)-, and reactive oxygen species-dependent pathway in HepG2 human hepatocellular carcinoma cells.

Licochalcone A (LicA), an estrogenic flavonoid, induces apoptosis in multiple types of cancer cells. In this study, the molecular mechanisms underlying the anti-cancer effects of LicA were investigated in HepG2 human hepatocellular carcinoma cells. LicA induced apoptotic cell death, activation of caspase-4, -9, and -3, and expression of endoplasmic reticulum (ER) stress-associated proteins, including C/EBP homologous protein (CHOP). Inhibition of ER stress by CHOP knockdown or treatment with the ER stress inhibitors, salubrinal and 4-phenylbutyric acid, reduced LicA-induced cell death. LicA also induced reactive oxygen species (ROS) accumulation and the anti-oxidant N-acetylcysteine reduced LicA-induced cell death and CHOP expression. In addition, LicA increased the levels of cytosolic Ca(2+), which was blocked by 2-aminoethoxydiphenyl borate (an antagonist of inositol 1,4,5-trisphosphate receptor) and BAPTA-AM (an intracellular Ca(2+) chelator). 2-Aminoethoxydiphenyl borate and BAPTA-AM inhibited LicA-induced cell death. Interestingly, LicA induced phosphorylation of phospholipase Cγ1 (PLCγ1) and inhibition of PLCγ1 reduced cell death and ER stress. Moreover, the multi-targeted receptor tyrosine kinase inhibitors, sorafenib and sunitinib, reduced LicA-induced cell death, ER stress, and cytosolic Ca(2+) and ROS accumulation. Finally, LicA induced phosphorylation of vascular endothelial growth factor receptor 2 (VEGFR2) and c-Met receptor and inhibition of both receptors by co-transfection with VEGFR2 and c-Met siRNAs reversed LicA-induced cell death, Ca(2+) increase, and CHOP expression. Taken together, these findings suggest that induction of ER stress via a PLCγ1-, Ca(2+)-, and ROS-dependent pathway may be an important mechanism by which LicA induces apoptosis in HepG2 hepatocellular carcinoma cells.

eNOS plays a major role in adiponectin synthesis in adipocytes.

Nitric oxide (NO) stimulates mitochondrial biogenesis. We recently reported that adiponectin synthesis is regulated by mitochondrial function in adipocytes. This study was undertaken to test the hypothesis that endothelial NO synthase (eNOS) plays an important role in adiponectin synthesis by producing NO and enhancing mitochondrial function in adipocytes. We examined the effects of eNOS knockdown on adiponectin synthesis in 3T3-L1 adipocytes and also examined plasma adiponectin levels and the mitochondria in adipose tissue of eNOS knockout (eNOS(-/-)) mice with and without chronic administration of a NO donor. In cultured 3T3-L1 adipocytes, eNOS siRNA decreased rosiglitazone-induced adiponectin secretion, which was associated with decreases in mitochondrial proteins and biogenesis factors. Plasma adiponectin concentrations were reduced in adult eNOS(-/-) mice compared with age-matched wild-type mice. Mitochondrial contents in adipose tissue were reduced in eNOS(-/-) mice, and this was associated with decreased expression of mitochondrial biogenesis factors, increased levels of 8-hydroxyguanosine, a biomarker of oxidative stress, and morphological abnormalities in mitochondria. Rosiglitazone-induced increases in adiponectin expression and mitochondrial content were also reduced significantly in eNOS(-/-) mice. Chronic administration of a NO donor reversed mitochondrial abnormalities and increased adiponectin expression in adipose tissue of eNOS(-/-) mice. eNOS plays an important role in adiponectin synthesis in adipocytes by increasing mitochondrial biogenesis and enhancing mitochondrial function.

Peroxisome proliferator-activated receptor-gamma coactivator 1-alpha overexpression prevents endothelial apoptosis by increasing ATP/ADP translocase activity.

OBJECTIVE: Fatty acids increase reactive oxygen species generation and cell apoptosis in endothelial cells. The peroxisome proliferator-activated receptor-gamma coactivator 1-alpha (PGC-1alpha) is a transcriptional coactivator that increases mitochondrial biogenesis and fatty acid oxidation in various cells. This study was undertaken to investigate the possible preventive effect of PGC-1alpha on endothelial apoptosis and its molecular mechanism. METHODS AND RESULTS: Treatment with linoleic acid in cultured human aortic endothelial cells increased reactive oxygen species generation and cell apoptosis. These effects appeared to be mediated by increases in cytosolic fat metabolites, ie, fatty acyl CoA, diacylglycerol, and ceramide, and consequent decreases in ATP/ADP translocase activity of adenine nucleotide translocator. Adenoviral overexpression of PGC-1alpha prevented linoleic acid-induced increases in reactive oxygen species generation and cell apoptosis in human aortic endothelial cells by increasing fatty acid oxidation, decreasing diacylglycerol and ceramide, and increasing ATP/ADP translocase activity. In isolated aorta, PGC-1alpha overexpression prevented linoleic acid-induced decrease in endothelium-dependent vasorelaxation, and this effect was abolished by adenine nucleotide translocator1 shRNA. CONCLUSIONS: PGC-1alpha regulates reactive oxygen species generation and apoptosis in endothelial cells by increasing fatty acid oxidation and enhancing ATP/ADP translocase activity. Measures to increase PGC-1alpha expression or ATP/ADP translocase activity in vascular cells may aid in the prevention or treatment of atherosclerosis.

Narrative review: evolving concepts in potassium homeostasis and hypokalemia.

Humans are intermittently exposed to large variations in potassium intake, which range from periods of fasting to ingestion of potassium-rich meals. These fluctuations would abruptly alter plasma potassium concentration if not for rapid mechanisms, primarily in skeletal muscle and the liver, that buffer the changes in plasma potassium concentration by means of transcellular potassium redistribution and feedback control of renal potassium excretion. However, buffers have capacity limits, and even robust feedback control mechanisms require that the perturbation occur before feedback can initiate corrective action. In contrast, feedforward control mechanisms sense the effect of disturbances on the system's homeostasis. This review highlights recent experimental insights into the participation of feedback and feedforward control mechanisms in potassium homeostasis. New data make clear that feedforward homeostatic responses activate when decreased potassium intake is sensed, even when plasma potassium concentration is still within the normal range and before frank hypokalemia ensues, in addition to the classic feedback activation of renal potassium conservation when plasma potassium concentration decreases. Given the clinical importance of dyskalemias in patients, these novel experimental paradigms invite renewed clinical inquiry into this important area.

Essential role of mitochondrial function in adiponectin synthesis in adipocytes.

OBJECTIVE: Adiponectin is an important adipocytokine that improves insulin action and reduces atherosclerotic processes. The plasma adiponectin level is paradoxically reduced in obese individuals, but the underlying mechanism is unknown. This study was undertaken to test the hypothesis that mitochondrial function is linked to adiponectin synthesis in adipocytes. RESEARCH DESIGN AND METHODS: We examined the effects of rosiglitazone and the measures that increase or decrease mitochondrial function on adiponectin synthesis. We also examined the molecular mechanism by which changes in mitochondrial function affect adiponectin synthesis. RESULTS: Adiponectin expression and mitochondrial content in adipose tissue were reduced in obese db/db mice, and these changes were reversed by the administration of rosiglitazone. In cultured adipocytes, induction of increased mitochondrial biogenesis (via adenoviral overexpression of nuclear respiratory factor-1) increased adiponectin synthesis, whereas impairment in mitochondrial function decreased it. Impaired mitochondrial function increased endoplasmic reticulum (ER) stress, and agents causing mitochondrial or ER stress reduced adiponectin transcription via activation of c-Jun NH(2)-terminal kinase (JNK) and consequent induction of activating transcription factor (ATF)3. Increased mitochondrial biogenesis reversed all of these changes. CONCLUSIONS: Mitochondrial function is linked to adiponectin synthesis in adipocytes, and mitochondrial dysfunction in adipose tissue may explain decreased plasma adiponectin levels in obesity. Impaired mitochondrial function activates a series of mechanisms involving ER stress, JNK, and ATF3 to decrease adiponectin synthesis.

Alpha-lipoic acid prevents endothelial dysfunction in obese rats via activation of AMP-activated protein kinase.

OBJECTIVE: Lipid accumulation in vascular endothelial cells may play an important role in the pathogenesis of atherosclerosis in obese subjects. We showed previously that alpha-lipoic acid (ALA) activates AMP-activated protein kinase (AMPK) and reduces lipid accumulation in skeletal muscle of obese rats. Here, we investigated whether ALA improves endothelial dysfunction in obese rats by activating AMPK in endothelial cells. METHODS AND RESULTS: Endothelium-dependent vascular relaxation was impaired, and the number of apoptotic endothelial cells was higher in the aorta of obese rats compared with control rats. In addition, triglyceride and lipid peroxide levels were higher, and NO synthesis was lower. Administration of ALA improved all of these abnormalities. AMPK activity was lower in aortic endothelium of obese rats, and ALA normalized it. Incubation of human aortic endothelial cells with ALA activated AMPK and protected cells from linoleic acid-induced apoptosis. Dominant-negative AMPK inhibited the antiapoptotic effects of ALA. CONCLUSIONS: Reduced AMPK activation may play an important role in the genesis of endothelial dysfunction in obese rats. ALA improves vascular dysfunction by normalizing lipid metabolism and activating AMPK in endothelial cells.

alpha-Lipoic acid prevents diabetes mellitus in diabetes-prone obese rats.

Several lines of evidence have suggested that triglyceride accumulation in skeletal muscle and pancreatic islets is causally related to type 2 diabetes mellitus. We recently showed that alpha-lipoic acid (ALA), a potent antioxidant and cofactor of mitochondrial respiratory enzymes, reduces body weight of rodents by suppressing food intake and increasing energy expenditure. We sought to determine if ALA can prevent the development of diabetes mellitus in obese Otsuka Long-Evans Tokushima Fatty (OLETF) rats. Most (78%) untreated OLETF rats showed glycosuria at 40 weeks of age, but this was completely prevented by ALA. Compared with untreated OLETF rats, ALA reduced body weight and protected pancreatic beta-cells from destruction. ALA also reduced triglyceride accumulation in skeletal muscle and pancreatic islets. These results indicate that ALA prevents diabetes mellitus in obese diabetes-prone rats by reducing lipid accumulation in non-adipose tissue as well as in adipose tissue.

Anti-obesity effects of alpha-lipoic acid mediated by suppression of hypothalamic AMP-activated protein kinase.

AMP-activated protein kinase (AMPK) functions as a fuel sensor in the cell and is activated when cellular energy is depleted. Here we report that alpha-lipoic acid (alpha-LA), a cofactor of mitochondrial enzymes, decreases hypothalamic AMPK activity and causes profound weight loss in rodents by reducing food intake and enhancing energy expenditure. Activation of hypothalamic AMPK reverses the effects of alpha-LA on food intake and energy expenditure. Intracerebroventricular (i.c.v.) administration of glucose decreases hypothalamic AMPK activity, whereas inhibition of intracellular glucose utilization through the administration of 2-deoxyglucose increases hypothalamic AMPK activity and food intake. The 2-deoxyglucose-induced hyperphagia is reversed by inhibiting hypothalamic AMPK. Our findings indicate that hypothalamic AMPK is important in the central regulation of food intake and energy expenditure and that alpha-LA exerts anti-obesity effects by suppressing hypothalamic AMPK activity.

Peroxisome proliferator-activated receptor (PPAR)-alpha activation prevents diabetes in OLETF rats: comparison with PPAR-gamma activation.

Lipid accumulation in nonadipose tissues is closely related to the development of type 2 diabetes in obese subjects. We examined the potential preventive effect of peroxisome proliferator-activated receptor (PPAR)-alpha and PPAR-gamma stimulation on the development of diabetes in obese diabetes-prone OLETF rats. Chronic administration of a PPAR-alpha agonist (0.5% [wt/wt] fenofibrate) or a PPAR-gamma agonist (3 mg x kg(-1) x day(-1) rosiglitazone) completely prevented the development of glycosuria. Pancreatic islets from untreated OLETF rats underwent sequential hypertrophy and atrophy, which was completely prevented by chronic fenofibrate treatment. In contrast, rosiglitazone treatment did not affect islet hypertrophy at earlier stages but prevented beta-cell atrophy at later stages. Fenofibrate treatment decreased body weight and visceral fat, whereas rosiglitazone treatment increased body weight. Despite the opposite effects on adiposity, both drugs were equally effective in improving insulin actions in skeletal muscle. Furthermore, both drugs significantly decreased the triglyceride content in the soleus muscle and pancreatic islets. The present study demonstrates that the PPAR-alpha agonist fenofibrate prevents the development of diabetes in OLETF rats by reducing adiposity, improving peripheral insulin action, and exerting beneficial effects on pancreatic beta-cells.