Paracellular pathway

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 

Transport of the compound AZ001 and markers for paracellular and transcellular pathways in the presence and absence of EDTA 

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]. 

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Y-L He, S Murby, G Warhurst, L Gifford, D Walker, J Ayrton, R Eastmond, M Rowland. Species differences in size discrimination in the paracellular pathway reflected by oral bioavailability of poly (ethylene glycol) and D-peptides. J Pharm Sci 87 626-633, 1998. 

Figure 5 Parallel transcellular and paracellular pathways through intestinal epithelial cell monolayer.

With respect to the size and charge selectivity of paracellular pathways, equivalent pore theory has been utilized to calculate an effective radius based on the membrane transport of uncharged hydrophilic molecules, while equivalent circuit theory has been used to separate mediated from paracellular membrane transport of small ions. The term equivalent should be emphasized, as selectivity parameters are obtained from membrane transport data, so phenomenological information is used to quantitate the magnitude of aqueous pathways... 

Ion transport across membranes can be evaluated by using mucosal and serosal electrodes to read transepithelial current (I) and potential difference OP). With these parameters, equivalent circuit analysis can be utilized to account for the relative contributions of transcellular and paracellular pathways. Ionic flux (J) is defined by the Nernst-Planck equation,... 

Additional epithelial aqueous pathways of significantly smaller radius (<3 A) have also been documented utilizing both equivalent pore and circuit theory [25], These pathways may correspond to specific channels through lipid membranes as opposed to paracellular pathways. Osmotically activated ion channels [35] and even specific water channels [36] have been characterized in renal epithelia. In intestinal epithelia, mucosal chloride channels have been studied in secreting crypt cells, and basolateral potassium channels in colonic epithelia serve cellular ion and volume homeostatic functions. 

Since the monolayer is composed of transcellular and paracellular pathways in parallel,... 

The tight junction is a component of the junctional complexes which join cells. Immediately basolateral to the tight junction is the zonula adherens (Figs. 6 and 7). Because the zonula adherens and the gap junctions are focal contact regions, they do not impact transport by the paracellular pathway. All of these junctions are specialized regions of the lateral cell membrane which demarcate the lateral space. In certain types of cells the lateral space is rather narrow and... 

The paracellular pathway consists of the tight junction (TJ) in series with the tortuous lateral space (LS) (Figs. 6 and 7), i.e., mass transfer resistances in series ... 

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... 

Upon taking PABL and Pe into account, the transmonolayer permeability coefficient (PM) was quantified and, in turn, delineated into its component permeabilities for the transcellular and paracellular pathways (Table 11). Specifically, Pparaceii was found as... 

The permeability of the cell monolayer consists of parallel transcellular and paracellular pathways. In passive diffusional transport, it is generally taken that uncharged molecules are capable of partitioning into the cell membrane and... 

Here, Pe takes into account the aqueous boundary layers (/W), transcellular diffusion with metabolism, the paracellular pathway (Pparaceu), and the filter support (PF). It is assumed that drug diffusing through the paracellular route escapes metabolism and contributes insignificantly to the appearance of intact drug in the receiver. 

Penetration enhancers are low molecular weight compounds that can increase the absorption of poorly absorbed hydrophilic drugs such as peptides and proteins from the nasal, buccal, oral, rectal, and vaginal routes of administration [186], Chelators, bile salts, surfactants, and fatty acids are some examples of penetration enhancers that have been widely tested [186], The precise mechanisms by which these enhancers increase drug penetration are largely unknown. Bile salts, for instance, have been shown to increase the transport of lipophilic cholesterol [187] as well as the pore size of the epithelium [188], indicating enhancement in both transcellular and paracellular transport. Bile salts are known to break down mucus [189], form micelles [190], extract membrane proteins [191], and chelate ions [192], While breakdown of mucus, formation of micelles, and lipid extraction may have contributed predominantly to the bile salt-induced enhancement of transcellular transport, chelation of ions possibly accounts for their effect on the paracellular pathway. In addition to their lack of specificity in enhancing mem-... 

There is growing evidence implicating Na+-dependent solute transporters and intracellular as well as extracellular Ca2+ in the physiological regulation of the paracellular pathway [81,203,204], Such modulation of paracellular permeability is especially important for drugs such as peptides and oligonucleotides that exhibit poor permeability characteristics across both the cornea and the conjunctiva [150,152,154,155], In addition, ion transporters such as Cl and Ca2+ channels have been implicated in macromolecular transport (see Sections IV.B.2 and IV.B.4). In the following discussion, some key ion transport processes and their possible roles in solute transport across epithelial tissues are summarized. 

B Hoffmann, I Nagel, W Clauss. (1990). Aldosterone regulates paracellular pathway resistance in rabbit distal colon. J Comp Physiol B 160 381-388. 

JL Madara, JR Pappenheimer. (1987). Structural basis for physiological regulation of paracellular pathways in intestinal epithelium. J Membr Biol 100 149-164. 

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. 

Sugano, K., N. Takata, M. Machida, K. Saitoh, and K. Terada. Prediction of passive intestinal absorption using bio-mimetic artificial membrane permeation assay and the paracellular pathway model, Int. J. Pharm. 2002, 241, 241-251... 

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 other principal pre-clinical PK model - the dog - is not thought to be such a useful model for prediction of absorption in man, because of larger pore size and greater pore frequency in the paracellular pathway of dog compared with rat [59]. 

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