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Conversion of zonulae occludentes from tight to leaky strand type by introducing claudin-2 into Madin-Darby canine kidney I cells.

Furuse M, Furuse K, Sasaki H, Tsukita S - J. Cell Biol. (2001)

Bottom Line: Interestingly, the TER values of MDCK I clones stably expressing claudin-2 (dCL2-MDCK I) fell to the levels of MDCK II cells (>20-fold decrease).Similar results were obtained in mouse epithelial cells, Eph4.These findings indicated that the addition of claudin-2 markedly decreased the tightness of individual claudin-1/4-based TJ strands, leading to the speculation that the combination and mixing ratios of claudin species determine the barrier properties of individual TJ strands.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, Faculty of Medicine, Kyoto University, Kyoto 606-8501, Japan.

ABSTRACT
There are two strains of MDCK cells, MDCK I and II. MDCK I cells show much higher transepithelial electric resistance (TER) than MDCK II cells, although they bear similar numbers of tight junction (TJ) strands. We examined the expression pattern of claudins, the major components of TJ strands, in these cells: claudin-1 and -4 were expressed both in MDCK I and II cells, whereas the expression of claudin-2 was restricted to MDCK II cells. The dog claudin-2 cDNA was then introduced into MDCK I cells to mimic the claudin expression pattern of MDCK II cells. Interestingly, the TER values of MDCK I clones stably expressing claudin-2 (dCL2-MDCK I) fell to the levels of MDCK II cells (>20-fold decrease). In contrast, when dog claudin-3 was introduced into MDCK I cells, no change was detected in their TER. Similar results were obtained in mouse epithelial cells, Eph4. Morphometric analyses identified no significant differences in the density of TJs or in the number of TJ strands between dCL2-MDCK I and control MDCK I cells. These findings indicated that the addition of claudin-2 markedly decreased the tightness of individual claudin-1/4-based TJ strands, leading to the speculation that the combination and mixing ratios of claudin species determine the barrier properties of individual TJ strands.

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Freeze-fracture replica images of TJs in neo-MDCK I (a), dCL2-MDCK I (b), and MDCK II cells (c). Cells (4 × 105 cells) were plated on 24-mm filters, cultured for 6 d, fixed with glutaraldehyde, and then processed for freeze-fracture replica electron microscopy. The number of TJ strands in neo-MDCK I cells was similar to those in dCL2-MDCK I as well as MDCK II cells (Table ), and the network pattern of TJ strands in neo-MDCK I cells did not appear to be more complex than that in dCL2-MDCK I or MDCK II cells. In neo-MDCK I cells (a), TJ strands were largely associated with the P-face and were mostly continuous, and on the E-face (inset) complementary continuous grooves were vacant. In dCL2-MDCK I cells (b), TJ strands were fairly discontinuous on the P-face, and on the E-face (inset) intramembranous particles were scattered within the grooves. The strands (c) and grooves (inset) of MDCK II cells were similar in appearance to those in dCL2-MDCK I cells. *Microvilli. Bar, 100 nm; inset, 50 nm.
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Figure 5: Freeze-fracture replica images of TJs in neo-MDCK I (a), dCL2-MDCK I (b), and MDCK II cells (c). Cells (4 × 105 cells) were plated on 24-mm filters, cultured for 6 d, fixed with glutaraldehyde, and then processed for freeze-fracture replica electron microscopy. The number of TJ strands in neo-MDCK I cells was similar to those in dCL2-MDCK I as well as MDCK II cells (Table ), and the network pattern of TJ strands in neo-MDCK I cells did not appear to be more complex than that in dCL2-MDCK I or MDCK II cells. In neo-MDCK I cells (a), TJ strands were largely associated with the P-face and were mostly continuous, and on the E-face (inset) complementary continuous grooves were vacant. In dCL2-MDCK I cells (b), TJ strands were fairly discontinuous on the P-face, and on the E-face (inset) intramembranous particles were scattered within the grooves. The strands (c) and grooves (inset) of MDCK II cells were similar in appearance to those in dCL2-MDCK I cells. *Microvilli. Bar, 100 nm; inset, 50 nm.

Mentions: Then, we examined TJs of the above clones of dCL2-MDCK I and neo-MDCK I cells by freeze–fracture electron microscopy. As shown in Fig. 5, the network pattern of TJ strands of dCL2-MDCK I clones was very similar to that of neo-MDCK I clones. To compare these networks quantitatively, the mean number of TJ strands in each clone was determined by making numerous counts of TJ strands along a line drawn perpendicular to the junctional axis according to the method described previously (Stevenson et al. 1988). As summarized in Fig. 6 and Table , although the mean number of TJ strands varied to some extent, there was no tendency for the number of strands in dCL2-MDCK I clones to be less than that of neo-MDCK I clones.


Conversion of zonulae occludentes from tight to leaky strand type by introducing claudin-2 into Madin-Darby canine kidney I cells.

Furuse M, Furuse K, Sasaki H, Tsukita S - J. Cell Biol. (2001)

Freeze-fracture replica images of TJs in neo-MDCK I (a), dCL2-MDCK I (b), and MDCK II cells (c). Cells (4 × 105 cells) were plated on 24-mm filters, cultured for 6 d, fixed with glutaraldehyde, and then processed for freeze-fracture replica electron microscopy. The number of TJ strands in neo-MDCK I cells was similar to those in dCL2-MDCK I as well as MDCK II cells (Table ), and the network pattern of TJ strands in neo-MDCK I cells did not appear to be more complex than that in dCL2-MDCK I or MDCK II cells. In neo-MDCK I cells (a), TJ strands were largely associated with the P-face and were mostly continuous, and on the E-face (inset) complementary continuous grooves were vacant. In dCL2-MDCK I cells (b), TJ strands were fairly discontinuous on the P-face, and on the E-face (inset) intramembranous particles were scattered within the grooves. The strands (c) and grooves (inset) of MDCK II cells were similar in appearance to those in dCL2-MDCK I cells. *Microvilli. Bar, 100 nm; inset, 50 nm.
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Figure 5: Freeze-fracture replica images of TJs in neo-MDCK I (a), dCL2-MDCK I (b), and MDCK II cells (c). Cells (4 × 105 cells) were plated on 24-mm filters, cultured for 6 d, fixed with glutaraldehyde, and then processed for freeze-fracture replica electron microscopy. The number of TJ strands in neo-MDCK I cells was similar to those in dCL2-MDCK I as well as MDCK II cells (Table ), and the network pattern of TJ strands in neo-MDCK I cells did not appear to be more complex than that in dCL2-MDCK I or MDCK II cells. In neo-MDCK I cells (a), TJ strands were largely associated with the P-face and were mostly continuous, and on the E-face (inset) complementary continuous grooves were vacant. In dCL2-MDCK I cells (b), TJ strands were fairly discontinuous on the P-face, and on the E-face (inset) intramembranous particles were scattered within the grooves. The strands (c) and grooves (inset) of MDCK II cells were similar in appearance to those in dCL2-MDCK I cells. *Microvilli. Bar, 100 nm; inset, 50 nm.
Mentions: Then, we examined TJs of the above clones of dCL2-MDCK I and neo-MDCK I cells by freeze–fracture electron microscopy. As shown in Fig. 5, the network pattern of TJ strands of dCL2-MDCK I clones was very similar to that of neo-MDCK I clones. To compare these networks quantitatively, the mean number of TJ strands in each clone was determined by making numerous counts of TJ strands along a line drawn perpendicular to the junctional axis according to the method described previously (Stevenson et al. 1988). As summarized in Fig. 6 and Table , although the mean number of TJ strands varied to some extent, there was no tendency for the number of strands in dCL2-MDCK I clones to be less than that of neo-MDCK I clones.

Bottom Line: Interestingly, the TER values of MDCK I clones stably expressing claudin-2 (dCL2-MDCK I) fell to the levels of MDCK II cells (>20-fold decrease).Similar results were obtained in mouse epithelial cells, Eph4.These findings indicated that the addition of claudin-2 markedly decreased the tightness of individual claudin-1/4-based TJ strands, leading to the speculation that the combination and mixing ratios of claudin species determine the barrier properties of individual TJ strands.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, Faculty of Medicine, Kyoto University, Kyoto 606-8501, Japan.

ABSTRACT
There are two strains of MDCK cells, MDCK I and II. MDCK I cells show much higher transepithelial electric resistance (TER) than MDCK II cells, although they bear similar numbers of tight junction (TJ) strands. We examined the expression pattern of claudins, the major components of TJ strands, in these cells: claudin-1 and -4 were expressed both in MDCK I and II cells, whereas the expression of claudin-2 was restricted to MDCK II cells. The dog claudin-2 cDNA was then introduced into MDCK I cells to mimic the claudin expression pattern of MDCK II cells. Interestingly, the TER values of MDCK I clones stably expressing claudin-2 (dCL2-MDCK I) fell to the levels of MDCK II cells (>20-fold decrease). In contrast, when dog claudin-3 was introduced into MDCK I cells, no change was detected in their TER. Similar results were obtained in mouse epithelial cells, Eph4. Morphometric analyses identified no significant differences in the density of TJs or in the number of TJ strands between dCL2-MDCK I and control MDCK I cells. These findings indicated that the addition of claudin-2 markedly decreased the tightness of individual claudin-1/4-based TJ strands, leading to the speculation that the combination and mixing ratios of claudin species determine the barrier properties of individual TJ strands.

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