Omega-3 Supplementation Prevents Short-Term High-Fat Diet Effects on the α7 Nicotinic Cholinergic Receptor Expression and Inflammatory Response.

The study is aimed at investigating if PUFA supplementation could prevent the effects of a short-term HFD on α7nAChR expression and on the severity of sepsis. Swiss mice were used for the in vivo experiments. For the in vitro experiments, we used a microglia cell line (BV-2) and a hepatoma cell line (Hepa-1c1c7) derived from mice. The animals were either fed standard chow, fed a short-term HFD (60%), or given supplementation with omega-3 fatty acid (2 g/kg or 4 g/kg bw) for 17 days, followed by a short-term HFD. Endotoxemia was induced with an intraperitoneal (i.p.) lipopolysaccharide injection (LPS, 5 or 12 mg/kg), and sepsis was induced by subjecting the animals to cecal ligation and puncture (CLP). BV-2 and Hepa-1c1c7 cells were treated with LPS (100 and 500 ng/mL, respectively) for 3 hours. RT-PCR or Western blotting was used to evaluate α7nAChR expression, inflammatory markers, DNMT1, and overall ubiquitination. LPS and HFD reduced the expression of α7nAChR and increased the expression of inflammatory markers. Omega-3 partially prevented the damage caused by the HFD to the expression of α7nAChR in the bone marrow and hypothalamus, decreased the inflammatory markers, and reduced susceptibility to sepsis-induced death. Exposing the BV-2 cells to LPS increased the protein content of DNMT1 and the overall ubiquitination and reduced the expression of α7nAChR. The inflammation induced by LPS in the BV-2 cell decreased α7nAChR expression and concomitantly increased DNMT1 expression and the ubiquitinated protein levels, indicating the participation of pre- and posttranscriptional mechanisms.

Maternal high fat diet consumption reduces liver alpha7 nicotinic cholinergic receptor expression and impairs insulin signalling in the offspring.

The activation of nicotinic acetylcholine receptor α7 subunit (α7nAChR) has been associated to anti-inflammatory response in macrophages. High-fat diet (HFD) consumption during pregnancy and lactation impairs the cholinergic anti-inflammatory pathway in liver and white adipose tissue of offspring. In order to evaluate the relationship between damage in the cholinergic anti-inflammatory pathway and insulin resistance (IR) development, the liver of offspring of obese dams was investigated. Additionally, the capacity of α7nAChR activation to reduce IR induced by saturated fatty acid was investigated in hepatoma cell line. Initially, female mice were subjected to either standard chow (SC) or HFD during pregnancy and lactation period. After weaning, only male offspring from HFD dams (HFD-O) and SC dams (SC-O) were fed with the SC diet. Hepatic α7nAChR expression was downregulated, and hepatic TNF-α, IL-1ß, and pIKK level, but not pJNK, were elevated in the HFD-O compared to SC-O mice. Besides, hepatic expression of TNF-α in response to lipopolysaccharide (LPS) was higher in HFD-O than SC-O mice. Insulin-stimulated phosphorylation of the AKT was lower in HFD-O compared to SC-O. Additionally, insulin-stimulated phosphorylation of the AKT in KOα7Alb-Cre mice fed HFD was lower than WT mice fed HFD. In hepatoma cell line, palmitate increased IL-6 and TNF-α expressions and pJNK level. These effects were accompanied by reduced capacity of insulin to stimulate AKT phosphorylation. PNU or nicotine reduced cytokine expression and JNK activation, but improved insulin resistance induced by palmitate. Our results suggest that maternal obesity impairs hepatic α7nAChR expression and AKT phosphorylation in the offspring. In vitro studies suggest that α7nAChR activation has potential to reduce deleterious effect of saturated fatty acids on insulin signalling.

Hypothalamic Neuropeptides Expression and Hypothalamic Inflammation in Adult Rats that Were Exposed to Tobacco Smoke during Breastfeeding: Sex-Related Differences.

The hypothalamus controls food intake and energy expenditure. In rats, maternal exposure to nicotine during breastfeeding alters the hypothalamic circuitry of the adult offspring, resulting in leptin resistance, neuropeptides changes and gliosis. Tobacco smoke exposure during lactation causes greater adiposity, hyperphagia and hyperleptinemia in the adult progeny. To understand the central mechanisms underlying the obese phenotype of adult rats that were directly and indirectly exposed to cigarette smoke during lactation, we investigated leptin signaling, orexigenic and anorexigenic neuropeptides expression, as well as astrocyte and microglia markers in hypothalamus. From postnatal day (PND) 3 to 21, Wistar lactating rat dams and their pups were divided into two groups: SE, smoke-exposed in a cigarette-smoking machine (four times/day); Crtl, exposed to filtered air. Offspring of both sexes were euthanized at PND180. The leptin pathway was not altered in SE animals from both sexes. SE males showed increased NPY (arcuate nucleus, ARC), CRH (paraventricular nucleus, PVN), as well as higher GFAP fiber density (ARC and PVN) and IL6 protein content. TRH (PVN) immunohistochemistry was reduced. SE females had lower CART-positive cells (ARC) and lower α-MSH immunostaining intensity (PVN and lateral hypothalamus), with no change of GFAP or IL-6. The protein contents of CX3CR1 (marker of activated microglia) and α7nAChR (anti-inflammatory marker) were not altered in both SE males and females. Neonatal cigarette smoke is deleterious to the hypothalamic circuitry, inducing changes in energy homeostasis favoring hyperphagia and decreased energy expenditure at adulthood in both sexes; however sex-dependent mechanisms were observed.

Hypothalamic AMPK activation blocks lipopolysaccharide inhibition of glucose production in mice liver.

Endotoxic hypoglycaemia has an important role in the survival rates of septic patients. Previous studies have demonstrated that hypothalamic AMP-activated protein kinase (hyp-AMPK) activity is sufficient to modulate glucose homeostasis. However, the role of hyp-AMPK in hypoglycaemia associated with endotoxemia is unknown. The aims of this study were to examine hyp-AMPK dephosphorylation in lipopolysaccharide (LPS)-treated mice and to determine whether pharmacological hyp-AMPK activation could reduce the effects of endotoxemia on blood glucose levels. LPS-treated mice showed reduced food intake, diminished basal glycemia, increased serum TNF-α and IL-1ß levels and increased hypothalamic p-TAK and TLR4/MyD88 association. These effects were accompanied by hyp-AMPK/ACC dephosphorylation. LPS-treated mice also showed diminished liver expression of PEPCK/G6Pase, reduction in p-FOXO1, p-AMPK, p-STAT3 and p-JNK level and glucose production. Pharmacological hyp-AMPK activation blocked the effects of LPS on the hyp-AMPK phosphorylation, liver PEPCK expression and glucose production. Furthermore, the effects of LPS were TLR4-dependent because hyp-AMPK phosphorylation, liver PEPCK expression and fasting glycemia were not affected in TLR4-mutant mice. These results suggest that hyp-AMPK activity may be an important pharmacological target to control glucose homeostasis during endotoxemia.

Serum levels and mesenteric fat tissue expression of adiponectin and leptin in patients with Crohn's disease.

Crohn's disease (CD) is characterized by inflammation and an aetiology that is still unknown. Hypertrophy of mesenteric fat is a reflection of disease activity, as this fat covers the entire length of the affected area. Adipocytes synthesize leptin and adiponectin, adipocytokines responsible for pro- and anti-inflammatory effects. Therefore, we evaluated serum levels of adiponectin and leptin, as well as mesenteral expression of adiponectin in active CD and those in remission. Sixteen patients with ileocaecal CD followed at the Outpatient Clinic, Coloproctology Unit of University of Campinas Clinical Hospital, participated in the study. Analysis of serum adiponectin and leptin by enzyme-linked immunosorbent assay was performed in patients with active CD (ACD group), remission CD (RCD group) and in six healthy controls. Ten patients with active ileocaecal CD (FCD group) and eight patients with non-inflammatory disease selected for surgery were also studied. The specimens were snap-frozen and the expression of adiponectin was determined by immunoblot of protein extracts. Serum C-reactive protein levels were higher in the ACD group when compared to the others and no difference of body mass index was observed between the groups. Serum adiponectin was lower in the ACD group when compared to control, but no differences were seen when comparing the ACD and RCD groups. Mesenteric adiponectin expression was lower in the FCD group when compared to the FC group. Serum leptin was similar in all groups. The lower levels of serum and mesenteric adiponectin in active CD suggest a defective regulation of anti-inflammatory pathways in CD pathogenesis.

Early activation of the multicomponent signaling complex associated with focal adhesion kinase induced by pressure overload in the rat heart.

Mechanical overload elicits functional and structural adaptive mechanisms in cardiac muscle. Signaling pathways linked to integrin/cytoskeleton complexes may have a function in mediation of the effects of mechanical stimulus in myocardial cells. We investigated the tyrosine phosphorylation and the assembly of the multicomponent signaling complex associated with focal adhesion kinase (Fak) and the actin cytoskeleton in the overloaded myocardium of rats. Pressure overload induced a 3-fold increase in Fak tyrosine phosphorylation within 3 minutes after a 60-mm Hg rise in aortic pressure. A pressure stimulus that lasted for 60 minutes was accompanied by a 5-fold increase in the amount of tyrosine-phosphorylated Fak, and a stimulus as low as 10 mm Hg doubled the amount of tyrosine-phosphorylated Fak in the myocardium within 10 minutes. Pressure overload also induced a time-dependent association of actin with Fak and an increase in the amount of Fak detected in the cytoskeletal fraction of the myocardium. These events were paralleled by c-Src activation and binding to Fak and by an association of Grb2 and p85 subunit of phosphatidylinositol 3-kinase with Fak. Erk1/2 and Akt, two possible downstream effectors of Fak via Grb2 and phosphatidylinositol 3-kinase, were also shown to be activated in parallel with Fak. These findings show that pressure overload induced a rapid activation of the Fak multiple signaling complex in the myocardium of rats, which suggests that this mechanism may have a role in mechanotransduction in the myocardium.