CHITOSANS AS ABSORPTION ENHANCERS FOR POORLY ABSORBABLE DRUGS .2. MECHANISM OF ABSORPTION ENHANCEMENT

Purpose. It has recently been shown that the absorption enhancing and toxic effects of chitosans are dependent on their chemical composition . In this study, the mechanisms underlying these effects were investig ated at the cellular level. Methods, The effects on epithelial cells o f chitosans with different chemical composition, absorption enhancing properties and toxicities were studied in Caco-2 monolayers. Chitosan C(1:31) has a low degree of acetylation (DA) (1%) and a low m.w. (31 k D), and displays dose-dependent absorption enhancement and cytotoxicit y; chitosan C(35:170) has a higher DA (35%) and a higher m.w. (170 kD) , is less dose-dependent in absorption enhancement, and is not cytotox ic. A third non-toxic chitosan C(49:22) with a high DA (49%), a low m. w. (22 kD), and no influence on epithelial permeability was used as co ntrol. Results, C(1:31)and C(35:170) bound tightly to the epithelium. Cellular uptake of the chitosans was not observed. Both chitosans incr eased apical but not basolateral cell membrane permeability and induce d a redistribution of cytoskeletal F-actin and the tight junction prot ein ZO-1. This resulted in increased paracellular permeability of hydr ophilic marker molecules of different molecular weights. Addition of n egatively charged heparin inhibited the cellular and the absorption en hancing effects of the chitosans, indicating that these effects are me diated via their positive charges. The onset of the effects of C(35:17 0) on apical membrane permeability and tight junction structure was mu ch faster than that of C(1:31). C(49:22) did not influence any of the properties of the Caco-2 cell monolayers studied. Conclusions, The bin ding and absorption enhancing effects of chitosans on epithelial cells are mediated through their positive charges. The interaction of chito sans with the cell membrane results in a structural reorganisation of tight junction-associated proteins which is followed by enhanced trans port through the paracellular pathway.