Pinolenic acid Mediated Anti-inflammatory and Immunometabolic Effects in Peripheral blood-derived Monocytes from patients with Rheumatoid Arthritis is Associated with Modulation of miRNA expression

Objectives: Pinolenic acid (PNLA), an omega-6 polyunsaturated fatty acid from pine nuts, has anti-inflammatory and anti-atherogenic effects. We aimed to investigate the actions of PNLA on activated purified monocytes from peripheral blood of patients with rheumatoid arthritis (RA).Methods: Flow cytometry was used to assess the intracellular expression of TNF-α, IL-6, IL-1β, and IL-8 in purified monocytes from patients with RA after lipopolysaccharide (LPS) stimulation with/without PNLA pre-treatment. The whole genomic transcriptomic (WGT) profile of PNLA-treated, and LPS-activated monocytes from patients with active RA was investigated by RNA-sequencing.Results: PNLA reduced percentage of monocytes expressing the cytokines TNF-α by 23% (p=0.048), IL-6 by 25% (p=0.011), IL-1β by 23% (p=0.050) and IL-8 by 19% (p=0.066). Canonical pathway analysis showed that PNLA inhibited oxidative phosphorylation (p= 9.14E-09) and mitochondrial dysfunction (p=4.18E-08), while the sirtuin (SIRTs) signalling pathway was activated (p=8.89E-06). Pathway analysis predicted upstream activation of peroxisome proliferator-activated receptors (PPARs), sirtuin3, and let7miRNA, which are anti-inflammatory and antioxidative. In contrast, DAP3, LIF and STAT3, which are involved in TNF-α, and IL-6 signal transduction, were inhibited. Many miRNAs were modulated by PNLA suggesting potential post-transcriptional regulation of metabolic and immune response that has not been described previously. Multiple miRNAs target pyruvate dehydrogenase kinase-4 (PDK4), single-immunoglobulin interleukin-1 receptor-related molecule (SIGIRR), mitochondrially encoded ATP synthase membrane subunit 6 (MT-ATP6) and acetyl-CoA acyltransferase 2 (ACAA2); genes implicated in cell metabolism, inflammation, and mitochondrial dysfunction.Conclusion: PNLA has anti-inflammatory and immune-metabolic effects on monocytes that are pathogenic in RA and atherosclerosis. Dietary PNLA supplementation may regulate key miRNAs that are involved in mitochondrial, metabolic, and inflammatory pathways.

Loss of PDK1 Induces Meiotic Defects in Oocytes From Diabetic Mice

Maternal diabetes has been shown to impair oocyte quality; however, the underlying mechanisms remain unclear. Here, using a streptozotocin (STZ)-induced diabetic mouse model, we first detected and reduced expression of pyruvate dehydrogenase kinase 1 (PDK1) in diabetic oocytes, accompanying with the lowered phosphorylation of serine residue 232 on α subunit of the pyruvate dehydrogenase (PDH) complex (Ser232-PDHE1α). Importantly, forced expression of PDK1 not only elevated the phosphorylation level of Ser232-PDHE1α, but also partly prevented the spindle disorganization and chromosome misalignment in oocytes from diabetic mice, with no beneficial effects on metabolic dysfunction. Moreover, a phospho-mimetic S232D-PDHE1α mutant is also capable of ameliorating the maternal diabetes-associated meiotic defects. In sum, our data indicate that PDK1-controlled Ser232-PDHE1α phosphorylation pathway mediates the effects of diabetic environment on oocyte competence.

MiR-130a-3p Regulates the Glycolysis via Targeting PDK1 in Hepatocellular Carcinoma

MiR-130a-3p has been certified to have low expression in several types of tumors. However, the function of miR-130a-3p in glucose metabolism and hepatocellular carcinoma progression is still elusive. Here we report that miR-130a-3p has explicitly low expression in human HCC tissues and cells and is closely related to the patient's tumor size and grade. Overexpression of miR-130a-3p significantly inhibits the glucose metabolism, proliferation and migration of HCC cells in vitro. In order to further study the effects of miR-130a-3p in the glucose metabolism of HCC cells, we found that overexpression of miR-130a-3p significantly inhibited the expression of pyruvate dehydrogenase kinase 1 (PDK1). Consistently, we confirmed that PDK1 is the target gene of miR-130a-3p through dual luciferase reporter gene assays. Cell rescue experiments showed that PDK1 inhibitors reversed the enhancement of cell proliferation, migration and glucose metabolism by miR-130a-3p inhibitor in Hep3B cells. In terms of mechanism, overexpression of miR-130a-3p targeted and inhibited the expression of PDK1, after which pyruvate dehydrogenase (PDH) is activated, thus glycolysis is inhibited, the production of lactic acid and ATP is reduced, and the ability to proliferate and migrate in HCC cells is weakened. In conclusion, our study highlights efforts to target PDK1 and miR-130a-3p as potential therapeutic strategies for the treatment of HCC.

Blood progenitor redox homeostasis through olfaction-derived systemic GABA in hematopoietic growth control in Drosophila

ABSTRACT The role of reactive oxygen species (ROS) in myeloid development is well established. However, its aberrant generation alters hematopoiesis. Thus, a comprehensive understanding of events controlling ROS homeostasis forms the central focus of this study. We show that, in homeostasis, myeloid-like blood progenitor cells of the Drosophila larvae, which reside in a specialized hematopoietic organ termed the lymph gland, use TCA to generate ROS. However, excessive ROS production leads to lymph gland growth retardation. Therefore, to moderate blood progenitor ROS, Drosophila larvae rely on olfaction and its downstream systemic GABA. GABA internalization and its breakdown into succinate by progenitor cells activates pyruvate dehydrogenase kinase (PDK), which controls inhibitory phosphorylation of pyruvate dehydrogenase (PDH). PDH is the rate-limiting enzyme that connects pyruvate to the TCA cycle and to oxidative phosphorylation. Thus, GABA metabolism via PDK activation maintains TCA activity and blood progenitor ROS homeostasis, and supports normal lymph gland growth. Consequently, animals that fail to smell also fail to sustain TCA activity and ROS homeostasis, which leads to lymph gland growth retardation. Overall, this study describes the requirement of animal odor-sensing and GABA in myeloid ROS regulation and hematopoietic growth control.

Cardioprotective Effects of Palmitoleic Acid (C16:1n7) in a Mouse Model of Catecholamine-Induced Cardiac Damage Are Mediated by PPAR Activation

Palmitoleic acid (C16:1n7) has been identified as a regulator of physiological cardiac hypertrophy. In the present study, we aimed to investigate the molecular pathways involved in C16:1n7 responses in primary murine cardiomyocytes (PCM) and a mouse model of isoproterenol (ISO)-induced cardiac damage. PCMs were stimulated with C16:1n7 or a vehicle. Afterwards, RNA sequencing was performed using an Illumina HiSeq sequencer. Confirmatory analysis was performed in PCMs and HL-1 cardiomyocytes. For an in vivo study, 129 sv mice were orally treated with a vehicle or C16:1n7 for 22 days. After 5 days of pre-treatment, the mice were injected with ISO (25 mg/kg/d s. c.) for 4 consecutive days. Cardiac phenotyping was performed using echocardiography. In total, 129 genes were differentially expressed in PCMs stimulated with C16:1n7, including Angiopoietin-like factor 4 (Angptl4) and Pyruvate Dehydrogenase Kinase 4 (Pdk4). Both Angptl4 and Pdk4 are proxisome proliferator-activated receptor α/δ (PPARα/δ) target genes. Our in vivo results indicated cardioprotective and anti-fibrotic effects of C16:1n7 application in mice. This was associated with the C16:1n7-dependent regulation of the cardiac PPAR-specific signaling pathways. In conclusion, our experiments demonstrated that C16:1n7 might have protective effects on cardiac fibrosis and inflammation. Our study may help to develop future lipid-based therapies for catecholamine-induced cardiac damage.

Impact of Sodium Dichloroacetate Alone and in Combination Therapies on Lung Tumor Growth and Metastasis

Metabolic reprogramming has been recognized as an essential emerging cancer hallmark. Dichloroacetate (DCA), an inhibitor of pyruvate dehydrogenase kinase (PDK), has been reported to have anti-cancer effects by reversing tumor-associated glycolysis. This study was performed to explore the anti-cancer potential of DCA in lung cancer alone and in combination with chemo- and targeted therapies using two non-small cell lung cancer (NSCLC) cell lines, namely, A549 and LNM35. DCA markedly caused a concentration- and time-dependent decrease in the viability and colony growth of A549 and LNM35 cells in vitro. DCA also reduced the growth of tumor xenografts in both a chick embryo chorioallantoic membrane and nude mice models in vivo. Furthermore, DCA decreased the angiogenic capacity of human umbilical vein endothelial cells in vitro. On the other hand, DCA did not inhibit the in vitro cellular migration and invasion and the in vivo incidence and growth of axillary lymph nodes metastases in nude mice. Treatment with DCA did not show any toxicity in chick embryos and nude mice. Finally, we demonstrated that DCA significantly enhanced the anti-cancer effect of cisplatin in LNM35. In addition, the combination of DCA with gefitinib or erlotinib leads to additive effects on the inhibition of LNM35 colony growth after seven days of treatment and to synergistic effects on the inhibition of A549 colony growth after 14 days of treatment. Collectively, this study demonstrates that DCA is a safe and promising therapeutic agent for lung cancer.

The Key Role of the WNT/β-Catenin Pathway in Metabolic Reprogramming in Cancers under Normoxic Conditions

The canonical WNT/β-catenin pathway is upregulated in cancers and plays a major role in proliferation, invasion, apoptosis and angiogenesis. Nuclear β-catenin accumulation is associated with cancer. Hypoxic mechanisms lead to the activation of the hypoxia-inducible factor (HIF)-1α, promoting glycolytic and energetic metabolism and angiogenesis. However, HIF-1α is degraded by the HIF prolyl hydroxylase under normoxia, conditions under which the WNT/β-catenin pathway can activate HIF-1α. This review is therefore focused on the interaction between the upregulated WNT/β-catenin pathway and the metabolic processes underlying cancer mechanisms under normoxic conditions. The WNT pathway stimulates the PI3K/Akt pathway, the STAT3 pathway and the transduction of WNT/β-catenin target genes (such as c-Myc) to activate HIF-1α activity in a hypoxia-independent manner. In cancers, stimulation of the WNT/β-catenin pathway induces many glycolytic enzymes, which in turn induce metabolic reprogramming, known as the Warburg effect or aerobic glycolysis, leading to lactate overproduction. The activation of the Wnt/β-catenin pathway induces gene transactivation via WNT target genes, c-Myc and cyclin D1, or via HIF-1α. This in turn encodes aerobic glycolysis enzymes, including glucose transporter, hexokinase 2, pyruvate kinase M2, pyruvate dehydrogenase kinase 1 and lactate dehydrogenase-A, leading to lactate production. The increase in lactate production is associated with modifications to the tumor microenvironment and tumor growth under normoxic conditions. Moreover, increased lactate production is associated with overexpression of VEGF, a key inducer of angiogenesis. Thus, under normoxic conditions, overstimulation of the WNT/β-catenin pathway leads to modifications of the tumor microenvironment and activation of the Warburg effect, autophagy and glutaminolysis, which in turn participate in tumor growth.

Sex differences in metabolic pathways are regulated by Pfkfb3 and Pdk4 expression in rodent muscle

Skeletal muscles display sexually dimorphic features. Biochemically, glycolysis and fatty acid β-oxidation occur preferentially in the muscles of males and females, respectively. However, the mechanisms of the selective utilization of these fuels remains elusive. Here, we obtain transcriptomes from quadriceps type IIB fibers of untreated, gonadectomized, and sex steroid-treated mice of both sexes. Analyses of the transcriptomes unveil two genes, Pfkfb3 (phosphofructokinase-2) and Pdk4 (pyruvate dehydrogenase kinase 4), that may function as switches between the two sexually dimorphic metabolic pathways. Interestingly, Pfkfb3 and Pdk4 show male-enriched and estradiol-enhanced expression, respectively. Moreover, the contribution of these genes to sexually dimorphic metabolism is demonstrated by knockdown studies with cultured type IIB muscle fibers. Considering that skeletal muscles as a whole are the largest energy-consuming organs, our results provide insights into energy metabolism in the two sexes, during the estrus cycle in women, and under pathological conditions involving skeletal muscles.