Retinoic acid and hydrocortisone strengthen the barrier function of human RPMI 2650 cells, a model for nasal epithelial permeability

Abstract

The nasal pathway represents an alternative route for non-invasive systemic administration of drugs. The main advantages of nasal drug delivery are the rapid onset of action, the avoidance of the first-pass metabolism in the liver and the easy applicability. In vitro cell culture systems offer an opportunity to model biological barriers. Our aim was to develop and characterize an in vitro model based on confluent layers of the human RPMI 2650 cell line. Retinoic acid, hydrocortisone and cyclic adenosine monophosphate, which influence cell attachment, growth and differentiation have been investigated on the barrier formation and function of the nasal epithelial cell layers. Real-time cell microelectronic sensing, a novel label-free technique was used for dynamic monitoring of cell growth and barrier properties of RPMI 2650 cells. Treatments enhanced the formation of adherens and tight intercellular junctions visualized by electron microscopy, the presence and localization of junctional proteins ZO-1 and b-catenin demonstrated by fluorescent immunohistochemistry, and the barrier function of nasal epithelial cell layers. The transepithelial resistance of the RPMI 2650 cell model reached 50 to 200 X 9 cm 2 , the permeability coefficient for 4.4 kDa FITC-dextran was 9.3 to 17 9 10 -6 cm/s, in agreement with values measured on nasal mucosa from in vivo and ex vivo experiments. Based on these results human RPMI 2650 cells seem to be a suitable nasal epithelial model to test different pharmaceutical excipients and various novel formulations, such as nanoparticles for toxicity and permeability.

Optimization of an oral mucosa in vitro model based on cell line TR146

“…EGF has shown to enhance differentiation of other epidermal barriers, 34,35 while HC is known for its promoting effects on the tightness of brain endothelial cells. [36][37][38] Addition of hydrocortisone alone at a physiological concentration to DMEM media (supplemented with 10% FCS, 1% P/S) improved the barrier upon airlift cultivation significantly in a concentration-dependent manner. 39,40 As further supplement, we tested retinoic acid (RA) in DMEM without serum as RA is often described to enhance the barrier.…”

Alpha-Melanocyte Stimulating Hormone Protects against Cytokine-Induced Barrier Damage in Caco-2 Intestinal Epithelial Monolayers

“…After the incubation samples from the upper and lower compartments were collected and the concentrations of the marker molecules were determined by a fluorescence multiwell plate reader (Fluostar Optima, BMG Labtechnologies, Germany; excitation wavelength: 485 nm, emission wavelength: 535 nm for fluorescein and excitation wavelength: 584 nm, emission wavelength: 680 nm for Evans-blue labeled albumin). The apparent permeability coefficients (P app ) were calculated by the following equation [24]. where [C] B is the concentration of the tracer in the basal (acceptor) compartment after 1 hour, [C] A is the concentration of the tracer in the apical (donor) compartment at 0 hour, V B is the volume of the basal compartment (530 μl) and A is the surface area available for permeability (0.33 cm 2 ).…”

Enhanced nasal drug delivery efficiency by increasing mechanical loading using hypergravity

“…RPMI 2650 cell line has shown consistent growth, and high stability in vitro without loss of normal diploid karyotype 32 , 33 . Further RPMI 2650 cells form a cellular barrier with junctional proteins, including ZO-1, OCLN, CLDN-1, CDH-1, and β-catenin 34 , 35 . They have been used as an in vitro model for nasal permeation studies in recent years, reporting that RPMI 2650 cells cultured at A-L conditions express sufficient barrier properties such as high TEER values and permeation coefficients 34 , 36 , 37 .…”