Markers of activated inflammatory cells are associated with disease severity and intestinal microbiota in adults with non­alcoholic fatty liver disease.

Several mechanisms contribute to the pathogenesis of non­alcoholic fatty liver disease (NAFLD). The intestinal microbiota (IM) and liver immune cells (LIC) may serve a role, but there has been no previous study assessing potential associations between IM and LIC. The aim of the present study was to investigate whether there are differences in LIC markers between patients with NAFLD and healthy controls (HC), and to determine whether these markers are associated with specific IM. The present prospective, cross­sectional study examined a cohort of adults with liver biopsy­confirmed NAFLD and HC. Clinical and laboratory data were collected. Fecal IM was assessed by quantitative polymerase chain reaction and LIC, by immunohistochemistry. NAFLD activity score (NAS) was used for disease severity. Liver immune cell counts were increased in patients with NAFLD (n=34) vs. HC (n=8) and this was associated with disease severity. Hematopoietic cell marker cluster of differentiation (CD)45+ and Kupffer cell marker CD163+ were higher in NAFLD compared with HC, and those with an NAS ≥5 had higher levels of CD20+ cells, a marker of B cells, vs. a NAS of 0 or 1­4. Additionally, from those patients (5 HC, 34 NAFLD), IM was measured. Specific immune cells in portal or lobular areas correlated with specific fecal IM, suggesting a potential association between IM and liver inflammation in patients with NAFLD. Specifically, Faecalibacterium prausnitzii was negatively correlated with CD45+ (r= ­0.394; P=0.015) and CD163+ (r= ­0.371; P=0.022) cells in the portal tract and Prevotella was negatively correlated with CD20+ (r= ­0.353; P=0.028) cells in the liver lobule. Other taxa exhibited no correlation. In conclusion, the present study demonstrated a potential association between IM and liver inflammation in NAFLD.

Nonalcoholic fatty liver disease is associated with dysbiosis independent of body mass index and insulin resistance.

This study aimed to determine if there is an association between dysbiosis and nonalcoholic fatty liver disease (NAFLD) independent of obesity and insulin resistance (IR). This is a prospective cross-sectional study assessing the intestinal microbiome (IM) of 39 adults with biopsy-proven NAFLD (15 simple steatosis [SS]; 24 nonalcoholic steatohepatitis [NASH]) and 28 healthy controls (HC). IM composition (llumina MiSeq Platform) in NAFLD patients compared to HC were identified by two statistical methods (Metastats, Wilcoxon). Selected taxa was validated using quantitative PCR (qPCR). Metabolites in feces and serum were also analyzed. In NAFLD, 8 operational taxonomic units, 6 genera, 6 families and 2 phyla (Bacteroidetes, Firmicutes) were less abundant and; 1 genus (Lactobacillus) and 1 family (Lactobacillaceae) were more abundant compared to HC. Lower abundance in both NASH and SS patients compared to HC were confirmed by qPCR for Ruminococcus, Faecalibacterium prausnitzii and Coprococcus. No difference was found between NASH and SS. This lower abundance in NAFLD (NASH+SS) was independent of BMI and IR. NAFLD patients had higher concentrations of fecal propionate and isobutyric acid and serum 2-hydroxybutyrate and L-lactic acid. These findings suggest a potential role for a specific IM community and functional profile in the pathogenesis of NAFLD.

Cocaine craving during protracted withdrawal requires PKCε priming within vmPFC.

In individuals with a history of drug taking, the capacity of drug-associated cues to elicit indices of drug craving intensifies or incubates with the passage of time during drug abstinence. This incubation of cocaine craving, as well as difficulties with learning to suppress drug-seeking behavior during protracted withdrawal, are associated with a time-dependent deregulation of ventromedial prefrontal cortex (vmPFC) function. As the molecular bases for cocaine-related vmPFC deregulation remain elusive, the present study assayed the consequences of extended access to intravenous cocaine (6 hours/day; 0.25 mg/infusion for 10 day) on the activational state of protein kinase C epsilon (PKCε), an enzyme highly implicated in drug-induced neuroplasticity. The opportunity to engage in cocaine seeking during cocaine abstinence time-dependently altered PKCε phosphorylation within vmPFC, with reduced and increased p-PKCε expression observed in early (3 days) and protracted (30 days) withdrawal, respectively. This effect was more robust within the ventromedial versus dorsomedial PFC, was not observed in comparable cocaine-experienced rats not tested for drug-seeking behavior and was distinct from the rise in phosphorylated extracellular signal-regulated kinase observed in cocaine-seeking rats. Further, the impact of inhibiting PKCε translocation within the vmPFC using TAT infusion proteins upon cue-elicited responding was determined and inhibition coinciding with the period of testing attenuated cocaine-seeking behavior, with an effect also apparent the next day. In contrast, inhibitor pretreatment prior to testing during early withdrawal was without effect. Thus, a history of excessive cocaine taking influences the cue reactivity of important intracellular signaling molecules within the vmPFC, with PKCε playing a critical role in the manifestation of cue-elicited cocaine seeking during protracted drug withdrawal.

A cross-sectional study assessing dietary intake and physical activity in Canadian patients with nonalcoholic fatty liver disease vs healthy controls.

BACKGROUND: Poor diet and a sedentary lifestyle can contribute to nonalcoholic fatty liver disease (NAFLD). OBJECTIVE: Our aim was to compare diet and physical activity of patients with NAFLD and healthy controls with current recommendations. DESIGN: This was a cross-sectional study. PARTICIPANTS/SETTINGS: Seventy-four patients with biopsy-proven NAFLD (33 simple steatosis and 41 steatohepatitis [NASH]) and 27 healthy controls participated between 2003 and 2011. MAIN OUTCOME MEASURES: Food records and activity logs were completed for 7 days. Results were compared with Dietary Reference Intakes and Canadian Physical Activity Guidelines. Plasma vitamin C was measured to assess food record accuracy. STATISTICAL ANALYSES PERFORMED: Intake/activity for each participant was compared with the recommendations and proportion of subjects not meeting the requirements was calculated. Groups were compared by Kruskal-Wallis and Mann-Whitney U test or z-test with Bonferroni adjustment. RESULTS: More patients with NASH (58.5%) were obese compared with patients with simple steatosis (24.2%) and healthy controls (7.4%; P<0.01). Patients with NAFLD showed more insulin resistance than healthy controls. The reported energy intake was below estimated requirements in all groups (P≤0.001). The proportion of subjects from each group exceeding acceptable energy intake from fat was as follows: simple steatosis: 27.3%; NASH: 46.3%; healthy controls: 63.0% (simple steatosis vs health controls; P<0.05) and from saturated fat: simple steatosis: 42.4%; NASH: 70.7%; healthy controls: 63.0% (simple steatosis vs. NASH; P<0.05). In each group, >80% of subjects did not consume enough linoleic or linolenic acid, vitamin D, and vitamin E, and >60% exceeded the upper intake level for sodium. Only 53.1% of patients with simple steatosis and 53.8% of patients with NASH, but 84.6% of healthy controls, met recommendations for physical activity (P=0.020). Plasma vitamin C was normal, similar among groups, and correlated with vitamin C intakes. CONCLUSIONS: All participants followed a similar Western diet with high fat and sodium intakes and suboptimal micronutrient intakes. However, physical activity was lower in NAFLD compared with healthy controls and was associated with higher body mass index and insulin resistance.

Deficits in ventromedial prefrontal cortex group 1 metabotropic glutamate receptor function mediate resistance to extinction during protracted withdrawal from an extensive history of cocaine self-administration.

Anomalies in prefrontal cortex (PFC) function are posited to underpin difficulties in learning to suppress drug-seeking behavior during abstinence. Because group 1 metabotropic glutamate receptors (mGluRs) regulate drug-related learning, we assayed the consequences of extended access to intravenous cocaine (6 h/d; 0.25 mg/infusion for 10 d) on the PFC expression of group 1 mGluRs and the relevance of observed changes for cocaine seeking. After protracted withdrawal, cocaine-experienced animals exhibited a time-dependent intensification of cue-induced cocaine-seeking behavior and an impaired extinction of this behavior. These behavioral phenomena were associated with a time-dependent reduction in mGluR1/5 expression within ventromedial PFC (vmPFC) of cocaine-experienced animals exposed to extinction testing but not in untested ones. Interestingly, pharmacological manipulations of vmPFC mGluR1/5 produced no immediate effects on cue-induced cocaine-seeking behavior but produced residual effects on a subsequent test for cocaine seeking. At 3 d withdrawal, cocaine-experienced rats infused intra-vmPFC with mGluR1/5 antagonists, either before or after an initial test for cocaine seeking, persisted in their cocaine seeking akin to cocaine-experienced rats in protracted withdrawal. Conversely, cocaine-experienced rats infused with an mGluR1/5 agonist before the initial test for cocaine-seeking at 30 d withdrawal exhibited a facilitation of extinction learning. These data indicate that cue-elicited deficits in vmPFC group 1 mGluR function mediate resistance to extinction during protracted withdrawal from a history of extensive cocaine self-administration and pose pharmacological stimulation of these receptors as a potential approach to facilitate learned suppression of drug-seeking behavior that may aid drug abstinence.