TRANSPORT OF PROTEOLYTIC-ENZYMES ACROSS CACO-2 CELL MONOLAYERS

Purpose. To investigate the mechanisms by which proteolytic enzymes, s uch as trypsin, chymotrypsin, papain, and bromelain, are able to cross the intestinal mucosal barrier after oral administration to man. Meth ods. Filter-grown Caco-2 cell monolayers were incubated with proteolyt ic enzymes and then the transepithelial electrical resistance (TEER) a nd the transport of the paracellular marker fluorescein were monitored . The effects of the enzymes on the cells were investigated by light m icroscopy and by biochemical assays. Transport of intact proteases acr oss the cells was verified by monitoring the proteolytic activity and MALDI-TOF mass spectroscopic identification of undegraded trypsin. Res ults. Depending on time, concentration, and side of exposure to Caco-2 cell monolayers, all proteases decreased the TEER and increased the t ransport of fluorescein. Some morphological and metabolic changes were observed. The effects were reversible, but until 24 hours after remov al of the proteases. Under the conditions of this in-vitro model, appr oximately 10% of the apically applied dose reached the basolateral com partment as biologically active, non-degraded molecules. Conclusions. Proteolytic enzymes were found to exert considerable effects on the ba rrier function of Caco-2 monolayers, facilitating the transport of nor mally non-absorbable compounds. This suggests the also reported, but s o far unexplained, systemic absorption of proteolytic enzymes after or al administration in vivo may occur by self-enhanced paracellular tran sport.