Intestinal permeability induced by lipopolysaccharide and measured by lactulose, rhamnose and mannitol sugars in chickens

Abstract

Increased intestinal permeability (IP) can lead to compromised health. Limited in vivo IP research has been conducted in chickens.The objectives of the current study were to develop a model of increased IP utilizing lipopolysaccharide (LPS Escherichia coli O55: B5) and to evaluate IP changes using the lactulose, mannitol and rhamnose (LMR) sugar permeability test. In addition, fluorescein isothiocyanate dextran (FITC-d), D-lactate, zonula occludens (ZO-1) and diamine oxidase (DAO) permeability tests were employed. Male Ross chickens were reared until day 14 on the floor in an animal care facility and then transferred to individual cages in three separate experiments. In each of experiments 1 and 2, 36 chicks were randomly allocated to receive either saline (control) or LPS (n = 18/group). Lactulose, mannitol and rhamnose sugar concentration in blood was measured at 0, 30, 60, 90, 120 and 180 min in experiment 1, at 60, 90 and 120 min in experiment 2 and at 90 min in experiment 3 (n = 16/group). Lipopolysaccharide was injected intraperitoneally at doses of 0.5, 1 and 1 mg/kg BW in experiments 1, 2 and 3, respectively, on days 16, 18 and 20, whereas control received sterile saline. On day 21, only birds in experiments 1 and 2 were fasted for 19.5 h. Chicks were orally gavaged with the LMR sugars (0.25 gL, 0.05 gM, 0.05 gR/bird) followed by blood collection (from the brachial vein) as per time point for each experiment. Only in experiment 3, were birds given an additional oral gavage of FITC-d (2.2 mg/ml per bird) 60 min after the first gavage. Plasma D-lactate, ZO-1 and DAO concentrations were also determined by ELISA in experiment 3 (n = 10). Administration of LPS did not affect IP as measured by the LMR sugar test compared with control. This was also confirmed by FITC-d and DAO levels in experiment 3 ( P > 0.05). The plasma levels of D-lactate were decreased ( P < 0.05). Plasma levels of ZO-1 were increased in the third experiment only and did not change in the first two experiments. Lipopolysaccharide at doses of 0.5 and 1 mg/kg did not increase IP in this model system. In conclusion, the LMR sugar can be detected in blood 90 min after the oral gavage. Further studies are needed for the applicability of LMR sugars tests.Keywords: lactulose, rhamnose, mannitol, lipopolysaccharide, FITC-d ImplicationsIncreased intestinal permeability (IP) can lead to health disorders and reduced feed efficiency in chickens. We aimed to develop a model of increased IP by administering gramnegative bacterial toxin lipopolysaccharide (LPS) to chickens. This model could then be applied in an attempt to develop a new lactulose, mannitol and rhamnose (LMR) sugar permeability method to determine IP. An established model of increased IP and novel biomarkers to monitor IP changes could be applied to the study of nutritional interventions aimed at improving the health and welfare of chickens.However, further research is required to determine the optimal LPS dose.

Effects of delayed feeding, sodium butyrate and glutamine on intestinal permeability in newly-hatched broiler chickens

“…Blood was collected from the jugular vein 150 min after gavage. Standards and serum samples were prepared and analysed in triplicate as previously described (Gilani et al 2016b). Standards were spiked with FITC-d at 0, 0.0001, 0.001, 0.01, 0.1, 1.0 and 10 µg/ml to obtain a standard curve utilizing a Synergy MX plate reader (Biotek Instruments, Bedfordshire, UK) at the excitation and emission wavelengths of 485 and 530 nm, respectively.…”

“…Standards were spiked with FITC-d at 0, 0.0001, 0.001, 0.01, 0.1, 1.0 and 10 µg/ml to obtain a standard curve utilizing a Synergy MX plate reader (Biotek Instruments, Bedfordshire, UK) at the excitation and emission wavelengths of 485 and 530 nm, respectively. Chicken specific antibodies of diamine oxidase and D-lactic acid Enzyme Linked Immunosorbent Assays (ELISA) were obtained from MyBioSource, (San Diego, USA).The procedure was performed as described previously (Gilani et al 2016b). Standards and plasma samples were measured in duplicate in a microplate reader at 450 nm wavelength (Bio-Rad laboratories, California, USA) following manufacturer's instructions.…”

Reduced fasting periods increase intestinal permeability in chickens

“…Standards and samples were prepared as previously described by Gilani et al. (); Gilani et al () with some modifications. Lactulose (1.28 mg), mannitol (0.68 mg) and rhamnose (0.68 mg) were each mixed with 30 ml fasted human plasma, to prepare standard solutions with concentrations of 125, 62.5 and 31.25 μ m for each sugar.…”

“…Standards and samples were prepared as previously described by Gilani et al. (); Gilani et al (). Briefly, standards (0, 0.0001, 0.001, 0.01, 0.1, 1.0 and 10 μg/ml) were prepared using the same FITC‐d as for the oral gavage.…”

Biomarkers for monitoring intestinal health in poultry: present status and future perspectives

“…In man, the clinical form of chronic, non-pathogen-induced intestinal inflammation, classified under the common denominator of inflammatory bowel disease (IBD), has been associated consistently with an acute phase response in the liver, resulting in a significant increase in acute phase proteins (especially C-reactive protein and lipopolysaccharide-binding protein (LBP)) in serum [ 36 , 37 ]. In the chicken three different experimental models have been used to increase the permeability of the intestinal mucosal barrier, but the effect on the intestinal barrier was insufficient to observe any changes in serum acute phase proteins [ 38 , 39 ]. Moreover, an acute phase response in the liver can also be seen in response to any inflammatory process, also to one taking place in other parts of the body, outside the intestinal tract, so one can doubt about the specificity of serum acute phase proteins as markers of (poor) intestinal health [ 40 ].…”