Caco paracellular pathway

Figure 8 Appearance kinetics of radiolabeled solutes that diffuse across Caco-2 cell monolayers via the paracellular pathway. The Transwell system consisted of a donor and receiver solution at pH 7.4. Stirring by planar rotation up to 100 rpm had no effect. The insert with filter, cell monolayer, and donor were transferred to a new receiver chamber at time intervals to maintain sink conditions.

Table 5 Permeability Coefficients of the Paracellular Pathway and Estimation of the Effective Pore Radius and Molecular Restriction Factor for the Caco-2 Cell Monolayer... 

Pade, V., Stavchansky, S., Estimation of the relative contribution of the transcellular and paracellular pathway to the transport of passively absorbed drugs in the Caco-2 cell culture model, Pharm. Res. 1997, 34, 1210-1215. 

The rat intestinal cell line IEC-18 has been evaluated as a model to study small intestinal epithelial permeability. This cell line forms very leaky monolayers with TER of 50 n cm2 and permeability to mannitol of 8 x 10-6 cm s 1. The IEC-18 model was proposed to be a better model than the Caco-2 monolayers for evaluating the small intestinal paracellular permeation of hydrophilic molecules. However, the leakier paracellular pathway is related to the poor differentiation level of the cells and an undeveloped paracellular barrier lacking peri-junctional actin-belt. In addition, due to the poor differentiation the cells have minute expression of transporters and are therefore not useful for studies of carrier-mediated transport [82, 84]... 

Another limitation of the Caco-2 monolayers is their colonic origin and tight paracellular pathway, which tend to lead to underestimations in permeability to paracellularly transported compounds [97]. This is likely to be correct for small compounds (MW < 150) - i.e., compounds smaller than normal drugs - but it remains to be seen to what extent the Caco-2 model gives false-negative predictions of the fraction absorbed for polar drugs of normal size in humans where para-... 

The paracellular pathway, between the epithelial cells, is both size- (MW, volume) and charge-dependent [60, 109, 110]. In general, compounds that are limited to paracellular transport are not efficiently absorbed due to the small available absorptive area and the restriction by tight junctions. The molecular weight cut-off seems to be around 400 g mol-1 and 300 g mol-1 for the small and large intestine respectively, and 300 g mol-1 for the Caco-2 cell monolayers [60], which shows the more colonic nature of the Caco-2 monolayer model. Compounds with a... 

Usually, PAMPA does not have any aqueous pores and is therefore not suitable for examining paracellular transport. Some cell models, for example, Caco-2 and MDCK, have a narrower tight junction than the in vivo human intestine and may underestimate paracellular transport. However, the contribution of the paracellular pathway can be added using an in silico approach [76-78]. 

The effect of size and charge on the passive diffusion of peptides across Caco-2 cell monolayers via paracellular pathways as well as on retention time has been studied using the IAM technique [38]. It was found that a positive net charge of the hydrophilic peptides favors permeation across the intestinal mucosa via the paracellular pathway. With an increase in size, the molecular sieving of the epithelial membrane dominates the transport of peptides and the influence of the net charge becomes less important. The permeability coefficient, Papp, of the capped amino acids and the model peptides can be calculated by ... 

Prognosis of a compounds permeability should be made stressing limitations of the model. There is no bioavailability prognosis from in vitro data - a cellular assay can provide only permeability potential through a biological membrane. The membrane, in most cases CACO-2 cells, is very similar to what we observe in vivo in the small intestine and resembles many characteristics to in vivo enterocytes. CACO-2 cells can be used for prediction of different pathways across intestinal cells. Best correlation occurs for passive transcellular route of diffusion. Passive paracellular pathway is less permeable in CACO-2 and correlations are rather qualitative than quantitative for that pathway. CACO-2 cells are an accepted model for identification of compounds with permeability problems, for ranking of compounds and selection of best compounds within a series. Carrier-mediated transport can be studied as well using careful characterization of transporters in the cell batch or clone as a prerequisite for transporter studies. 

On the other hand, artificial membranes represent a further simplification of the human GIT than Caco-2 monolayers as a number of features are not present, like the paracellular pathway, active transporters, or efflux systems. Caco-2 is a useful model to study actively transported compounds as well as substrates and inhibitors of efflux systems (PgP, MRP). Caco-2 monolayers also have some potential to model paracellular transport, although the junctions appear tighter than in the upper GIT. The consequence is that some paracellular transported compounds are properly transported while others are underestimated (Figure 15.6R). 

Kim M, Kometani T, Okada S, Shimuzu M. Permeation of hesperidin glycosides across Caco-2 monolayers via the paracellular pathway. Biosci Biotech Biochem 1999 63 2183-2188. 

Mukaizawa, F., et al., 2009. Novel oral absorption system containing polyamines and bile salts enhances drug transport via both transcellular and paracellular pathways across Caco-2 cell monolayers. hiL J. Pharm. 367 (1-2), 103-108. http //www.ncbi.nlm.nih.gov/ pubmed/18929635 (accessed 10.01.16.). 

Major transport pathways in Caco-2 monolayers. A Passive transcellular B Passive paracellular C Transporter-mediated apical uptake D Transporter-mediated apical efflux E Transporter-mediated basolateral efflux F Transporter-mediated basolateral uptake. 

The paracellular permeation pathway in the intestinal cell monolayer models is often limited. Therefore these models are not suitable for predicting permeability of paracellularly absorbed compounds. The average pore radius in Caco-2 cells (<6 A) is more representative of the colon than the small intestine (8-13 A)and paracellular transport can be up to 100-fold lower in Caco-2 cells than in the small intestine. Investigation of a rat intestinal cell line 2/4/Al, which forms polarized cell mono-layers and has an average pore radius (9 A) more representative of the small intestine, showed improved prediction of oral absorption for incompletely absorbed drugs [24, 25]. 

---