Dietary Intervention and Intestinal Microbiota in Non-alcoholic Fatty Liver

• abundance of fecal abundance of fecal Bacteroidetes [ Time Frame: thrice, at inclusion, day 21 and day 42 ] [ Designated as safety issue: No ]
  Bacterial cells/g caecal content
  
  

• liver fat content [ Time Frame: twice, at inclusion and day 21 ] [ Designated as safety issue: No ]
  picsel/uni MRI
  
  
• liver function tests [ Time Frame: four times, at screening, inclusion, day 21 and day 42 ] [ Designated as safety issue: No ]
  mmol/l
  
  
• CRP [ Time Frame: thrice, at screening, inclusion and day 21 ] [ Designated as safety issue: No ]
  mg/l
  
  
• serum cytokines [ Time Frame: twice, at inclusion and day 21 ] [ Designated as safety issue: No ]
  ng/ml
  
  
• serum LPS [ Time Frame: twice, at inclusion and day 21 ] [ Designated as safety issue: No ]
  pg/ml
  
  
• Breath test pullulation [ Time Frame: twice, at inclusion and day 21 ] [ Designated as safety issue: No ]
  
• intestinal permeability [ Time Frame: twice, at inclusion and day 21 ] [ Designated as safety issue: No ]
  Polyethylene glycol 3350/Polyethylene glycol 400 ratio
  
  
• body composition [ Time Frame: thrice, at inclusion, day 21 and day 42 ] [ Designated as safety issue: No ]
  body composition in kg and % of body weight
  
  
• liver lipid composition (biopsy) [ Time Frame: once, at screening ] [ Designated as safety issue: No ]
  

Study Hypothesis: In patients with NAFLD/NASH, changes in liver lipid composition and function tests following a short dietary intervention are associated with changes in gut microbiota

Study period:

• Study duration for the participant: 7-10 weeks (1-4 weeks screening period, 3 weeks of intervention + 3 weeks of follow-up)
• Expected study completion date: 30.04.2012 Study type: Single arm before-after study

Number of patients:

20 subjects with obesity and NAFLD / NASH

Main criteria for inclusion:

• Obesity defined as BMI>30
• Abnormal liver function tests defined as ALT > 1.5 times the upper limit of normal
• NAFLD present at liver biopsy
• Age > 18 years, < 60 years

Main exclusion criteria:

• Inability or unwillingness to give consent
• Parenteral nutrition or other ongoing dietary intervention
• Bulimia
• Other known cause of chronic liver disease, including hepatitis B or C, iron overload,
• Use of substances known to alter intestinal permeability, including alcohol and NSAIDs

Intervention:

The phase 2 Eurodiet® program will be used as standardized hypo-caloric diet during a 3-weeks intervention period. The products are commercially available and prescribed to reach 1000 kcal/day. These products will be offered free of charge.

Primary Objective:

To assess the impact of dietary intervention on the relative abundance of fecal Bacteroidetes (expressed as the bacteroidetes to firmicutes ratio) in patients with obesity, abnormal liver function tests and NAFLD

Secondary Objectives:

1. To compare fecal microbiota from patients with NAFLD or NASH at baseline
2. To assess fecal microbiota changes in patients with NAFLD or NASH after dietary intervention
3. To measure liver fat content at baseline and after dietary intervention
4. To assess changes in liver function tests and ultrasensitive CRP, cytokines and serum LPS in relation to changes in microbiota and liver lipid composition
5. To measure orocecal transit time, an index of intestinal pullulation, at baseline and after dietary intervention in patients with NAFLD or NASH
6. To measure intestinal permeability at baseline and after dietary intervention in patients with NAFLD or NASH
7. To assess body composition changes in relation to changes in microbiota and liver lipid composition

Statistical methods:

Baseline and end-of-treatment changes for both bacterial genders and subspecies will be compared using paired-sample Wilcoxon signed-rank test. ANOVA and paired t-test for comparison of other changes within groups. Pearson or Spearman tests will be used to assess correlations between changes in microbiota and changes in liver fat content, liver function tests, CRP, cytokines and intestinal pullulation and permeability.

Sample size:

20 patients with NAFLD/NASH will be studied before and after dietary intervention.

Assessment of end-points:

Fecal microbiota will be analysed with 454-Flex metagenomics Ultrasensitive CRP and serum LPS changes as compared with baseline Serum cytokines as measured with ELISA Liver fat content and composition will be measured using MR spectroscopy Small intestinal overgrowth and intestinal permeability will be assessed using standard 13C breath tests and polyethyleneglycol absorption test Body mass composition changes will be assessed using bioelectrical impedance analysis

Safety :

All adverse events will be recorded throughout the study, in compliance with GCP ICH E6 and national regulations.