Altered Na⁺/K⁺-ATPase expression plays a role in rumen epithelium adaptation in sheep fed hay ad libitum or a mixed hay/concentrate diet

Abstract

ABSTRACT:In this study we investigated rumen papillae morphology and the localization and expression of the Na + /K + -ATPase in eight sheep fed hay ad libitum (h) or hay ad libitum plus additional concentrate (h/c). Four sheep were provided with the ad libitum h-diet for the complete three-week experimental period. The second group of four sheep received the h-diet for only one week and was fed the mixed hay/concentrate (h/c) diet for another two weeks. The amount of concentrate supplement was stepwise increased from 150 to 1000 g/day and given in two meals. Following slaughter rumen papillae from the atrium ruminis (AR), the rumen ventralis (RV) and the ventral blind sac (BSV) were fixed and examined for morphological changes and Na + /K + -ATPase localization by morphometric methods and immunohistochemistry. Ruminal epithelial cells (REC) originating from the strata basale to granulosum were also isolated. Cellular Na + /K + -ATPase expression (mRNA and protein) and differentiation state were determined by RT-PCR, Western blot, and flow cytometry. Compared with data from h-fed sheep, morphometric analysis revealed an increased length and width of rumen papillae in h/c-fed sheep, resulting in a marked 41% and 62% increase in rumen papillae surface in AR and RV, respectively. The rumen mucosa of h/c-fed sheep was characterized by a predominant stratum corneum (42 ± 0.7 µm vs. 28 ± 0.5 µm), but the thickness of the metabolically active cell layers remained unchanged. REC suspensions from sheep fed the h/c diet generally contained more cells (7.30 ± 0.83 vs. 3.49 ± 0.52 × 10 7 /ml; P < 0.001) and an increased proportion of REC positive for basal cytokeratin and for the differentiation marker cytokeratin 10 (P < 0.05). Cellular (cell membrane) and epithelial (stratum basale to stratum granulosum) Na + /K + -ATPase localization was similar between rumen regions and was not changed by concentrate feeding. After two weeks on the h/c-diet, a 96% increase in the absolute number of Na + /K + -ATPasepositive REC (6.56 ± 0.84 vs. 3.35 ± 0.51 × 10 7 /ml; P = 0.003) and a 61% elevation (P = 0.043) in Na + /K + -ATPase protein expression in REC from the upper third of the suprabasal cell layers were found. Moreover, a two-fold (P = 0.001) elevation in cell membrane surface area accompanied by a reduction (1.19 × 10

Epithelial response to high-grain diets involves alteration in nutrient transporters and Na+/K+-ATPase mRNA expression in rumen and colon of goats1

“…Interestingly, dietary grain level did not modify the ATP1A1 expression in RE which is in contrast to recent observations in sheep (Kuzinski et al, 2011), where ATP1A1 expression in RE was upregulated with a hayconcentrate feeding compared with hay-feeding alone. Present correlations among ATP1A1 expression and ruminal pH and VFA concentration might indicate a positive relationship among ruminal fermentation and ATP1A1 expression in RE.…”

Modulation of vH+-ATPase is part of the functional adaptation of sheep rumen epithelium to high-energy diet

“…A downregulation of Na ϩ /K ϩ -ATPase-binding sites (25) can, thus, explain the reduced amiloride-sensitive component of H ϩ secretion seen in h/c-fed sheep. This is supported by results from our recent study (27), showing that concentrate feeding induced a decrease of the Na ϩ /K ϩ -ATPase protein amount per cm 2 cell membrane surface area, a decrease that equals the reduction of the amiloride effect seen after h/c feeding in this study (Ϫ31% and Ϫ33%). In the concentration used, foliomycin mainly affects vH ϩ -ATPases localized in the cell membrane (21) and thus, we hypothesize that the proportion of vH ϩ -ATPase localized in the cytosol or in intracellular organelles is increased in REC from h/c-fed compared with h-fed sheep.…”

The mRNA and protein expression of ruminal MCT1 is increased by feeding a mixed hay/concentrate diet compared with hay &lt;i&gt;ad libitum&lt;/i&gt; diet (Short Communication)

“…Functionally, these changes are reflected by an increased butyrate uptake. In the present study, the ruminal butyrate concentration was 6.8±0.4 mmol/l in h-fed sheep and increased to 17.3±1.2 mmol/l (P<0.001) after feeding the h/c diet (Kuzinski et al 2011). Thus, elevated butyrate concentrations could be a trigger for the upregulation of MCT1 mRNA (1.8-and 2.2-fold in G2 and G3, respectively) and protein expression (1.3-fold) observed in h/c-fed sheep.…”

“…The experimental design and methods have been described elsewere in detail (Kuzinski et al 2011). In preparation to the experiment, 8 male castrated lambs were fed the h diet for one week.…”