Transepithelial transport of PAMAM dendrimers across isolated rat jejunal mucosae in ussing chambers

Abstract

Oral delivery remains a challenge for poorly permeable hydrophilic macromolecules. Poly(amido amine) (PAMAM) dendrimers have shown potential for their possible oral delivery. Transepithelial transport of carboxyl-terminated G3.5 and amine-terminated G4 PAMAM dendrimers was assessed using isolated rat jejunal mucosae mounted in Ussing chambers. The 1 mM FITC-labeled dendrimers were added to the apical side of mucosae. Apparent permeability coefficients (Papp) from the apical to the basolateral side were significantly increased for FITC when conjugated to G3.5 PAMAM dendrimer compared to FITC alone. Minimal signs of toxicity were observed when mucosae were exposed to both dendrimers with respect to transepithelial electrical resistance changes, carbachol-induced short circuit current stimulation, and histological changes. [(14)C]-mannitol fluxes were not altered in the presence of 1 mM dendrimers, suggesting that the paracellular pathway was not affected at this concentration in this model. These results give insight into the mechanism of PAMAM dendrimer transepithelial rat jejunal transport, as well as toxicological considerations important for oral drug delivery.

Figure 1

Size exclusion chromatograms of G3.5-FITC and G4-FITC conjugates before and after fractionation. Gray line = before fractionation, black line = after fractionation.

Figure 2

P_app of FITC-PAMAM (1.0 mM), FITC-dextran 4 kDa (0.625 mM), FITC-dextran 10 kDa (0.25 mM), and free FITC (0.02 mM) across isolated rat jejunum. FITC-G3.5 dendrimers had significantly increased P_app compared to free FITC (*p < 0.05).

Figure 3

P_app of [¹⁴C]-mannitol through isolated rat jejunum. No significant difference was observed for G4.0 and G3.5 dendrimer (1.0 mM) treatments vs free FITC, indicating no enhanced paracellular transport in the presence of either dendrimer.

Figure 4

Percent TEER changes of isolated jejunal tissue: control (●), G4 dendrimers 1.0 mM (■), G3.5 dendrimers 1.0 mM (◆), FITC-dextran 4 kDa (Δ), FITC-dextran 10 kDa (∇). Percent TEER values were calculated as a percentage of the initial TEER at t = 0 in each group. Significant differences from TEER of each individual group at t = 0 are marked with open circles (G3.5 dendrimers), star (control, G3.5 dendrimers, FITC-dextran 4 kDa), and asterisk (all groups).

Figure 5

ΔI_SC response to basolateral additions of carbachol to jejunal mucosae. No significant difference in response was observed for test groups: control (●), G4 dendrimers 1.0 mM (■), G3.5 dendrimers 1.0 mM (◆), FITC-dextran 4 kDa (Δ), FITC-dextran 10 kDa (∇).

Figure 6

H&E staining of isolated jejunal tissue after 120 min incubation in Ussing chambers. No difference in histology was observed between controls and dendrimer-treated tissue: 1 mM G3.5 dendrimer (upper), 1 mM G4 dendrimer (middle), control (lower). Scale bar = 100 μm for all figures.