Cuban Policosanol Prevents the Apoptosis and the Mitochondrial Dysfunction Induced by Lipopolysaccharide in C2C12 Myoblast via Activation of Akt and Erk Pathways.

Skeletal muscle plays crucial roles in locomotion, protein reservoir, and maintenance of metabolic homeostasis. Loss of muscle, known as muscle atrophy, causes the metabolic diseases such as type 2 diabetes mellitus, hypertension, and so on. Therefore, great efforts have been devoted to prevent the muscle atrophy. Policosanols are a mixture of long chain fatty alcohols extracted from various natural sources. They have long been used as functional foods to lower the level of serum lipids, including triacylglycerol and cholesterol, and to protect against inflammatory stress. In this study, we examine the protective effect and molecular mechanism of Cuban policosanol on skeletal muscle cell death and mitochondrial dysfunction using lipopolysaccharide-treated C2C12 cells. Our results demonstrated that policosanol significantly rescued cell survival (40% vs. 88%; LPS vs. LPS+policosanol) via activation of the Akt pathway, resulting in inhibition of apoptosis (p<0.05). Moreover, policosanol restored the LPS-induced repression of collagen by two fold (0.33±0.04 vs. 0.67±0.03 compared to that of control; LPS vs. LPS+policosanol) via activation of ERK-mTOR-p70S6K pathways. In addition, policosanol increased the mitochondrial fusion by regulating the activities of DRP1 and Mfn2, leading to ameliorate the mitochondrial dysfunction induced by LPS. Improved mitochondria function increased the oxygen consumption rate with glucose as fuel source, indicating that policosanol could shift the glucose metabolism from lactate fermentation, induced by lipopolysaccharide, to oxidative phosphorylation. Thus, policosanol is a promising agent for preventing the inflammation-induced muscle cell death and mitochondrial dysfunction.

Cryptotanshinone Prevents the Binding of S6K1 to mTOR/Raptor Leading to the Suppression of mTORC1-S6K1 Signaling Activity and Neoplastic Cell Transformation.

Cryptotanshinone is known for its inhibitory activity against tumorigenesis in various human cancer cells. However, exact mechanisms underlying the anticancer effects of cryptotanshinone are not fully elucidated. Here, we propose a plausible molecular mechanism, wherein cryptotanshinone represses rapamycin-sensitive mTORC1/S6K1 mediated cancer cell growth and cell transformation. We investigated the various effects of cryptotanshinone on the mTORC1/S6K1 axis, which is associated with the regulation of cell growth in response to nutritional and growth factor signals. We found that cryptotanshinone specifically inhibited the mTORC1-mediated phosphorylation of S6K1, which consequently suppressed the clonogenicity of SK-Hep1 cells and the neoplastic transformation of JB6 Cl41 cells induced by insulin-like growth factor-1. Finally, we observed that cryptotanshinone prevented S6K1 from binding to the Raptor/mTOR complex, rather than regulating mTOR and its upstream pathway. Overall, our findings provide a novel mechanism underlying anti-cancer effects cryptotanshinone targeting mTORC1 signaling, contributing to the development of anticancer agents involving metabolic cancer treatment.

The effect of autocrine motility factor alone and in combination with methyl jasmonate on liver cancer cell growth.

Neoplastic cells secrete autocrine motility factor (AMF) to stimulate the motility of cancer cells. In this study, AMF secreted from HT-29 colorectal cancer cells selectively suppressed liver cancer cells by downregulating pAKT and ß-catenin. In addition, HT-29 AMF significantly augmented the activity of methyl jasmonate against liver cancer cells and is a promising alternative for liver cancer therapy.

Synergistic effects of autocrine motility factor and methyl jasmonate on human breast cancer cells.

Autocrine motility factor (AMF) stimulates the motility of cancer cells via an autocrine route and has been implicated in tumor progression and metastasis. Overexpression of AMF is correlated with the aggressive nature of breast cancer and is negatively associated with clinical outcomes. In contrast, AMF also has the ability to suppress cancer cells. In this study, AMFs from different cancer cells were demonstrated to have suppressive activity against MCF-7 and MDA-MB-231 breast cancer cells. In a growth and colony formation assay, AMF from AsPC-1 pancreatic cancer cells (ASPC-1:AMF) was determined to be more suppressive compared to other AMFs. It was also demonstrated that AsPC-1:AMF could arrest breast cancer cells at the G0/G1 cell cycle phase. Quantified by Western blot analysis, AsPC-1:AMF lowered levels of the AMF receptor (AMFR) and G-protein-coupled estrogen receptor (GPER), concomitantly regulating the activation of the AKT and ERK signaling pathways. JAK/STAT activation was also decreased. These results were found in estrogen receptor (ER)-positive MCF-7 cells but not in triple-negative MDA-MB-231 cells, suggesting that AsPC-1:AMF could work through multiple pathways led to apoptosis. More importantly, AsPC-1:AMF and methyl jasmonate (MJ) cooperatively and synergistically acted against breast cancer cells. Thus, AMF alone or along with MJ may be a promising breast cancer treatment option.

Autocrine motility factor secreted by HeLa cells inhibits the growth of many cancer cells by regulating AKT/ERK signaling.

In cell competition, a secreted death signal can determine cell fate. However, the nature of such a signal remains unclear. In this study, conditioned medium from HeLa cells (HeLa CM) inhibited growth of A549 and MCF-7 cells. Through HeLa CM fractionation, glucose 6-phosphate isomerase/autocrine motility factor (GPI/AMF) was identified as the main growth inhibitor. Previously, AMF was known for its mitogenic, motogenic, and differentiation functions and was implicated in tumor progression and metastasis. HeLa CM lost its growth inhibitory property after treatment with erythrose-4-phosphate (E4P) or anti-GPI antibody. Purified HeLa recombinant AMF (rAMF) proteins inhibited the growth of A549, MDA-MB-232, MCF-7, AsPC-1, DU145, Hep-2, Hep G2, and HT-29 cells. However, growth of HL-60, SKOV3, U-87 MG, SNU-484, U-87 MG, and 3T3-L1 cells was little affected. In a Transwell assay, HeLa rAMF effectively reduced A549 cell migration and invasion. HeLa rAMF effectively induced apoptosis in A549 cells, apparently by reducing the levels of Bcl-2, GPI, and poly(ADP-ribose) polymerase (PARP)14 and activating caspase-3 and p53. HeLa rAMF antagonized HER2 and the AMF receptor (AMFR or GP78) in relation to the AKT/EKT signaling pathway. These results suggest that HeLa AMF could act as a diffusible death signal that could induce cancer cell-selective growth inhibition and apoptosis.

PDK4 Deficiency Suppresses Hepatic Glucagon Signaling by Decreasing cAMP Levels.

In fasting or diabetes, gluconeogenic genes are transcriptionally activated by glucagon stimulation of the cAMP-protein kinase A (PKA)-CREB signaling pathway. Previous work showed pyruvate dehydrogenase kinase (PDK) inhibition in skeletal muscle increases pyruvate oxidation, which limits the availability of gluconeogenic substrates in the liver. However, this study found upregulation of hepatic PDK4 promoted glucagon-mediated expression of gluconeogenic genes, whereas knockdown or inhibition of hepatic PDK4 caused the opposite effect on gluconeogenic gene expression and decreased hepatic glucose production. Mechanistically, PDK4 deficiency decreased ATP levels, thus increasing phosphorylated AMPK (p-AMPK), which increased p-AMPK-sensitive phosphorylation of cyclic nucleotide phosphodiesterase 4B (p-PDE4B). This reduced cAMP levels and consequently p-CREB. Metabolic flux analysis showed that the reduction in ATP was a consequence of a diminished rate of fatty acid oxidation (FAO). However, overexpression of PDK4 increased FAO and increased ATP levels, which decreased p-AMPK and p-PDE4B and allowed greater accumulation of cAMP and p-CREB. The latter were abrogated by the FAO inhibitor etomoxir, suggesting a critical role for PDK4 in FAO stimulation and the regulation of cAMP levels. This finding strengthens the possibility of PDK4 as a target against diabetes.

Pyruvate dehydrogenase kinase 4 deficiency attenuates cisplatin-induced acute kidney injury.

Clinical prescription of cisplatin, one of the most widely used chemotherapeutic agents, is limited by its side effects, particularly tubular injury-associated nephrotoxicity. Since details of the underlying mechanisms are not fully understood, we investigated the role of pyruvate dehydrogenase kinase (PDK) in cisplatin-induced acute kidney injury. Among the PDK isoforms, PDK4 mRNA and protein levels were markedly increased in the kidneys of mice treated with cisplatin, and c-Jun N-terminal kinase activation was involved in cisplatin-induced renal PDK4 expression. Treatment with the PDK inhibitor sodium dichloroacetate (DCA) or genetic knockout of PDK4 attenuated the signs of cisplatin-induced acute kidney injury, including apoptotic morphology of the kidney tubules along with numbers of TUNEL-positive cells, cleaved caspase-3, and renal tubular injury markers. Cisplatin-induced suppression of the mitochondrial membrane potential, oxygen consumption rate, expression of electron transport chain components, cytochrome c oxidase activity, and disruption of mitochondrial morphology were noticeably improved in the kidneys of DCA-treated or PDK4 knockout mice. Additionally, levels of the oxidative stress marker 4-hydroxynonenal and mitochondrial reactive oxygen species were attenuated, whereas superoxide dismutase 2 and catalase expression and glutathione synthetase and glutathione levels were recovered in DCA-treated or PDK4 knockout mice. Interestingly, lipid accumulation was considerably attenuated in DCA-treated or PDK4 knockout mice via recovered expression of peroxisome proliferator-activated receptor-α and coactivator PGC-1α, which was accompanied by recovery of mitochondrial biogenesis. Thus, PDK4 mediates cisplatin-induced acute kidney injury, suggesting that PDK4 might be a therapeutic target for attenuating cisplatin-induced acute kidney injury.

Inhibition of Pyruvate Dehydrogenase Kinase 2 Protects Against Hepatic Steatosis Through Modulation of Tricarboxylic Acid Cycle Anaplerosis and Ketogenesis.

Hepatic steatosis is associated with increased insulin resistance and tricarboxylic acid (TCA) cycle flux, but decreased ketogenesis and pyruvate dehydrogenase complex (PDC) flux. This study examined whether hepatic PDC activation by inhibition of pyruvate dehydrogenase kinase 2 (PDK2) ameliorates these metabolic abnormalities. Wild-type mice fed a high-fat diet exhibited hepatic steatosis, insulin resistance, and increased levels of pyruvate, TCA cycle intermediates, and malonyl-CoA but reduced ketogenesis and PDC activity due to PDK2 induction. Hepatic PDC activation by PDK2 inhibition attenuated hepatic steatosis, improved hepatic insulin sensitivity, reduced hepatic glucose production, increased capacity for ß-oxidation and ketogenesis, and decreased the capacity for lipogenesis. These results were attributed to altered enzymatic capacities and a reduction in TCA anaplerosis that limited the availability of oxaloacetate for the TCA cycle, which promoted ketogenesis. The current study reports that increasing hepatic PDC activity by inhibition of PDK2 ameliorates hepatic steatosis and insulin sensitivity by regulating TCA cycle anaplerosis and ketogenesis. The findings suggest PDK2 is a potential therapeutic target for nonalcoholic fatty liver disease.

Inflammation increases pyruvate dehydrogenase kinase 4 (PDK4) expression via the Jun N-Terminal Kinase (JNK) pathway in C2C12 cells.

Chronic inflammation augments the deleterious effects of several diseases, particularly diabetes, cancer, and sepsis. It is also involved in the process of metabolic shift from glucose oxidation to lactate production. Although several studies suggest that the change in activity of the pyruvate dehydrogenase complex (PDC) is a major factor causing this metabolic change, the exact mechanism of the inflammatory state remains unclear. In this study, we investigated the effect of lipopolysaccharide (LPS) on the expression of pyruvate dehydrogenase kinase 4 (PDK4), which is strongly associated with inactivation of the PDC in C2C12 myoblasts. In C2C12 myoblasts, LPS exposure led to increased PDK4 mRNA and protein expression levels as well as lactate production in culture medium. However, the expression levels of other PDK isoenzymes (PDK1 - 3) remained unchanged. Additionally, we observed that LPS treatment induced phosphorylation of Jun N-Terminal Kinases (JNK). To confirm the role of JNK, we inhibited the JNK pathway and observed that PDK4 expression and lactate production were decreased, but p38 and ERK were not significantly changed. Taken together, our results suggest that LPS induces PDK4 expression and alters glucose metabolism via the JNK pathway.

Effect of silymarin on gluconeogenesis and lactate production in exercising rats.

In this study, we investigated the effects of silymarin (SM) on gluconeogenesis during exercise in rats. After 4 weeks of exercise, blood samples, liver, and skeletal muscle tissues were collected, and the levels of triglycerides (TG), lactate, peroxisome proliferator activated receptor gamma (PPARγ), phosphoenol pyruvate carboxykinase (PEPCK), pyruvate dehydrogenase kinase 4 (PDK4), and phosphorylated 5-AMP activated protein kinase (AMPK) were measured. The TG and lactate level of the serum were reduced. In addition, the expression of Akt, PEPCK, and PPARγ in liver was decreased as well as the expression of AMPK in muscle. On the contrary, the level of PDK4 in muscle was increased. These results showed that that administration of SM ameliorated exerciseinduced gluconeogenesis and ß-oxidation through the regulation of PPARγ, PEPCK, and PDK4. Thus, intake of SM during exercise may improve endurance by modulating of the metabolism of glucose, lipids, and lactate.

Metabolic Connection of Inflammatory Pain: Pivotal Role of a Pyruvate Dehydrogenase Kinase-Pyruvate Dehydrogenase-Lactic Acid Axis.

Pyruvate dehydrogenase kinases (PDK1-4) are mitochondrial metabolic regulators that serve as decision makers via modulation of pyruvate dehydrogenase (PDH) activity to convert pyruvate either aerobically to acetyl-CoA or anaerobically to lactate. Metabolic dysregulation and inflammatory processes are two sides of the same coin in several pathophysiological conditions. The lactic acid surge associated with the metabolic shift has been implicated in diverse painful states. In this study, we investigated the role of PDK-PDH-lactic acid axis in the pathogenesis of chronic inflammatory pain. Deficiency of Pdk2 and/or Pdk4 in mice attenuated complete Freund's adjuvant (CFA)-induced pain hypersensitivities. Likewise, Pdk2/4 deficiency attenuated the localized lactic acid surge along with hallmarks of peripheral and central inflammation following intraplantar administration of CFA. In vitro studies supported the role of PDK2/4 as promoters of classical proinflammatory activation of macrophages. Moreover, the pharmacological inhibition of PDKs or lactic acid production diminished CFA-induced inflammation and pain hypersensitivities. Thus, a PDK-PDH-lactic acid axis seems to mediate inflammation-driven chronic pain, establishing a connection between metabolism and inflammatory pain. SIGNIFICANCE STATEMENT: The mitochondrial pyruvate dehydrogenase (PDH) kinases (PDKs) and their substrate PDH orchestrate the conversion of pyruvate either aerobically to acetyl-CoA or anaerobically to lactate. Lactate, the predominant end product of glycolysis, has recently been identified as a signaling molecule for neuron-glia interactions and neuronal plasticity. Pathological metabolic shift and subsequent lactic acid production are thought to play an important role in diverse painful states; however, their contribution to inflammation-driven pain is still to be comprehended. Here, we report that the PDK-PDH-lactic acid axis constitutes a key component of inflammatory pain pathogenesis. Our findings establish an unanticipated link between metabolism and inflammatory pain. This study unlocks a previously ill-explored research avenue for the metabolic control of inflammatory pain pathogenesis.

Pyruvate Dehydrogenase Kinases: Therapeutic Targets for Diabetes and Cancers.

Impaired glucose homeostasis is one of the risk factors for causing metabolic diseases including obesity, type 2 diabetes, and cancers. In glucose metabolism, pyruvate dehydrogenase complex (PDC) mediates a major regulatory step, an irreversible reaction of oxidative decarboxylation of pyruvate to acetyl-CoA. Tight control of PDC is critical because it plays a key role in glucose disposal. PDC activity is tightly regulated using phosphorylation by pyruvate dehydrogenase kinases (PDK1 to 4) and pyruvate dehydrogenase phosphatases (PDP1 and 2). PDKs and PDPs exhibit unique tissue expression patterns, kinetic properties, and sensitivities to regulatory molecules. During the last decades, the up-regulation of PDKs has been observed in the tissues of patients and mammals with metabolic diseases, which suggests that the inhibition of these kinases may have beneficial effects for treating metabolic diseases. This review summarizes the recent advances in the role of specific PDK isoenzymes on the induction of metabolic diseases and describes the effects of PDK inhibition on the prevention of metabolic diseases using pharmacological inhibitors. Based on these reports, PDK isoenzymes are strong therapeutic targets for preventing and treating metabolic diseases.

Physiological effect and therapeutic application of alpha lipoic acid.

Reactive oxygen species and reactive nitrogen species promote endothelial dysfunction in old age and contribute to the development of cardiovascular diseases such as atherosclerosis, diabetes, and hypertension. α-Lipoic acid was identified as a catalytic agent for oxidative decarboxylation of pyruvate and α-ketoglutarate in 1951, and it has been studied intensively by chemists, biologists, and clinicians who have been interested in its role in energetic metabolism and protection from reactive oxygen species-induced mitochondrial dysfunction. Consequently, many biological effects of α-lipoic acid supplementation can be attributed to the potent antioxidant properties of α-lipoic acid and dihydro α-lipoic acid. The reducing environments inside the cell help to protect from oxidative damage and the reduction-oxidation status of α-lipoic acid is dependent upon the degree to which the cellular components are found in the oxidized state. Although healthy young humans can synthesize enough α-lipoic acid to scavenge reactive oxygen species and enhance endogenous antioxidants like glutathione and vitamins C and E, the level of α-lipoic acid significantly declines with age and this may lead to endothelial dysfunction. Furthermore, many studies have reported α-lipoic acid can regulate the transcription of genes associated with anti-oxidant and anti-inflammatory pathways. In this review, we will discuss recent clinical studies that have investigated the beneficial effects of α-lipoic acid on endothelial dysfunction and propose possible mechanisms involved.

Dimethylfumarate attenuates restenosis after acute vascular injury by cell-specific and Nrf2-dependent mechanisms.

Excessive proliferation of vascular smooth muscle cells (VSMCs) and incomplete re-endothelialization is a major clinical problem limiting the long-term efficacy of percutaneous coronary angioplasty. We tested if dimethylfumarate (DMF), an anti-psoriasis drug, could inhibit abnormal vascular remodeling via NF-E2-related factor 2 (Nrf2)-NAD(P)H quinone oxidoreductase 1 (NQO1) activity. DMF significantly attenuated neointimal hyperplasia induced by balloon injury in rat carotid arteries via suppression of the G1 to S phase transition resulting from induction of p21 protein in VSMCs. Initially, DMF increased p21 protein stability through an enhancement in Nrf2 activity without an increase in p21 mRNA. Later on, DMF stimulated p21 mRNA expression through a process dependent on p53 activity. However, heme oxygenase-1 (HO-1) or NQO1 activity, well-known target genes induced by Nrf2, were dispensable for the DMF induction of p21 protein and the effect on the VSMC proliferation. Likewise, DMF protected endothelial cells from TNF-α-induced apoptosis and the dysfunction characterized by decreased eNOS expression. With knock-down of Nrf2 or NQO1, DMF failed to prevent TNF-α-induced cell apoptosis and decreased eNOS expression. Also, CD31 expression, an endothelial specific marker, was restored in vivo by DMF. In conclusion, DMF prevented abnormal proliferation in VSMCs by G1 cell cycle arrest via p21 upregulation driven by Nrf2 and p53 activity, and had a beneficial effect on TNF-α-induced apoptosis and dysfunction in endothelial cells through Nrf2-NQO1 activity suggesting that DMF might be a therapeutic drug for patients with vascular disease.

Regulation of pyruvate metabolism in metabolic-related diseases.

Pyruvate is an obligatory intermediate in the oxidative disposal of glucose and a major precursor for the synthesis of glucose, glycerol, fatty acids, and non-essential amino acids. Stringent control of the fate of pyruvate is critically important for cellular homeostasis. The regulatory mechanisms for its metabolism are therefore of great interest. Recent advances include the findings that (a) the mitochondrial pyruvate carrier is sensitive to inhibition by thiazolidinediones; (b) pyruvate dehydrogenase kinases induce the Warburg effect in many disease states; and (c) pyruvate carboxylase is an important determinate of the rates of gluconeogenesis in humans with type 2 diabetes. These enzymes are potential therapeutic targets for several diseases.

Pyruvate dehydrogenase kinase-4 contributes to the recirculation of gluconeogenic precursors during postexercise glycogen recovery.

During recovery from glycogen-depleting exercise, there is a shift from carbohydrate oxidation to glycogen resynthesis. The activity of the pyruvate dehydrogenase (PDH) complex may decrease to reduce oxidation of carbohydrates in favor of increasing gluconeogenic recycling of carbohydrate-derived substrates for this process. The precise mechanism behind this has yet to be elucidated; however, research examining mRNA content has suggested that the less-abundant pyruvate dehydrogenase kinase-4 (PDK4) may reduce PDH activation during exercise recovery. To investigate this, skeletal muscle and liver of wild-type (WT) and PDK4-knockout (PDK4-KO) mice were analyzed at rest (Rest), after exercise to exhaustion (Exh), and after 2 h of recovery with ad libitum feeding (Rec). Although there were no differences in exercise tolerance between genotypes, caloric consumption was doubled by PDK4-KO mice during Rec. Because of this, PDK4-KO mice at Rec supercompensated muscle glycogen to 120% of resting stores. Therefore, an extra group of PDK4-KO mice were pair-fed (PF) with WT mice during Rec for comparison. PF mice fully replenished muscle glycogen but recovered only 50% of liver glycogen stores. Concentrations of muscle lactate and alanine were also lower in PF than in WT mice, indicating that this decrease may lead to a potential reduction of recycled gluconeogenic substrates, due to oxidation of their carbohydrate precursors in skeletal muscle, leading to observed reductions in hepatic glucose and glycogen concentrations. Because of the impairments seen in PF PDK4-KO mice, these results suggest a role for PDK4 in regulating the PDH complex in muscle and promoting gluconeogenic precursor recirculation during recovery from exhaustive exercise.

Dimethylfumarate suppresses adipogenic differentiation in 3T3-L1 preadipocytes through inhibition of STAT3 activity.

The excessive accumulation of adipocytes contributes to the development of obesity and obesity-related diseases. The interactions of several transcription factors, such as C/EBPß, PPARγ, C/EBPα, Nrf2, and STAT3, are required for adipogenic differentiation. Dimethylfumarate (DMF), an immune modulator and antioxidant, may function as an inhibitor of STAT3 and an activator of Nrf2. This study examined whether DMF inhibits adipogenic differentiation of 3T3-L1 preadipocytes by inhibiting STAT3 or activating Nrf2. DMF suppressed 3T3-L1 preadipocyte differentiation to mature adipocytes in a dose-dependent manner as determined by Oil Red O staining. The mRNA and protein levels of adipogenic genes, including C/EBPß, C/EBPα, PPARγ, SREBP-1c, FAS, and aP2, were significantly lower in DMF-treated 3T3-L1 preadipocytes. Suppression of adipogenic differentiation by DMF treatment resulted primarily from inhibition of the early stages of differentiation. DMF inhibits clonal expansion during adipogenic differentiation through induction of a G1 cell cycle arrest. Additionally, DMF regulates cell cycle-related proteins, such as p21, pRb, and cyclin D. DMF treatment markedly inhibited differentiation medium-induced STAT3 phosphorylation and inhibited STAT3 transcriptional activation of a reporter construct composed of four synthetic STAT3-response elements. Moreover, inhibition of endogenous Nrf2 activity using a dominant negative Nrf2 did not abolish the DMF-induced inhibition of adipogenic differentiation of 3T3-L1 preadipocytes. In summary, DMF is a negative regulator of adipogenic differentiation based on its regulation of adipogenic transcription factors and cell cycle proteins. This negative regulation by DMF is mediated by STAT3 inhibition, but is unlikely to involve Nrf2 activation.

Transcriptional regulation of pyruvate dehydrogenase kinase.

The pyruvate dehydrogenase complex (PDC) activity is crucial to maintains blood glucose and ATP levels, which largely depends on the phosphorylation status by pyruvate dehydrogenase kinase (PDK) isoenzymes. Although it has been reported that PDC is phosphorylated and inactivated by PDK2 and PDK4 in metabolically active tissues including liver, skeletal muscle, heart, and kidney during starvation and diabetes, the precise mechanisms by which expression of PDK2 and PDK4 are transcriptionally regulated still remains unclear. Insulin represses the expression of PDK2 and PDK4 via phosphorylation of FOXO through PI3K/Akt signaling pathway. Several nuclear hormone receptors activated due to fasting or increased fat supply, including peroxisome proliferator-activated receptors, glucocorticoid receptors, estrogen-related receptors, and thyroid hormone receptors, also participate in the up-regulation of PDK2 and PDK4; however, the endogenous ligands that bind those nuclear receptors have not been identified. It has been recently suggested that growth hormone, adiponectin, epinephrine, and rosiglitazone also control the expression of PDK4 in tissue-specific manners. In this review, we discuss several factors involved in the expressional regulation of PDK2 and PDK4, and introduce current studies aimed at providing a better understanding of the molecular mechanisms that underlie the development of metabolic diseases such as diabetes.

Metformin inhibits growth hormone-mediated hepatic PDK4 gene expression through induction of orphan nuclear receptor small heterodimer partner.

Growth hormone (GH) is a counter-regulatory hormone that plays an important role in preventing hypoglycemia during fasting. Because inhibition of the pyruvate dehydrogenase complex (PDC) by pyruvate dehydrogenase kinase 4 (PDK4) conserves substrates for gluconeogenesis, we tested whether GH increases PDK4 expression in liver by a signaling pathway sensitive to inhibition by metformin. The effects of GH and metformin were determined in the liver of wild-type, small heterodimer partner (SHP)-, PDK4-, and signal transducer and activator of transcription 5 (STAT5)-null mice. Administration of GH in vivo increased PDK4 expression via a pathway dependent on STAT5 phosphorylation. Metformin inhibited the induction of PDK4 expression by GH via a pathway dependent on AMP-activated protein kinase (AMPK) and SHP induction. The increase in PDK4 expression and PDC phosphorylation by GH was reduced in STAT5-null mice. Metformin decreased GH-mediated induction of PDK4 expression and metabolites in wild-type but not in SHP-null mice. In primary hepatocytes, dominant-negative mutant-AMPK and SHP knockdown prevented the inhibitory effect of metformin on GH-stimulated PDK4 expression. SHP directly inhibited STAT5 association on the PDK4 gene promoter. Metformin inhibits GH-induced PDK4 expression and metabolites via an AMPK-SHP-dependent pathway. The metformin-AMPK-SHP network may provide a novel therapeutic approach for the treatment of hepatic metabolic disorders induced by the GH-mediated pathway.