Exercise and Omega-3 Polyunsaturated Fatty Acid Supplementation for the Treatment of Hepatic Steatosis in Hyperphagic OLETF Rats

Background and Aims. This study examined if exercise and omega-3 fatty acid (n3PUFA) supplementation is an effective treatment for hepatic steatosis in obese, hyperphagic Otsuka Long-Evans Tokushima Fatty (OLETF) rats. Methods. Male OLETF rats were divided into 4 groups (n=8/group): (1) remained sedentary (SED), (2) access to running wheels; (EX) (3) a diet supplemented with 3% of energy from fish oil (n3PUFA-SED); and (4) n3PUFA supplementation plus EX (n3PUFA+EX). The 8 week treatments began at 13 weeks, when hepatic steatosis is present in OLETF-SED rats. Results. EX alone lowered hepatic triglyceride (TAG) while, in contrast, n3PUFAs failed to lower hepatic TAG and blunted the ability of EX to decrease hepatic TAG levels in n3PUFAs+EX. Insulin sensitivity was improved in EX animals, to a lesser extent in n3PUFA+EX rats, and did not differ between n3PUFA-SED and SED rats. Only the EX group displayed higher complete hepatic fatty acid oxidation (FAO) to CO2 and carnitine palmitoyl transferase-1 activity. EX also lowered hepatic fatty acid synthase protein while both EX and n3PUFA+EX decreased stearoyl CoA desaturase-1 protein. Conclusions. Exercise lowers hepatic steatosis through increased complete hepatic FAO, insulin sensitivity, and reduced expression of de novo fatty acid synthesis proteins while n3PUFAs had no effect

Discovery of a small molecule probe that post-translationally stabilizes the survival motor neuron protein for the treatment of spinal muscular atrophy.

Spinal muscular atrophy (SMA) is the leading genetic cause of infant death. We previously developed a high-throughput assay that employs an SMN2-luciferase reporter allowing identification of compounds that act transcriptionally, enhance exon recognition, or stabilize the SMN protein. We describe optimization and characterization of an analog suitable for in vivo testing. Initially, we identified analog 4m that had good in vitro properties but low plasma and brain exposure in a mouse PK experiment due to short plasma stability; this was overcome by reversing the amide bond and changing the heterocycle. Thiazole 27 showed excellent in vitro properties and a promising mouse PK profile, making it suitable for in vivo testing. This series post-translationally stabilizes the SMN protein, unrelated to global proteasome or autophagy inhibition, revealing a novel therapeutic mechanism that should complement other modalities for treatment of SMA

Beta 3 Adrenergic Receptor Activation Rescues Metabolic Dysfunction in Female Estrogen Receptor Alpha-Null Mice

Metabolic disease risk escalates following menopause. The mechanism is not fully known, but likely involves reduced signaling through estrogen receptor alpha (ERα), which is highly expressed in brown and white adipose tissue (BAT and WAT).Objective: Test the hypothesis that uncoupling protein (UCP1) activation mitigates metabolic dysfunction caused by loss of signaling through ERα.Methods: At 8 weeks of age, female ERα knock out (KO) and wild-type mice were housed at 28°C and fed a Western-style high-fat, high sucrose diet (HFD) or a normal low-fat chow diet (NC) for 10 weeks. During the final 2 weeks, they received daily injections of CL 316,256 (CL), a selective β3 adrenergic agonist, or vehicle control (CTRL), creating eight groups: WT-CTRL, WT-CL, KO-CTRL, and KO-CL on HFD or NC; n = 4–10/group.Results: ERαKO demonstrated exacerbated HFD-induced adiposity gain (P < 0.001) and insulin resistance (P = 0.006). CL treatment improved insulin sensitivity (P < 0.05) and normalized ERαKO-induced adiposity increase (P < 0.05). In both genotypes, CL increased resting energy expenditure (P < 0.05) and induced WAT beiging indicated by increased UCP1 protein in both perigonadal (PGAT) and subcutaneous (SQAT) depots. These effects were attenuated under HFD conditions (P < 0.05). In KO, CL reduced HFD energy consumption compared to CTRL (P < 0.05). Remarkably, CL increased WAT ERβ protein levels of both WT and KO (P < 0.001), revealing CL-mediated changes in estrogen signaling may have protective metabolic effects.Conclusion: CL completely restored metabolic dysfunction in ERαKO mice. Thus, UCP1 may be a therapeutic target for treating metabolic dysfunction following loss of estrogen receptor signaling

Novel Common Genetic Susceptibility Loci for Colorectal Cancer

BACKGROUND: Previous genome-wide association studies (GWAS) have identified 42 loci (P < 5 × 10-8) associated with risk of colorectal cancer (CRC). Expanded consortium efforts facilitating the discovery of additional susceptibility loci may capture unexplained familial risk. METHODS: We conducted a GWAS in European descent CRC cases and control subjects using a discovery-replication design, followed by examination of novel findings in a multiethnic sample (cumulative n = 163 315). In the discovery stage (36 948 case subjects/30 864 control subjects), we identified genetic variants with a minor allele frequency of 1% or greater associated with risk of CRC using logistic regression followed by a fixed-effects inverse variance weighted meta-analysis. All novel independent variants reaching genome-wide statistical significance (two-sided P < 5 × 10-8) were tested for replication in separate European ancestry samples (12 952 case subjects/48 383 control subjects). Next, we examined the generalizability of discovered variants in East Asians, African Americans, and Hispanics (12 085 case subjects/22 083 control subjects). Finally, we examined the contributions of novel risk variants to familial relative risk and examined the prediction capabilities of a polygenic risk score. All statistical tests were two-sided. RESULTS: The discovery GWAS identified 11 variants associated with CRC at P < 5 × 10-8, of which nine (at 4q22.2/5p15.33/5p13.1/6p21.31/6p12.1/10q11.23/12q24.21/16q24.1/20q13.13) independently replicated at a P value of less than .05. Multiethnic follow-up supported the generalizability of discovery findings. These results demonstrated a 14.7% increase in familial relative risk explained by common risk alleles from 10.3% (95% confidence interval [CI] = 7.9% to 13.7%; known variants) to 11.9% (95% CI = 9.2% to 15.5%; known and novel variants). A polygenic risk score identified 4.3% of the population at an odds ratio for developing CRC of at least 2.0. CONCLUSIONS: This study provides insight into the architecture of common genetic variation contributing to CRC etiology and improves risk prediction for individualized screenin

Unveiling anti-oxidative and anti-inflammatory effects of docosahexaenoic acid and its lipid peroxidation product on lipopolysaccharide-stimulated BV-2 microglial cells

Abstract Background Phospholipids in the central nervous system are enriched in n-3 and n-6 polyunsaturated fatty acids (PUFA), especially docosahexaenoic acid (DHA) and arachidonic acid (ARA). These PUFA can undergo enzymatic reactions to produce lipid mediators, as well as reaction with oxygen free radicals to produce 4-hydroxyhexenal (4-HHE) from DHA and 4-hydroxynonenal (4-HNE) from ARA. Recent studies demonstrated pleiotropic properties of these peroxidation products through interaction with oxidative and anti-oxidant response pathways. In this study, BV-2 microglial cells were used to investigate ability for DHA, 4-HHE, and 4-HNE to stimulate the anti-oxidant stress responses involving the nuclear factor erythroid-2-related factor 2 (Nrf2) pathway and synthesis of heme oxygenase (HO-1), as well as to mitigate lipopolysaccharide (LPS)-induced nitric oxide (NO), reactive oxygen species (ROS), and cytosolic phospholipase A2 (cPLA2). In addition, LC-MS/MS analysis was carried out to examine effects of exogenous DHA and LPS stimulation on endogenous 4-HHE and 4-HNE levels in BV-2 microglial cells. Methods Effects of DHA, 4-HHE, and 4-HNE on LPS-induced NO production was determined using the Griess reagent. LPS-induced ROS production was measured using CM-H2DCFDA. Western blots were used to analyze expression of p-cPLA2, Nrf2, and HO-1. Cell viability and cytotoxicity were measured using the WST-1 assay, and cell protein concentrations were measured using the BCA protein assay kit. An ultra-high-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis was used to determine levels of free 4-HHE and 4-HNE in cells. Results DHA (12.5–100 μM), 4-HHE (1.25–10 μM), and 4-HNE (1.25–10 μM) dose dependently suppressed LPS-induced production of NO, ROS, and as p-cPLA2 in BV-2 microglial cells. With the same concentrations, these compounds could enhance Nrf2 and HO-1 expression in these cells. Based on the estimated IC50 values, 4-HHE and 4-HNE were five- to tenfold more potent than DHA in inhibiting LPS-induced NO, ROS, and p-cPLA2. LC-MS/MS analysis indicated ability for DHA (10–50 μM) to increase levels of 4-HHE and attenuate levels of 4-HNE in BV-2 microglial cells. Stimulation of cells with LPS caused an increase in 4-HNE which could be abrogated by cPLA2 inhibitor. In contrast, bromoenol lactone (BEL), a specific inhibitor for the Ca2+-independent phospholipase A2 (iPLA2), could only partially suppress levels of 4-HHE induced by DHA or DHA + LPS. Conclusions This study demonstrated the ability of DHA and its lipid peroxidation products, namely, 4-HHE and 4-HNE at 1.25–10 μM, to enhance Nrf2/HO-1 and mitigate LPS-induced NO, ROS, and p-cPLA2 in BV-2 microglial cells. In addition, LC-MS/MS analysis of the levels of 4-HHE and 4-HNE in microglial cells demonstrates that increases in production of 4-HHE from DHA and 4-HNE from LPS are mediated by different mechanisms