Cytokeratin expression in primary epithelial cell culture from bovine conjunctiva

Paladino, Grazia; Marino, Clara; Mulè, Simona La Terra; Civiale, C.; Rusciano, Dario; Enea, Vincenzo

doi:10.1016/j.tice.2004.05.003

Cited by 18 publications

(19 citation statements)

References 22 publications

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“…The observed kinetics of promoter activity, peaking at the second passage (P2) and decreasing to basal levels at P4, is certainly intriguing. In our earlier work on conjunctival epithelial cell cultures [14,15] , we showed that, under air-lift culture conditions and without subculturing, their phenotype remains quite stable for many days [15] . Conjunctival cells grown on transwells fi lters under submerged conditions tend to degenerate faster [14] .…”

Section: Discussionmentioning

confidence: 90%

“…Corneal and conjunctival epithelial tissues and their derived primary cell cultures were prepared from bovine eyes provided by a local slaughterhouse, as previously described [14,15] . Primary BCEC were either grown as monolayers on collagen type I precoated plastic dishes, or as multilayers on Transwell ® culture chamber inserts (Corning, Milan, Italy) at 37 ° C and 5% CO 2 .…”

Section: Tissue Collection and Cell Culturementioning

confidence: 99%

“…Primary BCEC were either grown as monolayers on collagen type I precoated plastic dishes, or as multilayers on Transwell ® culture chamber inserts (Corning, Milan, Italy) at 37 ° C and 5% CO 2 . Cells, when grown as multilayers, were used for RNA extraction when their transepithelial electrical resistance was calculated to be at plateau values, indicating the best physiological state of the culture [14,15] . Sirc (from rabbit cornea, epithelial/fi broblastic mixed morphology) and clone 1-5c-4 (human epithelial cells, originally derived from conjunctiva) continuous cell lines (ATCC, Manassas, Va., USA) were cultured according to ATCC instructions.…”

Section: Tissue Collection and Cell Culturementioning

confidence: 99%

“…In our laboratory we have recently established a method for culturing bovine conjunctival epithelial cells (BCEC) that, as shown by morphological examination and specifi c marker detection, maintain structural and functional features of the original tissue [14,15] . Thus, the purpose of this study was to investigate LF gene expression in conjunctival and corneal epithelial tissues as well as in BCEC.…”

Section: Introductionmentioning

confidence: 99%

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Lactoferrin Expression by Bovine Ocular Surface Epithelia: A Primary Cell Culture Model to Study Lactoferrin Gene Promoter Activity

Santagati¹,

Mulè²,

Amico³

et al. 2005

Ophthalmic Res

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Tear lactoferrin, mainly secreted by the lachrymal glands, exerts a protective effect on the ocular surface, and an abnormal decrease of its production may lead to an increased risk of infection and pathological alterations of ocular surface epithelia. In this study we analyzed whether corneal and conjunctival epithelia could be an additional source of tear lactoferrin, and whether conjunctival epithelial cells in culture could be a suitable model system to address regulation of lactoferrin gene expression. Real-time PCR and Western immunoblotting showed that in bovines lactoferrin is indeed produced by these epithelia, and that the human lactoferrin promoter can direct the expression of a CAT reporter gene, thus indicating that these cells are a true source of lactoferrin, and may be used in vitro to study the regulation of lactoferrin expression.

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Section: Discussionmentioning

confidence: 90%

Section: Tissue Collection and Cell Culturementioning

confidence: 99%

Section: Tissue Collection and Cell Culturementioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Lactoferrin Expression by Bovine Ocular Surface Epithelia: A Primary Cell Culture Model to Study Lactoferrin Gene Promoter Activity

Santagati¹,

Mulè²,

Amico³

et al. 2005

Ophthalmic Res

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“…In the human conjunctiva, goblet cells secreting gel-forming mucins, can occur singly or in greater numbers lining epithelial infoldings or crypts (Shatos et al, 2003). Decrease or loss of mucin/ glycocalix production (Paladino et al, 2004;Pflugfelder et al, 1997) generate squamous metaplasia, which may lead to dry eye and ocular surface diseases. In the conjunctiva, putative epithelial stem cells are concentrated mainly in the fornix region, but can be found also along the whole inner epithelium layer (Lavker et al, 1998).…”

Section: Abstract: Epithelial Basal Precursor Stem Cell Niche Epimentioning

confidence: 99%

Human conjunctival epithelial precursor cells and their progeny in 3D organotypic culture

Rosellini¹,

Papini²,

Giannarini³

et al. 2007

Int. J. Dev. Biol.

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KEY WORDS: epithelial basal precursor, stem cell, niche, epithelium regeneration, interlamellar leukocytesThe conjunctival epithelium is a stratified, non-keratinized epithelium containing goblet epithelial cells. It can be distinguished by the corneal epithelium by the expression of different cytokeratins, mucins and by the presence of glycocalix (Meller et al., 2002). In the human conjunctiva, goblet cells secreting gel-forming mucins, can occur singly or in greater numbers lining epithelial infoldings or crypts (Shatos et al., 2003). Decrease or loss of mucin/ glycocalix production (Paladino et al., 2004;Pflugfelder et al., 1997) generate squamous metaplasia, which may lead to dry eye and ocular surface diseases. In the conjunctiva, putative epithelial stem cells are concentrated mainly in the fornix region, but can be found also along the whole inner epithelium layer (Lavker et al., 1998). Conjunctival epithelial precursor cells are bipotent cells giving rise to either goblet and non-goblet keratinocytes and good candidate to be conjunctival stem/precursor cells (Wei et al., 1997;Pellegrini et al., 1999). Conjunctival epithelial stem/precursor cells can be maintained and expanded ex vivo, if cultivated upon appropriate matrix components, on which both goblet and Int. J. Dev. Biol. 51: 739-743 (2007) doi: 10.1387/ijdb.062198ar non-goblet keratinocytes could be expanded (Santos et al., 2005;Connon et al., 2006).Intact small fragments of conjunctiva were cultivated in an appropriate medium and analysed at different time intervals [prior to culture (T0), after 7 days (T1) and 14 days(T2)] for histology and for the expression of different cellular markers. 3D-organotypic cultures on spongesConjunctival fragments on gelatin-supports, revealed good maintenance of 3D-tissue-architecture and cellular heterogeneity throughout the whole period of cultures. At each time in culture, the stromal compartment appeared well maintained, with few capillaries that remained apparently intact and remarkably unAbbreviations used in this paper: 3D, three dimensional; RT-PCR, reverse transcriptase-polymerase chain reaction. 740 A. Rosellini et al.

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Development and characterization of an organotypic model of Barrett's esophagus

Kosoff

Gardiner

Merlo

et al. 2012

Journal Cellular Physiology

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Understanding the molecular and cellular processes underlying the development, maintenance, and progression of Barrett’s esophagus (BE) presents an empirical challenge because there are no simple animal models and standard 2D cell culture can distort cellular processes. Here we describe a three-dimensional (3D) cell culture system to study Barrett’s esophagus. BE cell lines (CP-A, CP-B, CP-C, and CP-D) and esophageal squamous keratinocytes (EPC2) were cultured on a matrix consisting of esophageal fibroblasts and collagen. Comparison of growth and cytokeratin expression in the presence of all-trans retinoic acid or hydrochloric acid was made by immunohistochemistry and Alcian Blue staining to determine which treatments produced a BE phenotype of columnar cytokeratin expression in 3D culture. All-trans retinoic acid differentially affected the growth of BE cell lines in 3D culture. Notably, the non-dyplastic metaplasia-derived cell line (CP-A) expressed reduced squamous cytokeratins and enhanced columnar cytokeratins upon ATRA treatment. ATRA altered the EPC2 squamous cytokeratin profile towards a more columnar expression pattern. Cell lines derived from patients with high grade dysplasia already expressed columnar cytokeratins and therefore did not show a systematic shift toward a more columnar phenotype with ATRA treatment. ATRA treatment however did reduce the squamoid-like multilayer stratification observed in all cell lines. As the first study to demonstrate long term 3D growth of BE cell lines, we have determined that BE cells can be cultured for at least three weeks on a fibroblast/collagen matrix and that the use of ATRA causes a general reduction in squamous-like multilayered growth and an increase in columnar phenotype with the specific effects cell-line dependent.

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Cytokeratin expression in primary epithelial cell culture from bovine conjunctiva

Cited by 18 publications

References 22 publications

Lactoferrin Expression by Bovine Ocular Surface Epithelia: A Primary Cell Culture Model to Study Lactoferrin Gene Promoter Activity

Lactoferrin Expression by Bovine Ocular Surface Epithelia: A Primary Cell Culture Model to Study Lactoferrin Gene Promoter Activity

Human conjunctival epithelial precursor cells and their progeny in 3D organotypic culture

Development and characterization of an organotypic model of Barrett's esophagus

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