Fat Pad Accumulation and Diet Purification Alter Gastrointestinal Integrity Markers in C57Bl/6 Mice
Authors: Price Tara R. , Moncada Kristin , Nolasco Liliana , Walzem Rosemary 
Published: November 2023

Diet composition and increased adiposity are often implicated as either causative or explanatory mechanisms leading to altered gastrointestinal (GI) function including changes in permeability. GI permeability (GI‐P) is thought to be a key factor in systemic inflammation due to microbial inflammogen passage into the systemic circulation. The intestine's ability to exclude FITC‐dextran as well as measurement of lipopolysaccharide (LPS) in serum or plasma are frequently used indicators of gut permeability. Nutritionally adequate, closed‐formula non‐purified “chow‐type” diet is often thought to provide a “healthy‐diet” comparison to purified diets in rodent studies. We recently compared FITC‐dextran and LPS outcomes from age‐matched male C57Bl/6 mice fed either purified diets providing 20% EN PRO, 30% EN fat, and 50% EN CHO or chow. Unexpectedly FITC determined GI‐P of mice fed purified diets containing either isolated soy protein (ISP) or dried whole milk powder (DWMP) was some 68‐fold lower (p<0.02) than that of chow fed mice at 4 hours following a 600 mg 4 kDa MW FITC‐dextran/kg BW oral dose. Similarly, plasma endotoxin was 3‐fold higher in chow‐fed mice compared to ISP‐ or DWMP‐fed animals (p<0.000). To determine if these differences would persist if mice from the same age cohort (i.e. all born in the same week) were fed purified and non‐purified versions of soy and dairy based diets differing in PRO and CHO complexity. Each diet provided 20%EN PRO, 30% EN fat, and 50% EN CHO. Purified diets contained 14% neutral detergent fiber (NDF) as cellulose, with cornstarch:maltodextrin (1:1.8) and sucrose (soy) or native milk sugars (dairy) providing remainder of CHO. Non‐purified soy and dairy diets provided 11.4% and 8.4% NDF, respectively, from wheat mids, corn, corn gluten meal and soybean meal (soy diet). Weanling C57Bl/6 mice (n=15/diet) were fed either purified diets containing isolated soy protein (ISP) or dried whole milk powder (DWMP) as the protein source, or non‐purified diets containing soybean meal (SPC), or dried whole milk powder (DMC) as protein sources in a 14 week feeding trial started after a 2‐week acclimation period in which all were fed the facility's standard chow. At the study's end, ISP‐fed mice were heaviest (p<0.000) and had gained the most body weight (p<0.000). Feed intake was lowest in DMC, being different from DWMP and SPC (p<0.04). Overall, purified diets slowed gut motility (6.6 hrs) compared to non‐purified diets (4.1 hrs, p<0.0001). Fat pads (epidydimal, mesenteric, retroperitoneal and flank subcutaneous) were heaviest in ISP‐fed mice (p<0.002), being 93% heavier than SPC. GI‐P was assessed by FITC‐dextran and was greatest in ISP‐fed mice (p<0.008). Increasing FITC‐dextran permeability was positively correlated with increasing fat mass percentage independent of diet source (p<0.02, FITC = 0.00714 +0.0488* fat pad g/100g BW). Mice fed either dairy protein containing diet had numerically lower FITC‐dextran GI‐P in both purified (10.2 ng/mL v. 13.6 ng/mL, p <0.11) and non‐purified diet (9ng/mL v.10.2ng/mL, p <0.000) settings. Results suggest that age‐matching is not sufficient to ensure comparable FITC‐dextran GI‐P assessments in mice fed different diets. In temporally contiguous feeding, adiposity appears to be a key factor in apparent GI permeability; less refined diets reduce gut permeability. Outcomes from soy‐containing diets particularly suggest that protein and carbohydrate refinement impact weight gain and GI permeability.