Toll-like receptor-2 deficiency enhances non-alcoholic steatohepatitis.
BACKGROUND
Previously we reported that mice deficient in toll-like receptor 4 (TLR-4) signalling were protected from diet-induced non-alcoholic steatohepatitis (NASH). Another member of the toll-like receptor family, TLR-2, has been shown to play a role in lipid trafficking via uptake of diacylated lipoproteins. However, a role for TLR-2 in NASH has not been elucidated. The objectives of the current study were to examine the influence of dietary fat quality and TLR-2 on NASH pathogenesis.


METHODS
Steatohepatitis was induced in male Db, C57BL/6 and TLR-2(-/-) mice by feeding an L-amino acid-defined diet that was deficient in methionine and choline (MCDD). Mice fed the base diet supplemented with methionine and choline (control diet; CD) were used as controls. To determine the role of fat quality, MCDD was enriched with polyunsaturated corn oil (PUFA) or coconut oil that is comprised mostly of saturated fat (SAFA); the total amount of each fat was 112.9 g/kg of diet. After 8 weeks of feeding CD or MCDD, hepatic steatosis, inflammation and necrosis were evaluated in histological sections. Total RNA was extracted from frozen liver samples and mRNA expression of TNFalpha, collagen alpha1, IL-10, peroxisome proliferator-activated receptor-gamma (PPAR-gamma), TLR-4, and CD14, was analyzed via real-time PCR. Protein levels of TLR-2 were analyzed by western blot.


RESULTS
Panlobular macrovessicular steatosis and diffuse leukocyte infiltration were noted in PUFA-fed Db mice. Histological scores demonstrated significantly less steatosis, inflammation and necrosis in SAFA-fed mice of all mouse strains. However, compared to wild type mice, hepatocellular damage was notably more severe in TLR-2(-/-) mice. Consistent with histological findings, mRNA expression of TNFalpha was elevated by approximately 3-fold in TLR-2(-/-) mice; PPAR-gamma expression was blunted in this strain compared to wild type. Expression of the matrix protein collagen alphaI was also significantly higher in TLR-2(-/-) mice, indicating a pro-fibrogenic state. Sensitivity to steatohepatitis due to dietary fat or TLR-2 deficiency correlated significantly with alterations in the expression of TLR-4 as well as the co-receptor CD-14.


CONCLUSIONS
Our findings suggest that dietary saturated fat plays a protective role against MCDD-induced steatohepatitis, whereas TLR-2 deficiency exacerbated NASH. The mechanism underlying the response to dietary fat and TLR-2 likely involves altered signalling via the TLR-4 pathway.
Department of Molecular and Cellular Physiology, Louisiana State University Health Sciences Center, Shreveport, LA, USA. crive1@lsuhsc.edu
Chantal A
CA
Rivera
LaTausha
L
Gaskin
Monique
M
Allman
Jia
J
Pang
Kristen
K
Brady
Patrick
P
Adegboyega
Kevin
K
Pruitt
D000818:Animals; D002796:Choline Deficiency; D024042:Collagen Type I / Q000378:metabolism; D004041:Dietary Fats / Q000009:adverse effects*; D004042:Dietary Fats, Unsaturated / Q000009:adverse effects; D004195:Disease Models, Animal; D005234:Fatty Liver / Q000209:etiology* / Q000378:metabolism; D008297:Male; D008715:Methionine / Q000172:deficiency*; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D047495:PPAR gamma / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology; D051195:Toll-Like Receptor 2 / Q000172:deficiency* / Q000235:genetics; D051197:Toll-Like Receptor 4 / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
D024042:Collagen Type I; D004041:Dietary Fats; D004042:Dietary Fats, Unsaturated; D047495:PPAR gamma; C493486:Tlr2 protein, mouse; C493487:Tlr4 protein, mouse; D051195:Toll-Like Receptor 2; D051197:Toll-Like Receptor 4; D014409:Tumor Necrosis Factor-alpha; D008715:Methionine