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Increased platelet-activating factor receptor gene expression by corneal epithelial wound healing.
Invest Ophthalmol Vis Sci. 2000 Jun; 41(7):1696-702.IO

Abstract

PURPOSE

Platelet activating factor (PAF) is a potent inflammatory mediator the synthesis of which increases in the cornea after injury. The effects of PAF are mediated by receptors (PAF-R), which are present in target cells. This study was undertaken to investigate the effects of wound healing, PAF, and growth factors on modulating PAF-R mRNA levels in corneal epithelial cells.

METHODS

Cultures of rabbit corneal epithelial (RCE), rabbit limbal epithelial (RLE), rabbit corneal fibroblast (RCF), and rabbit corneal endothelial (RCEn) cells, as well as rabbit corneal keratocytes (RCKs) were used. For the in vivo wound-healing experiments, a 7-mm central corneal deepithelialization was performed in anesthetized rabbits. For the in vitro experiments, wounded rabbit corneas were maintained in organ culture. Corneas were stimulated with 120 nM PAF or preincubated with PAF antagonists, cyclohexamide (CHX) or actinomycin D (AcD) before adding PAF. RCE cells were stimulated with transforming growth factor (TGF)-beta1, -beta2, and, -beta3, basic fibroblast growth factor (bFGF), keratinocyte growth factor (KGF); and hepatocyte growth factor (HGF). Total RNA was isolated and PAF-R expression evaluated by reverse transcription-polymerase chain reaction (RT-PCR), Northern blot analysis, and quantitative RT-PCR.

RESULTS

PAF-R mRNA was expressed in RCE, RLE, and RCEn cells and RCKs, but not in RCFs. After epithelial injury, PAF-R expression increased from 2.5 to 4 times, both in vitro and in vivo. Addition of cPAF further stimulated PAF-R gene expression in epithelium, which was abolished by PAF antagonists. Quantitative RT-PCR revealed that PAF stimulated PAF-R mRNA threefold after injury. The induction of PAF-R by its agonist required previous injury and was inhibited by AcD but not by CHX. Treatment of RCE cells with TGF-beta1, -beta2, or -beta3, HGF, and KGF increased mRNA in PAF-R; however, bFGF had no effect.

CONCLUSIONS

Corneal injury produces changes in PAF-R mRNA expression. Whereas stroma fibroblastic cells lost the PAF-R gene expression found in keratocytes, corneal epithelial injury upregulated PAF-R mRNA. These results suggest that activation of selective growth factors and increases in PAF synthesis after injury stimulate PAF-R gene transcription and constitute important feedback mechanisms needed to maintain the inflammatory process and regulate epithelial wound healing.

Authors+Show Affiliations

Department of Ophthalmology and Neuroscience Center, Louisiana State University Health Sciences Center, New Orleans, USA.No affiliation info available

Pub Type(s)

Journal Article
Research Support, U.S. Gov't, P.H.S.

Language

eng

PubMed ID

10845588

Citation

Ma, X, and H E. Bazan. "Increased Platelet-activating Factor Receptor Gene Expression By Corneal Epithelial Wound Healing." Investigative Ophthalmology & Visual Science, vol. 41, no. 7, 2000, pp. 1696-702.
Ma X, Bazan HE. Increased platelet-activating factor receptor gene expression by corneal epithelial wound healing. Invest Ophthalmol Vis Sci. 2000;41(7):1696-702.
Ma, X., & Bazan, H. E. (2000). Increased platelet-activating factor receptor gene expression by corneal epithelial wound healing. Investigative Ophthalmology & Visual Science, 41(7), 1696-702.
Ma X, Bazan HE. Increased Platelet-activating Factor Receptor Gene Expression By Corneal Epithelial Wound Healing. Invest Ophthalmol Vis Sci. 2000;41(7):1696-702. PubMed PMID: 10845588.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Increased platelet-activating factor receptor gene expression by corneal epithelial wound healing. AU - Ma,X, AU - Bazan,H E, PY - 2000/6/14/pubmed PY - 2000/6/17/medline PY - 2000/6/14/entrez SP - 1696 EP - 702 JF - Investigative ophthalmology & visual science JO - Invest Ophthalmol Vis Sci VL - 41 IS - 7 N2 - PURPOSE: Platelet activating factor (PAF) is a potent inflammatory mediator the synthesis of which increases in the cornea after injury. The effects of PAF are mediated by receptors (PAF-R), which are present in target cells. This study was undertaken to investigate the effects of wound healing, PAF, and growth factors on modulating PAF-R mRNA levels in corneal epithelial cells. METHODS: Cultures of rabbit corneal epithelial (RCE), rabbit limbal epithelial (RLE), rabbit corneal fibroblast (RCF), and rabbit corneal endothelial (RCEn) cells, as well as rabbit corneal keratocytes (RCKs) were used. For the in vivo wound-healing experiments, a 7-mm central corneal deepithelialization was performed in anesthetized rabbits. For the in vitro experiments, wounded rabbit corneas were maintained in organ culture. Corneas were stimulated with 120 nM PAF or preincubated with PAF antagonists, cyclohexamide (CHX) or actinomycin D (AcD) before adding PAF. RCE cells were stimulated with transforming growth factor (TGF)-beta1, -beta2, and, -beta3, basic fibroblast growth factor (bFGF), keratinocyte growth factor (KGF); and hepatocyte growth factor (HGF). Total RNA was isolated and PAF-R expression evaluated by reverse transcription-polymerase chain reaction (RT-PCR), Northern blot analysis, and quantitative RT-PCR. RESULTS: PAF-R mRNA was expressed in RCE, RLE, and RCEn cells and RCKs, but not in RCFs. After epithelial injury, PAF-R expression increased from 2.5 to 4 times, both in vitro and in vivo. Addition of cPAF further stimulated PAF-R gene expression in epithelium, which was abolished by PAF antagonists. Quantitative RT-PCR revealed that PAF stimulated PAF-R mRNA threefold after injury. The induction of PAF-R by its agonist required previous injury and was inhibited by AcD but not by CHX. Treatment of RCE cells with TGF-beta1, -beta2, or -beta3, HGF, and KGF increased mRNA in PAF-R; however, bFGF had no effect. CONCLUSIONS: Corneal injury produces changes in PAF-R mRNA expression. Whereas stroma fibroblastic cells lost the PAF-R gene expression found in keratocytes, corneal epithelial injury upregulated PAF-R mRNA. These results suggest that activation of selective growth factors and increases in PAF synthesis after injury stimulate PAF-R gene transcription and constitute important feedback mechanisms needed to maintain the inflammatory process and regulate epithelial wound healing. SN - 0146-0404 UR - https://www.unboundmedicine.com/medline/citation/10845588/Increased_platelet_activating_factor_receptor_gene_expression_by_corneal_epithelial_wound_healing_ L2 - https://iovs.arvojournals.org/article.aspx?volume=41&issue=7&page=1696 DB - PRIME DP - Unbound Medicine ER -