Clinical trial NCT02148471: Fatty Acids, Genes and Microbiota in Fatty Liver
Official title: Non-alcoholic Steatohepatitis Versus Simple Hepatic Steatosis: Is There a Difference in the Nutritional Factors Influencing Lipid Perioxidation and Inflammation?
Study type: OBSERVATIONAL
Overall status: COMPLETED
Conditions: Nonalcoholic Fatty Liver Disease
Steatosis
Nonalcoholic Steatohepatitis
Keywords: Nonalcoholic fatty liver disease
Steatosis
Nonalcoholic steatohepatitis
Lead sponsor: University Health Network, Toronto
Sponsor class: OTHER
Collaborators: name: Canadian Liver Foundation
name: Canadian Institutes of Health Research (CIHR)
name: American College of Gastroenterology
First posted: 2014-05-28
Enrollment: 205
Sex: ALL
Minimum age (years): 18
Maximum age (years): 70
Standard ages: ADULT
OLDER_ADULT
Accepts healthy volunteers: yes
Observational model: COHORT
Time perspective: CROSS_SECTIONAL
Sampling method: NON_PROBABILITY_SAMPLE
Study population: Healthy living liver donors from the Multiorgan transplant program at the University Health Network Patients with nonalcoholic fatty liver disease (SS or NASH on liver biopsy) or patients with elevated liver enzymes but no significant findings on liver biopsy.

Brief summary: The first aim of this study is to assess oxidative stress and nutritional status in patients with elevated liver enzymes who were found to have either simple steatosis (SS) or nonalcoholic steatohepatitis (NASH) or normal histological findings on liver biopsy by measuring liver lipid peroxides and tumor necrosis factor (TNF)-α, liver pathology and immunohistochemistry, liver function tests, liver and red blood cell membrane fatty composition, insulin resistance (IR) parameters, plasma lipid peroxides, plasma antioxidant vitamins and antioxidant power, lipid profile, subject demographics, medical history and medication use. The second aim is to detect differences in hepatic gene expression (messenger RNA, mRNA) and epigenetic regulation (micro RNA, miRNA) between patients with SS or NASH and healthy controls, in addition to determine in patients with non-alcoholic fatty liver disease (NAFLD = SS+NASH combined) whether there is an association between hepatic n-3 PUFA content and gene expression. The third aim is to determine the intestinal microbiome (microbial composition and metagenome) in patients with SS or NASH and healthy controls.

Detailed description: NASH is associated with obesity, diabetes and hyperlipidemia. Fat accumulation in the liver is likely due to variable degrees of disordered fatty-acid metabolism and insulin resistance (IR). Liver steatosis, especially polyunsaturated fatty acids (PUFA) in the liver, increases lipid peroxidation and is associated with a reduction in the antioxidant defense system. This oxidative stress can lead to increased production of pro-inflammatory cytokines (TNF-α, transforming growth factor-beta) contributing to the development of steatohepatitis and fibrosis.TNF-α - may further contribute to IR. In addition, changes in fatty acid composition within the liver may influence lipid metabolism and inflammation. In particular, n-3 PUFA have an effect on the insulin sensitivity, transcription of antioxidant genes, inflammatory response and production of reactive oxygen species. Differences might be seen on the gene expression level (mRNA) and also in epigenetic regulation (miRNA).

Microbiota composition might influence energy metabolism, and inflammatory tone and IR through increased endotoxemia and therefore could also play a role in the development of NAFLD.

Eligibility criteria: Inclusion criteria:

* Male and female patients, age \>18 y
* A liver biopsy with a diagnosis of SS or NASH OR No signs of steatosis, fibrosis or any other kind of liver disease on histology (minimal findings) OR For healthy control subjects, those with normal liver enzymes and normal liver imaging on ultrasound
* alcohol consumption (\<20g of ethanol per day);
* absence of any other possible cause for liver dysfunction.

Exclusion criteria:

* any other liver disease apart from NAFLD
* anticipated need for liver transplantation in one year or complications of liver disease;
* any reasons contraindicating a liver biopsy (patients) or liver donation (healthy donors)
* chronic gastrointestinal diseases, previous gastrointestinal surgery modifying the anatomy, patients with diabetes requiring insulin.
* medications known to precipitate steatohepatitis (corticosteroids, high dose estrogens, methotrexate, amiodarone, spironolactone, sulfasalazine, perhexiline maleate, diethylamino- ethoxyhexestrol (DH), tamoxifen, diethylstilbestrol, naproxen or oxacillin) or regular intake of non-steroidal anti-inflammatory drugs (except for low dose aspirin), use of ursodeoxycholic acid or any experimental drug in the 6 months prior to entry.
* regular intake of prebiotics, probiotics, antibiotics, or laxatives; in the 3 months prior to study entry
* Pregnant or lactating

Primary outcomes:
measure: Hepatic fatty acid composition in total lipids in liver biopsy
timeFrame: Baseline
measure: Hepatic gene expression
timeFrame: Baseline
measure: Intestinal microbiota composition
timeFrame: Baseline
Secondary outcomes:
measure: Lipid peroxides in the liver
timeFrame: Baseline
measure: Hepatic liver antioxidant power
timeFrame: Baseline
measure: Hepatic microRNA expression in the liver
timeFrame: Baseline
measure: Intestinal microbiota - specific organisms and groups
timeFrame: Baseline
measure: Intestinal microbiome on a genetic level
timeFrame: Baseline
measure: Short-chain fatty acids in stool
timeFrame: Baseline
measure: Plasma endotoxin
timeFrame: Baseline
Other outcomes:
measure: Hepatic phospholipid composition
timeFrame: Baseline
measure: Red blood cell fatty acid and phospholipid composition
timeFrame: Baseline
measure: Plasma fatty acid composition
timeFrame: Baseline
measure: Plasma lipid peroxides
timeFrame: Baseline
measure: Plasma antioxidant vitamins
timeFrame: Baseline
measure: Serum antioxidant power
timeFrame: Baseline
measure: TNF-alpha in the liver
timeFrame: Baseline
measure: Immunohistochemistry
timeFrame: Baseline
measure: Free choline in serum
timeFrame: Baseline
measure: Bacterial DNA in plasma
timeFrame: Baseline
measure: Insulin resistance
timeFrame: Baseline
measure: Plasma ethanol
timeFrame: Baseline
measure: Anthropometry
timeFrame: Baseline
measure: Food intake
timeFrame: Baseline
measure: Physical activity
timeFrame: Baseline
measure: Factors influencing intestinal microbiota
timeFrame: Baseline
measure: Liver function tests
timeFrame: Baseline
Locations:
city: Toronto
country: Canada
facility: Toronto General Hospital
state: Ontario
Study officials:
affiliation: University Health Network, Toronto
name: Johane Allard, MD,FRCPC
role: PRINCIPAL_INVESTIGATOR
References:
citation: Schwenger KJ, Allard JP. Clinical approaches to non-alcoholic fatty liver disease. World J Gastroenterol. 2014 Feb 21;20(7):1712-23. doi: 10.3748/wjg.v20.i7.1712.
citation: Monteiro J, Leslie M, Moghadasian MH, Arendt BM, Allard JP, Ma DW. The role of n - 6 and n - 3 polyunsaturated fatty acids in the manifestation of the metabolic syndrome in cardiovascular disease and non-alcoholic fatty liver disease. Food Funct. 2014 Mar;5(3):426-35. doi: 10.1039/c3fo60551e.
citation: Mouzaki M, Allard JP. The role of nutrients in the development, progression, and treatment of nonalcoholic fatty liver disease. J Clin Gastroenterol. 2012 Jul;46(6):457-67. doi: 10.1097/MCG.0b013e31824cf51e.
citation: Mouzaki M, Allard J. Non-alcoholic steatohepatitis: the therapeutic challenge of a global epidemic. Ann Gastroenterol. 2012;25(3):207-217.
citation: Da Silva HE, Arendt BM, Noureldin SA, Therapondos G, Guindi M, Allard JP. A cross-sectional study assessing dietary intake and physical activity in Canadian patients with nonalcoholic fatty liver disease vs healthy controls. J Acad Nutr Diet. 2014 Aug;114(8):1181-94. doi: 10.1016/j.jand.2014.01.009. Epub 2014 Mar 14.
citation: Arendt BM, Ma DW, Simons B, Noureldin SA, Therapondos G, Guindi M, Sherman M, Allard JP. Nonalcoholic fatty liver disease is associated with lower hepatic and erythrocyte ratios of phosphatidylcholine to phosphatidylethanolamine. Appl Physiol Nutr Metab. 2013 Mar;38(3):334-40. doi: 10.1139/apnm-2012-0261. Epub 2012 Oct 15.
citation: Mouzaki M, Comelli EM, Arendt BM, Bonengel J, Fung SK, Fischer SE, McGilvray ID, Allard JP. Intestinal microbiota in patients with nonalcoholic fatty liver disease. Hepatology. 2013 Jul;58(1):120-7. doi: 10.1002/hep.26319. Epub 2013 May 14.
citation: Allard JP, Aghdassi E, Mohammed S, Raman M, Avand G, Arendt BM, Jalali P, Kandasamy T, Prayitno N, Sherman M, Guindi M, Ma DW, Heathcote JE. Nutritional assessment and hepatic fatty acid composition in non-alcoholic fatty liver disease (NAFLD): a cross-sectional study. J Hepatol. 2008 Feb;48(2):300-7. doi: 10.1016/j.jhep.2007.09.009. Epub 2007 Nov 20.
citation: Arendt BM, Comelli EM, Ma DW, Lou W, Teterina A, Kim T, Fung SK, Wong DK, McGilvray I, Fischer SE, Allard JP. Altered hepatic gene expression in nonalcoholic fatty liver disease is associated with lower hepatic n-3 and n-6 polyunsaturated fatty acids. Hepatology. 2015 May;61(5):1565-78. doi: 10.1002/hep.27695. Epub 2015 Feb 27.
citation: Pettinelli P, Arendt BM, Schwenger KJP, Sivaraj S, Bhat M, Comelli EM, Lou W, Allard JP. Relationship Between Hepatic Gene Expression, Intestinal Microbiota, and Inferred Functional Metagenomic Analysis in NAFLD. Clin Transl Gastroenterol. 2022 Jul 1;13(7):e00466. doi: 10.14309/ctg.0000000000000466. Epub 2022 Feb 10.