Oral Supplementation of Glutamine Attenuates the Progression of Nonalcoholic Steatohepatitis in C57BL/6J Mice

Abstract

Universally accepted therapeutic strategies for the treatment of nonalcoholic steatohepatitis (NASH) are still lacking. Studies suggest a preventive effect of oral Gln supplementation on the development of NASH; however, whether Gln also has therapeutic potential for pre-existing NASH has not yet been clarified. The aim of the present study was to determine whether Gln prevents the progression of diet-induced NASH in mice. For 8 wk, female C57BL/6J mice (6-8 wk old) were pair-fed a liquid Western-style diet [WSD, 25% of energy from fat, 50% wt:wt fructose, 0.16% wt:wt cholesterol] or control diet (C diet) to induce liver damage. From week 8 to 13, they were pair-fed the C diet or WSD alone or supplemented with l-Gln to provide 2.1 g/kg body weight (C diet + Gln or WSD + Gln). Energy intake was adjusted to the group with the lowest energy intake. Indexes of liver damage and inflammation, intestinal barrier function, and toll-like receptor 4 () signaling in the liver were determined. The liver histology scores significantly increased from 8 to 13 wk (+31%) in WSD-fed mice and were significantly higher than in controls ( ≤ 0.05 for both time comparisons), whereas scores did not differ between C diet-fed and WSD + Gln-fed mice after 13 wk of feeding. The occludin protein concentrations in the small intestinal tissue were similarly reduced in both WSD-fed groups when compared with controls [WSD compared with C diet (-53%) and C diet + Gln (-42%), ≤ 0.05; WSD + Gln compared with C diet + Gln (-34%), ≤ 0.05] after 13 wk, whereas the expression of myeloid differentiation primary response gene 88 mRNA and concentration of inducible nitric oxide synthase and 4-hydroxynonenal protein adducts were significantly higher only in livers of WSD-fed mice ( ≤ 0.05 for the WSD group compared with all other groups; WSD + Gln group compared with the C diet groups: NS). Taken together, our data suggest that oral Gln supplementation protects mice from the progression of pre-existing, WSD-induced NASH.

Oral Supplementation of Sodium Butyrate Attenuates the Progression of Non-Alcoholic Steatohepatitis

“…To induce early stages of NASH, mice were pair-fed a liquid fat-, fructose-and cholesterol-rich diet (FFC; 17.8 MJ/kg diet: 60E% from carbohydrates, 25E% from fat and 15E% from protein with 50% wt/wt fructose and 0.16% wt/wt cholesterol; Ssniff, Soest, Germany). Control animals were fed a standard liquid diet (C; 15.7 MJ/kg diet: 69E% from carbohydrates, 12E% from fat and 19E% from protein; Ssniff, Soest, Germany) as detailed previously [22]. For the pair-feeding of mice, liquid diet was administered in bottles with ball nipples.…”

“…To achieve equal caloric intakes, the liquid diet intake of mice in each group was assessed daily and mean caloric intake per group per day 3 of 19 was calculated. The amount of diet and calories in the different groups were then adjusted to the group with the lowest caloric intake at the next day, whereas the group with the lowest caloric intake was fed ad libitum (pair-feeding model) [22]. After 8 weeks of feeding, mice were assigned to the following groups (n = 8/group): C-fed, FFC-fed, C-fed mice receiving C supplemented with 0.6 g SoB/kg bw (C+SoB; Sigma-Aldrich, Steinheim, Germany) or FFC-fed mice receiving FFC supplemented with 0.6 g SoB/kg bw (FFC+SoB).…”

“…The study design is summarized in Figure 1. At week 11, mice were fasted for 6h followed by a glucose tolerance test (GTT) as detailed previously [22]. After 13 weeks, mice were anesthetized through the intraperitoneal injection of 100 mg ketamine + 16 mg xylazine/kg bw.…”

“…Liver and intestinal tissue were fixed in neutral-buffered formalin or snap frozen in liquid nitrogen. For comparison of the effects of the different diets on the liver and body weight found after 8 weeks, data from a mouse experiment [22] run in parallel in the mouse facility in which mice were pair-fed C or FFC, were included in the present study (Table 1, Figure 2). ball nipples.…”

“…Endotoxin levels were measured in portal plasma using a commercially available limulus amebocyte lysate assay (Charles River, France) as previously described [22]. Recovery rates were 90-124%.…”