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[Expression of nuclear transcription factor NF-kappaB in human corneal fibroblasts]
1Department of Ophthalmology, The First Hospital of Beijing Medical University, Beijing 100034.
Objective:
To explore the constitutive expression and activation of nuclear transcription factor NF-kappaB in corneal fibroblasts.
Methods:
2 or 3 passage corneal fibroblasts cultured in vitro were divided into two groups after synchronization. One group was cultured in DEME free of serum; the other in DEME with 10 microg/ml lipopolysaccharides for 6 hours. NF-kappaB was studied by flow cytometry (FCM) technique using immunofluorescent staining, and by gel shift assay (EMSA) using gamma-(32)P labeled NF-kappaB oligonucleotide probe.
Results:
According to FCM results, there were constitutively 9.4% NF-kappaB positive corneal fibroblasts, and 2.2 folds more (30.0%) after stimulated with lipopolysaccharides. Using EMSA method, we demonstrated the existence of NF-kappaB with DNA binding activity in the stimulated nuclei.
Conclusion:
There is nuclear transcription factor NF-kappaB in human corneal fibroblasts. When activated, cells express more transcription factor which can enter nuclei, bind itself to the related DNAs and play a regulatory role of gene transcription.