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Home » Archive » 2014

TDK conference 2014

The effect of selective matriptase inhibitors on paracellular permeability
Barna Réka Fanni - year 5
SZIU Faculty of Veterinary Science, Department of Pharmacology and Toxicology
Supervisor: Erzsébet Pásztiné Gere

Abstract:

Matriptase is a type II transmembrane serine protease, and it is expressed in non-transformed and cancerous epithel cells. Some of the selective matriptase inhibitors have been found to be efficient drug candidates in the treatment of osteoarthritis, human influenza, and tumor metastases. In this study we used 3- aminidophenylalanine- structure based inhibitors to investigate putative changes in the integrity of IPEC-J2 monolayer and to examine role of matriptase in modulation of paracellular permeability.

Our aim was to establish a cell modell suitable for tracing transport processes between the apical and basolateral compartments mimicking physiological processes in small intestine. We used non- tumorgenic, porcine jejunal cell monolayer, IPEC-J2 cells cultured on membrane insert. IPEC-J2 cells were treated with non- selective, serine proetase inhibitor 4-(2-aminoethyl)-benzosulphonylfluorid (AEBSF ), and selective 3- amidinophenylalanine-structure based matriptase/TMPRSS2 inhibitors. Significant decrease was found in transepithelial electrical resistance (TER) between apical and basolateral compartments of treated IPEC-J2 cell monolayer . The rate and reversibility of decrease in TER was dependent on the inhibitor concentration and the cell differentiaiton. Based on findings from apico-basolateral transport of fluorescein isothiocyanate- labelled dextran 4 kDa (FD4) significant increase in paracellular permeability of cell monolayer exposed to matriptase inhibition was confirmed as a result of barrier dysfunction. Neutral red uptake assay proved that cell viability did not alter in cells treated with matriptase inhibitors compared to that of control groups. Hydrogen- peroxide level analyzed with Amplex red assay did not indicate oxidative stress during and after application of selective matriptase inhibitors. Immunofluorescent staining revealed changes in the distribution pattern of occludin in the dichlorobiphenyl- sulfonamidobenzimidamide - treated cells as a consequence of the effect of matriptase inhibitor on structural alterations of tight junction protein assembly.

In conclusion, it was found that the matriptase inhibitors decreased TER and simultaneously increased the apico-basolateral transport of FD4, with relocalization of occludin from cell membran to cytoplasm. Based on these results matriptase could play significant role in regulation of the paracellular permeability by modulating tight junction assembly. In the future, the in-depth understanding of matriptase effect may contribute to establishment of new therapeutical options in the treatment of Crohn-disease, colitis ulcerosa or to invent defensive strategies in veterinary practice to restore barrier dysfunction in infectious diseases in livestock.



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