European Journal of Pharmaceutical Sciences 25 (2005) 113–121 Permeability of pig urinary bladder wall: the effect of chitosan and the role of calcium Mojca Kerec a , Marija Bogataj a,∗ , Peter Veraniˇ c b , Aleˇ s Mrhar a a FacultyofPharmacy,UniversityofLjubljana,Aˇ skerˇ ceva 7, Ljubljana 1000, Slovenia b InstituteofCellBiology,MedicalFaculty,Lipiˇ ceva 2, Ljubljana, Slovenia Received 17 September 2004; received in revised form 21 January 2005; accepted 7 February 2005 Available online 9 March 2005 Abstract Chitosan is a cationic polysaccharide widely employed as an absorption enhancer. The aim of this work was to examine the effect of chitosan on the permeability of isolated pig urinary bladder wall as well as to determine the role of calcium ions in this process. Besides permeability studies, scanning electron microscopy and fluorescent microscopy were applied to get an insight into the mechanism by which chitosan increases the permeability of urinary bladder wall. Additionally, the obtained findings were compared to the mechanism proposed for Caco-2 cells. The results show that 0.5% (w/v) chitosan increases the permeability of urinary bladder wall by causing the desquamation of the urothelium. Calcium ions, when applied to the luminal surface of the urinary bladder at the same time as chitosan, decreases the effect of chitosan on permeation of the model drug moxifloxacin into the bladder wall in concentration dependent way. The desquamation of urothelium cells caused by chitosan was reduced in the presence of calcium, but not to such extent as it would be expected from the permeability studies. When present, calcium obviously interferes directly in the interactions between chitosan and the surface of urothelium. © 2005 Elsevier B.V. All rights reserved. Keywords: Chitosan; Urinary bladder; Calcium; Permeability; Desquamation; Microscopy 1. Introduction Chitosan is a polysaccharide composed of glucosamine and N-acetyl glucosamine. It is obtained by the partial deacetylation of chitin. Different types of chitosan vary in the degree of deacetylation and viscosity. Chitosan is regarded as a biocompatible, biodegradable and non-toxic polymer. It also possesses bioadhesive properties and is known as an ab- sorption enhancer. All these properties make this polymer a promising agent in the development of controlled drug deliv- ery systems (Felt et al., 1998; Ravi Kumar, 2000; Singla and Chawla, 2001). The ability of chitosan to work as an absorption enhancer was proven on Caco-2 cells, which serve as a model of in- testinal epithelium (Artursson et al., 1994; Borchard et al., 1996; Dodane et al., 1999; Smith et al., 2004), as well as in ∗ Corresponding author. Tel.: +386 1 476 95 10; fax: +386 1 425 80 31. E-mailaddress: marija.bogataj@ffa.uni-lj.si (M. Bogataj). “in vitro” experiments on nasal, buccal, vaginal and urinary bladder mucosae of different animals (Natsume et al., 1999; S ¸ enel and Hincal, 2001; Grabnar et al., 2003; Sinswat and Tengamnuay, 2003; Sandri et al., 2004). Chitosan also in- creased the bioavailabilty of a peptide drug buserelin, which was intraduodenally applied in the “in vivo” experiments in rats (Lueßen et al., 1996). Chitosan decreases transepithelial electrical resistance (TEER) of Caco-2 cells in a dose-dependent way. However, the lowest concentrations of chitosan that still produced a significant decline in TEER differ a lot from one study to an- other (Ranaldi et al., 2002; Smith et al., 2004). The influence of chitosan on the permeability of Caco-2 cells is reported to be reversible. After 1-h exposure of Caco-2 cells to chi- tosan (from 0.00125 to 0.01%, w/v), TEER returned to the baseline values throughout the remaining 24 h (Dodane et al., 1999). Smith et al. (2004) reported that 48 h were necessary for TEER to reach 80% of the baseline value after the Caco- 2 cells were exposed to 0.5% chitosan for 1 h. On the other 0928-0987/$ – see front matter © 2005 Elsevier B.V. All rights reserved. doi:10.1016/j.ejps.2005.02.003