Expression of claudins -2 and -4 and cingulin is coordinated with the start of stratification and differentiation in corneal epithelial cells: retinoic acid reversibly disrupts epithelial barrier
Summary Although tight junctions (TJ) have been extensively studied in simple epithelial cells, it is still unknown whether their organization is coupled to cell differentiation in stratified epithelia. We studied the expression of TJ in RCE1(5T5) cells, an in vitro model which mimics the sequential...
| Main Authors: | , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
The Company of Biologists
2012-11-01
|
| Series: | Biology Open |
| Subjects: | |
| Online Access: | http://bio.biologists.org/content/2/2/132 |
| _version_ | 1818740291565256704 |
|---|---|
| author | María Teresa Ortiz-Melo Erika Sánchez-Guzmán Arturo González-Robles Jesús Valdés Eber Gómez-Flores Federico Castro-Muñozledo |
| author_facet | María Teresa Ortiz-Melo Erika Sánchez-Guzmán Arturo González-Robles Jesús Valdés Eber Gómez-Flores Federico Castro-Muñozledo |
| author_sort | María Teresa Ortiz-Melo |
| collection | DOAJ |
| description | Summary
Although tight junctions (TJ) have been extensively studied in simple epithelial cells, it is still unknown whether their organization is coupled to cell differentiation in stratified epithelia. We studied the expression of TJ in RCE1(5T5) cells, an in vitro model which mimics the sequential steps of rabbit corneal epithelial differentiation. RCE1(5T5) cells expressed TJ components which were assembled once cells constituted differentiated epithelia, as suggested by the increase of transepithelial electrical resistance (TER) which followed a similar kinetic to the expression of the early differentiation marker Pax-6. TJ were functional as indicated by the establishment of an epithelial barrier nonpermeable to ruthenium red or a biotin tracer. In immunostaining experiments, TJ were located at the superficial cells from the suprabasal layers; Western blot and RT-PCR suggested that TJ were composed of claudins (cldn) -1, -2, -4, cingulin (cgn), occludin (ocln) and ZO-1. Semi-quantitative RT-PCR and TER measurements showed that TJ became organized when cells began to form a 3–5 layers stratified epithelium; TER increased once cells reached confluence, with a time course comparable to the raise in the expression of cgn, cldn-2 and -4. Nevertheless, cldn-1, -2, ZO-1 and ocln were present in the cells from the beginning of cultivation, suggesting that TER increases mainly depend on TJ assembly. While EGF increased epithelial barrier strength, retinoic acid disrupted it, increasing paracellular flux about 2-fold; this effect was concentration dependent and completely reversible. Our results suggest that TJ assembly is tightly linked to the expression of corneal epithelial terminal phenotype. |
| first_indexed | 2024-12-18T01:38:24Z |
| format | Article |
| id | doaj.art-df6d78faf961428f9f0b4e2d645fb145 |
| institution | Directory Open Access Journal |
| issn | 2046-6390 |
| language | English |
| last_indexed | 2024-12-18T01:38:24Z |
| publishDate | 2012-11-01 |
| publisher | The Company of Biologists |
| record_format | Article |
| series | Biology Open |
| spelling | doaj.art-df6d78faf961428f9f0b4e2d645fb1452022-12-21T21:25:25ZengThe Company of BiologistsBiology Open2046-63902012-11-012213214310.1242/bio.2012314520123145Expression of claudins -2 and -4 and cingulin is coordinated with the start of stratification and differentiation in corneal epithelial cells: retinoic acid reversibly disrupts epithelial barrierMaría Teresa Ortiz-Melo0Erika Sánchez-Guzmán1Arturo González-Robles2Jesús Valdés3Eber Gómez-Flores4Federico Castro-Muñozledo5 Departamento de Biología Celular, Centro de Investigación y de Estudios Avanzados del Instituto Politécnico Nacional, Apartado Postal 14-740, México City 07000, México Departamento de Biología Celular, Centro de Investigación y de Estudios Avanzados del Instituto Politécnico Nacional, Apartado Postal 14-740, México City 07000, México Departamento de Infectómica y Patogénesis Molecular, Centro de Investigación y de Estudios Avanzados del Instituto Politécnico Nacional, México City 07000, México Departamento de Bioquímica, Centro de Investigación y de Estudios Avanzados del Instituto Politécnico Nacional, México City 07000, México Departamento de Biología Celular, Centro de Investigación y de Estudios Avanzados del Instituto Politécnico Nacional, Apartado Postal 14-740, México City 07000, México Departamento de Biología Celular, Centro de Investigación y de Estudios Avanzados del Instituto Politécnico Nacional, Apartado Postal 14-740, México City 07000, México Summary Although tight junctions (TJ) have been extensively studied in simple epithelial cells, it is still unknown whether their organization is coupled to cell differentiation in stratified epithelia. We studied the expression of TJ in RCE1(5T5) cells, an in vitro model which mimics the sequential steps of rabbit corneal epithelial differentiation. RCE1(5T5) cells expressed TJ components which were assembled once cells constituted differentiated epithelia, as suggested by the increase of transepithelial electrical resistance (TER) which followed a similar kinetic to the expression of the early differentiation marker Pax-6. TJ were functional as indicated by the establishment of an epithelial barrier nonpermeable to ruthenium red or a biotin tracer. In immunostaining experiments, TJ were located at the superficial cells from the suprabasal layers; Western blot and RT-PCR suggested that TJ were composed of claudins (cldn) -1, -2, -4, cingulin (cgn), occludin (ocln) and ZO-1. Semi-quantitative RT-PCR and TER measurements showed that TJ became organized when cells began to form a 3–5 layers stratified epithelium; TER increased once cells reached confluence, with a time course comparable to the raise in the expression of cgn, cldn-2 and -4. Nevertheless, cldn-1, -2, ZO-1 and ocln were present in the cells from the beginning of cultivation, suggesting that TER increases mainly depend on TJ assembly. While EGF increased epithelial barrier strength, retinoic acid disrupted it, increasing paracellular flux about 2-fold; this effect was concentration dependent and completely reversible. Our results suggest that TJ assembly is tightly linked to the expression of corneal epithelial terminal phenotype.http://bio.biologists.org/content/2/2/132Corneal epitheliumTight junctionRetinoic acidClaudinOccludinCingulinCell differentiationCell culture |
| spellingShingle | María Teresa Ortiz-Melo Erika Sánchez-Guzmán Arturo González-Robles Jesús Valdés Eber Gómez-Flores Federico Castro-Muñozledo Expression of claudins -2 and -4 and cingulin is coordinated with the start of stratification and differentiation in corneal epithelial cells: retinoic acid reversibly disrupts epithelial barrier Biology Open Corneal epithelium Tight junction Retinoic acid Claudin Occludin Cingulin Cell differentiation Cell culture |
| title | Expression of claudins -2 and -4 and cingulin is coordinated with the start of stratification and differentiation in corneal epithelial cells: retinoic acid reversibly disrupts epithelial barrier |
| title_full | Expression of claudins -2 and -4 and cingulin is coordinated with the start of stratification and differentiation in corneal epithelial cells: retinoic acid reversibly disrupts epithelial barrier |
| title_fullStr | Expression of claudins -2 and -4 and cingulin is coordinated with the start of stratification and differentiation in corneal epithelial cells: retinoic acid reversibly disrupts epithelial barrier |
| title_full_unstemmed | Expression of claudins -2 and -4 and cingulin is coordinated with the start of stratification and differentiation in corneal epithelial cells: retinoic acid reversibly disrupts epithelial barrier |
| title_short | Expression of claudins -2 and -4 and cingulin is coordinated with the start of stratification and differentiation in corneal epithelial cells: retinoic acid reversibly disrupts epithelial barrier |
| title_sort | expression of claudins 2 and 4 and cingulin is coordinated with the start of stratification and differentiation in corneal epithelial cells retinoic acid reversibly disrupts epithelial barrier |
| topic | Corneal epithelium Tight junction Retinoic acid Claudin Occludin Cingulin Cell differentiation Cell culture |
| url | http://bio.biologists.org/content/2/2/132 |
| work_keys_str_mv | AT mariateresaortizmelo expressionofclaudins2and4andcinguliniscoordinatedwiththestartofstratificationanddifferentiationincornealepithelialcellsretinoicacidreversiblydisruptsepithelialbarrier AT erikasanchezguzman expressionofclaudins2and4andcinguliniscoordinatedwiththestartofstratificationanddifferentiationincornealepithelialcellsretinoicacidreversiblydisruptsepithelialbarrier AT arturogonzalezrobles expressionofclaudins2and4andcinguliniscoordinatedwiththestartofstratificationanddifferentiationincornealepithelialcellsretinoicacidreversiblydisruptsepithelialbarrier AT jesusvaldes expressionofclaudins2and4andcinguliniscoordinatedwiththestartofstratificationanddifferentiationincornealepithelialcellsretinoicacidreversiblydisruptsepithelialbarrier AT ebergomezflores expressionofclaudins2and4andcinguliniscoordinatedwiththestartofstratificationanddifferentiationincornealepithelialcellsretinoicacidreversiblydisruptsepithelialbarrier AT federicocastromunozledo expressionofclaudins2and4andcinguliniscoordinatedwiththestartofstratificationanddifferentiationincornealepithelialcellsretinoicacidreversiblydisruptsepithelialbarrier |