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Epidermal growth factor receptor in cultured human retinal pigment epithelial cells.
Ophthalmologica. 2007; 221(4):244-50.O

Abstract

BACKGROUND

Migration and proliferation of retinal pigment epithelial (RPE) cells play an important role in proliferative vitreoretinopathy. Epidermal growth factor receptor (EGFR) is a cell surface receptor with intrinsic tyrosine kinase activity. The engagement of the receptor by its ligand can induce intracellular mitogenic signal transduction pathways and stimulate proliferation, migration and differentiation of cells. This experiment aimed to investigate the activation and role of EGFR signal transduction pathway in proliferation of human RPE cells.

METHODS

Cultured human RPE cells of the 3rd to 6th passages were studied by colorimetric assay for cellular growth and survival (MTT assay) to test the effects of EGF (0.1, 1, 10, 50, and 100 ng/ml) and fetal bovine serum (FBS) on proliferation of human RPE cells. An in vitro wound healing model was also set up, and the number of cells that had entered the denuded area was counted. The human RPE cells were cultured for 3 days with 0.1% FBS, 10% FBS, 10 ng/ml EGF + 0.1% FBS and a combination of EGF and 10% FBS, respectively. Immunohistochemical staining and in situ hybridization were used to observe the expressions of EGFR protein and mRNA, respectively. Activation of mitogen-activated protein kinase (MAPK) was detected by immunohistochemical method with specific antiphosphorylated extracellular signal-regulated kinase (ERK)1/2 antibody.

RESULTS

EGF stimulated proliferation and migration of cultured human RPE cells in a concentration-dependent manner. The maximum of the proliferation rate of RPE cells was 81.8% with EGF at a concentration of 10-100 ng/ml of EGF in serum-free Dulbecco's modified essential medium (DMEM) and 122.7% at a concentration of 1-10 ng/ml of EGF in 5% FBS DMEM (p < 0.001); there was a significant difference between serum-free DMEM groups and 5% FBS DMEM groups. The maximum of the migration rate of the cells was 438.9% at a concentration of 10-100 ng/ml of EGF in 10% FBS DMEM, 147% with 10% FBS, and only 36% with EGF in 0.1% FBS at the concentration of 10 ng/ml (p < 0.001). EGF promoted the expression of EGFR protein and mRNA in RPE cells. FBS cooperated with EGF in the stimulation of EGFR expression, and it had a stronger effect in the process than EGF alone. After 3 days of incubation with EGF, phosphorylated ERK1/2 was detectable in the nucleus of RPE cells, whereas cells presented immunostaining positive for phosphorylated ERK1/2 in the cytoplasm before stimulation, indicating that EGF could induce MAPK nuclear translocation.

CONCLUSION

EGF could induce EGF-EGFR-MAPK signal transduction pathway in human RPE cells in a concentration-dependent manner in vitro, which may play a key role in the activation of human RPE cell proliferation and migration.

Authors+Show Affiliations

Department of Ophthalmology, Xijing Hospital, The Fourth Military Medical University, Chang-le Road 17, 710032 Xi'an, Shaanxi, PR China.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

17579290

Citation

Yan, Feng, et al. "Epidermal Growth Factor Receptor in Cultured Human Retinal Pigment Epithelial Cells." Ophthalmologica. Journal International D'ophtalmologie. International Journal of Ophthalmology. Zeitschrift Fur Augenheilkunde, vol. 221, no. 4, 2007, pp. 244-50.
Yan F, Hui YN, Li YJ, et al. Epidermal growth factor receptor in cultured human retinal pigment epithelial cells. Ophthalmologica. 2007;221(4):244-50.
Yan, F., Hui, Y. N., Li, Y. J., Guo, C. M., & Meng, H. (2007). Epidermal growth factor receptor in cultured human retinal pigment epithelial cells. Ophthalmologica. Journal International D'ophtalmologie. International Journal of Ophthalmology. Zeitschrift Fur Augenheilkunde, 221(4), 244-50.
Yan F, et al. Epidermal Growth Factor Receptor in Cultured Human Retinal Pigment Epithelial Cells. Ophthalmologica. 2007;221(4):244-50. PubMed PMID: 17579290.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Epidermal growth factor receptor in cultured human retinal pigment epithelial cells. AU - Yan,Feng, AU - Hui,Yan-nian, AU - Li,Yang-jun, AU - Guo,Chang-mei, AU - Meng,Hao, PY - 2006/03/16/received PY - 2006/09/22/accepted PY - 2007/6/21/pubmed PY - 2007/7/27/medline PY - 2007/6/21/entrez SP - 244 EP - 50 JF - Ophthalmologica. Journal international d'ophtalmologie. International journal of ophthalmology. Zeitschrift fur Augenheilkunde JO - Ophthalmologica VL - 221 IS - 4 N2 - BACKGROUND: Migration and proliferation of retinal pigment epithelial (RPE) cells play an important role in proliferative vitreoretinopathy. Epidermal growth factor receptor (EGFR) is a cell surface receptor with intrinsic tyrosine kinase activity. The engagement of the receptor by its ligand can induce intracellular mitogenic signal transduction pathways and stimulate proliferation, migration and differentiation of cells. This experiment aimed to investigate the activation and role of EGFR signal transduction pathway in proliferation of human RPE cells. METHODS: Cultured human RPE cells of the 3rd to 6th passages were studied by colorimetric assay for cellular growth and survival (MTT assay) to test the effects of EGF (0.1, 1, 10, 50, and 100 ng/ml) and fetal bovine serum (FBS) on proliferation of human RPE cells. An in vitro wound healing model was also set up, and the number of cells that had entered the denuded area was counted. The human RPE cells were cultured for 3 days with 0.1% FBS, 10% FBS, 10 ng/ml EGF + 0.1% FBS and a combination of EGF and 10% FBS, respectively. Immunohistochemical staining and in situ hybridization were used to observe the expressions of EGFR protein and mRNA, respectively. Activation of mitogen-activated protein kinase (MAPK) was detected by immunohistochemical method with specific antiphosphorylated extracellular signal-regulated kinase (ERK)1/2 antibody. RESULTS: EGF stimulated proliferation and migration of cultured human RPE cells in a concentration-dependent manner. The maximum of the proliferation rate of RPE cells was 81.8% with EGF at a concentration of 10-100 ng/ml of EGF in serum-free Dulbecco's modified essential medium (DMEM) and 122.7% at a concentration of 1-10 ng/ml of EGF in 5% FBS DMEM (p < 0.001); there was a significant difference between serum-free DMEM groups and 5% FBS DMEM groups. The maximum of the migration rate of the cells was 438.9% at a concentration of 10-100 ng/ml of EGF in 10% FBS DMEM, 147% with 10% FBS, and only 36% with EGF in 0.1% FBS at the concentration of 10 ng/ml (p < 0.001). EGF promoted the expression of EGFR protein and mRNA in RPE cells. FBS cooperated with EGF in the stimulation of EGFR expression, and it had a stronger effect in the process than EGF alone. After 3 days of incubation with EGF, phosphorylated ERK1/2 was detectable in the nucleus of RPE cells, whereas cells presented immunostaining positive for phosphorylated ERK1/2 in the cytoplasm before stimulation, indicating that EGF could induce MAPK nuclear translocation. CONCLUSION: EGF could induce EGF-EGFR-MAPK signal transduction pathway in human RPE cells in a concentration-dependent manner in vitro, which may play a key role in the activation of human RPE cell proliferation and migration. SN - 0030-3755 UR - https://www.unboundmedicine.com/medline/citation/17579290/Epidermal_growth_factor_receptor_in_cultured_human_retinal_pigment_epithelial_cells_ L2 - https://www.karger.com?DOI=10.1159/000101926 DB - PRIME DP - Unbound Medicine ER -