Hyaluronic acid 35 normalizes TLR4 signaling in Kupffer cells from ethanol-fed rats via regulation of microRNA291b and its target Tollip

Abstract

TLR4 signaling in hepatic macrophages is increased after chronic ethanol feeding. Treatment of hepatic macrophages after chronic ethanol feeding with small-specific sized hyaluronic acid 35 (HA35) normalizes TLR4 signaling; however, the mechanisms for HA35 action are not completely understood. Here we used Next Generation Sequencing of microRNAs to identify negative regulators of TLR4 signaling reciprocally modulated by ethanol and HA35 in hepatic macrophages. Eleven microRNAs were up-regulated by ethanol; only 4 microRNAs, including miR291b, were decreased by HA35. Bioinformatics analysis identified Tollip, a negative regulator of TLR4, as a target of miR291b. Tollip expression was decreased in hepatic macrophages from ethanol-fed rats, but treatment with HA35 or transfection with a miR291b hairpin inhibitor restored Tollip expression and normalized TLR4-stimulated TNFα expression. In peripheral blood monocytes isolated from patients with alcoholic hepatitis, expression of TNFα mRNA was robustly increased in response to challenge with lipopolysaccharide. Importantly, pre-treatment with HA35 reduced TNFα expression by more than 50%. Taken together, we have identified miR291b as a critical miRNA up-regulated by ethanol. Normalization of the miR291b → Tollip pathway by HA35 ameliorated ethanol-induced sensitization of TLR4 signaling in macrophages/monocytes, suggesting that HA35 may be a novel therapeutic agent in the treatment of ALD.

Specifically Sized Hyaluronan (35 kDa) Prevents Ethanol‐Induced Disruption of Epithelial Tight Junctions Through a layilin‐Dependent Mechanism in Caco‐2 Cells

“…Intestinal permeability increases early in the progression of ALD and involves the disruption of epithelial tight junctions within the gastrointestinal tract (Elamin et al., ). Protection of tight junctions in the proximal colon of mice by treatment with HA35 was associated with protection from EtOH‐induced liver injury in mice (Saikia et al., ). Here, we have extended this analysis to determine the interactions between EtOH and HA35 on the tight junction proteins ZO‐1 and occludin in other intestinal regions.…”

“…HA35, a small specific‐sized hyaluronan species, has protective effects on intestinal homeostasis in both development and disease (Hill et al., ; Kim et al., ). Importantly, oral provision of HA35 during short‐term EtOH feeding to mice prevented EtOH‐induced loss of tight junction integrity in the proximal colon, reduced the concentration of endotoxin in the portal circulation, and protected from liver injury (Saikia et al., ). Here, we investigated the impact of HA35 preventing EtOH‐induced loss of colocalization of ZO‐1 and occludin along the entire gastrointestinal tract.…”

“…In animal models, EtOH feeding impairs the integrity of tight junctions. The colon is consistently targeted in response to both short‐term and chronic EtOH feeding (Chaudhry et al., ; Cresci et al., ; Saikia et al., ); however, the region(s) affected by EtOH likely depends on both the dose and duration of EtOH exposure (Patel et al., ). Importantly, strategies that restore gut integrity in the colon are associated with a protection from EtOH‐induced liver injury, including supplementation with glutamine and tributyrin (Chaudhry et al., ; Cresci et al., ).…”

“…Maltose dextrin was isocalorically substituted in place of EtOH for pair‐fed controls. During the last 3 days of the study, mice were treated with 15 mg/kg HA35 or an equivalent volume of saline by gavage at 1:30 pm (Saikia et al., ). The morning after the last HA35 treatment, EtOH‐ and pair‐fed mice were anesthetized and samples collected prior to euthanasia.…”

“…Recent data from our laboratory demonstrate that HA35 also protects mice from short‐term EtOH‐induced liver and gut injury; protection was associated with maintenance of tight junctions in the proximal colon during short‐term EtOH feeding (Saikia et al., ). However, it is not known whether HA35 protects multiple regions of the intestine and whether its therapeutic efficacy is due to direct and/or indirect interactions with the intestinal epithelium.…”