(171f) Ex-Vivo Analysis of Lipid Impact On Intestinal Mucosa Barrier

Ex-vivo Analysis of Lipid
Impact on Intestinal Mucosa Barrier

Hasan Yildiz, yildiz.h@husky.neu.edu

Rebecca L. Carrier,
rebecca@coe.neu.edu

Introduction:

A heterogeneous mucus layer has
evolved to protect exposed epithelial surfaces by trapping environmental
microorganisms while allowing the passage of certain compounds (e.g., nutrients)
across the epithelium. Our previous results indicate that exogenous lipids that
may be associated with food or drug delivery systems significantly impact
intestinal mucus barrier properties. However, these studies were conducted
using collected (scraped) porcine intestinal mucus, and recent literature
indicates that intestinal mucus is not isotropic. The main objective of this work
is to analyze the impact of lipids on particle transport across intact mucus on
an intestinal explant.

Materials
and Methods:

Transport of model drug
carriers was investigated using 3 week old wild type FVB/N mouse small
intestine. Mice were euthanized via CO₂. Intestine fragments were
dissected, cut open to expose the intestinal lumen, and placed into a chamber
on a concavity slide. 200 nm carboxylate- modified polystyrene microspheres
were diluted in maleate buffer or simulated
fed state intestinal contents (?FED state?) including maleate buffer, bile
salts, phospholipids (lecithin) and a lipid mixture comprised of 35 mM
soybean oil, 30 mM sodium oleate
and 15 mM monoglycerol for a final particle concentration of 0.0025 wt.-%. 5 µl of diluted particle solutions were then added on
top of tissue segments and the system was equilibrated for 1 h at 25 ⁰C
prior to microscopy in a humid chamber. Particle
diffusion was measured by tracking the positions of diluted microspheres using
real- time multiple particle tracking technique (MPT). Positions of particle
centroids were used to calculate time-averaged mean squared displacements (MSD)
and effective diffusivities (D_eff): MSD =
[x(t+τ)-x(t)]² + [y(t+τ)-y(t)]² and D_eff
= MSD/(4τ) where x and y are positional data and τ is the time scale.

Results:

Intestinal
contents characteristic of the fed state, including model bile and lipids,
markedly reduced particle transport rate through intact ex vivo intestinal mucus.
At a time scale of 10
s, transport rates of 200 nm carboxylate-modified particles in FED State were
nearly 5-fold lower than those of same-size particles in maleate buffer. Intermolecular
interactions, potentially including lipid-particle and/or lipid-mucus
interactions, possibly retard particle diffusion in the gastrointestinal tract.
Furthermore, effective diffusivities in maleate buffer in scraped mucus and
intestinal explant were on the same order, whereas in Fed State diffusivities
in explant tissue are an order of magnitude higher than collected porcine
intestine mucus.

Conclusions:

Food-associated lipids strengthen the intestinal
mucus barrier to particulate matter. These results motivate in vivo in-depth
investigation of the impact of lipids on mucosal barriers to facilitate
lipid-based delivery of drugs and potentially enable control of pathogen
invasion in disease.