An experiment was conducted to uncover the effects of increasing dietary grain levels on expression of thiamine transporters in ruminal epithelium, and to assess the protective effects of thiamine against high-grain-induced inflammation in dairy cows. Six rumen-fistulated, lactating Holstein dairy cows (627 ± 16.9 kg of body weight, 180 ± 6 d in milk; mean ± standard deviation) were randomly assigned to a replicated 3 × 3 Latin square design trial. Three treatments were control (20% dietary starch, dry matter basis), high-grain diet (HG, 33.2% dietary starch, DM basis), and HG diet supplemented with 180 mg of thiamine/kg of dry matter intake. On d 19 and 20 of each period, milk performance was measured. On d 21, ruminal pH, endotoxic lipopolysaccharide (LPS), and thiamine contents in rumen and blood, and plasma inflammatory cytokines were detected; a rumen papillae biopsy was taken on d 21 to determine the gene and protein expression of toll-like receptor 4 (TLR4) signaling pathways. The HG diet decreased ruminal pH (5.93 vs. 6.49), increased milk yield from 17.9 to 20.2 kg/d, and lowered milk fat and protein from 4.28 to 3.83%, and from 3.38 to 3.11%, respectively. The HG feeding reduced thiamine content in rumen (2.89 vs. 8.97 μg/L) and blood (11.66 vs. 17.63 μg/L), and the relative expression value of thiamine transporter-2 (0.37-fold) and mitochondrial thiamine pyrophosphate transporter (0.33-fold) was downregulated by HG feeding. The HG-fed cows exhibited higher endotoxin LPS in rumen fluid (134,380 vs. 11,815 endotoxin units/mL), and higher plasma concentrations of lipopolysaccharide binding protein and pro-inflammatory cytokines when compared with the control group. The gene and protein expression of tumor necrosis factor α (TNFα), IL1B, and IL6 in rumen epithelium increased when cows were fed the HG diet, indicating that local inflammation occurred. The depressions in ruminal pH, milk fat, and protein of HG-fed cows were reversed by thiamine supplementation. Thiamine supplementation increased thiamine contents in rumen and blood, and also upregulated the relative expression of thiamine transporters compared with the HG group. Thiamine supplementation decreased ruminal LPS (49,361 vs. 134,380 endotoxin units/mL) and attenuated the HG-induced inflammation response as indicated by a reduction in plasma IL6, and decreasing gene and protein expression of pro-inflammatory cytokines in rumen epithelium. Western bottling analysis showed that thiamine suppressed the protein expression of TLR4 and the phosphorylation of nuclear factor kappa B (NFκB) unit p65. In conclusion, HG feeding inhibits thiamine transporter expression in ruminal epithelium. Thiamine could attenuate the epithelial inflammation during high-grain feeding, and the protective effects may be due to its ability to suppress TLR4-mediated NFκB signaling pathways.
