Enhanced porcine gut barrier functioning and reduced inflammation from a combination of seaweed bioactives

Abstract

Background: Microbial and toxin-related challenges are well-documented causes of impaired animal performance. The gastro-intestinal tract is the single largest organ that interfaces with numerous challenges including pathogenic organisms, toxins and other immune activating stimuli. The integrity of the intestinal mucosal barrier is the first and most critical line of defence. There are a number of reports demonstrating the potential of naturally derived feed additives to mitigate the damage derived from toxin or infectious agents on cell culture barrier integrity models but little information on target species tissue. Natural ingredients based on Lithothamnion glaciale (LG) and an extract of Ascophyllum nodosum (ANE) were tested in an ex vivo model using porcine ileal tissue, in the absence and presence of an infectious challenge derived from Salmonella enterica enteriditis (SEE). Read-outs included various parameters on the barrier integrity, tissue histology as well as the immune status of the tissue.

Results: The SEE challenge significantly impaired barrier integrity (P<0.05), as demonstrated by increased paracellular ([3H]-mannitol) relative to transcellular ([14C]-caffeine) transport. LG and ANE were tested individually at multiple doses and in combination to explore potential synergistic effects. The barrier integrity was positively impacted from the combination of LG and ANE at the low ANE dose, especially following the SEE challenge (P<0.001). The SEE challenge reduced TNF-α expression in the control treatment, which is caused by the downregulation of the inflammatory response immediately after a challenge (P>0.01). The relative expression of the gut barrier protein Cadherin-17 was increased when LG and/or ANE was included (P<0.001), both with and without the SEE challenge. The inflammatory markers tumour necrosis factor-α, Caspase-1, Interleukin-22 and regenerating islet-derived protein-3 γ were affected by the inclusion of LG and/or ANE (P<0.05). A synergistic effect between the two marine bioactives was evident and appears to be dose dependent, with the low dose rate of ANE and the low and medium LG rate being most optimal.

Conclusions: These results suggest that an optimal combination of marine bioactives can have a significant effect in enhancing gut barrier integrity and immune reactivity when challenged with an intestinal pathogenic bacteria in porcine ileal tissue.

Keywords: Ascophyllum nodosum; Lithothamnion sp.; Gut barrier; Gut integrity; Marine bioactives; Piglet health; Salmonella infection.

Fig. 1

Apparent permeability (Papp) of mannitol transport for time intervals 0–120, 120–240, and 240–300 min for 3 different control groups. Presented are the least square mean values with their standard errors, 4 replicates per treatment

Fig. 2

LDH leakage (%) into apical and basolateral compartment after 300 min incubation as cumulative values. Unchallenged: not exposed to Salmonella enterica enteriditis challenge; Challenged: exposed to Salmonella enterica enteriditis challenge; LG: Lithothamnion glaciale; ANE: extract. Presented are the least square mean values with their standard errors, 4 replicates per treatment. Effects of treatment (P<0.001) and the interaction between condition x treatment (P<0.05) were observed

Fig. 3

FD4 permeability for time intervals 0–60, 60–120, 120–240, and 240–300 minutes. A control treatment groups; B single components LG and ANE; C combined components LG + ANE without and with Salmonella enterica enteriditis challenge. LG: Lithothamnion glaciale; ANE: Ascophyllum Nodosum extract. Presented are the least square mean values with their standard errors, 4 replicates per treatment

Fig. 4

Apparent permeability P_app of mannitol transport for the combined LG and ANE treatments without and with SEE challenge at 240 minutes. A Treatment groups in unchallenged conditions; B Treatment groups in challenged conditions. LG: Lithothamnion glaciale, ANE Ascophyllum Nodosum extract, SEE Salmonella enterica enteriditis. Presented are the least square mean values with their standard errors, 4 replicates per treatment

Fig. 5

Regression analysis for LG and ANE treatment groups apparent permeability (P_app) without and with SEE challenge. A Contour plot; B Interaction profiles. Predicted RMSE for the linear regression: 3.1626; R² = 0.45; P-value < 0.01. LG: Lithothamnion glaciale; ANE: Ascophyllum Nodosum extract; SEE: Salmonella enterica enteriditis

Fig. 6

Histological analysis of tissue following inTESTineTM incubation. A Blanc at 0 hr; B Control at 5 hrs; C LG 63.2/ANE 2.3 treatment at 5 hrs; D Control + SEE challenge at 5 hrs; E LG 63.2/ANE 2.3 + SEE challenge at 5 hrs. SEE: Salmonella enterica enteriditis; LG: Lithothamnion glaciale; ANE: Ascophyllum Nodosum extract