Effects of promyelocytic leukemia zinc finger protein on the proliferation of cultured human corneal endothelial cells.
OBJECTIVE
To determine whether the promyelocytic leukemia zinc finger (PLZF) protein, a transcriptional repressor and negative regulator during cell cycling, plays a role in the proliferation of cultured human corneal endothelial cells (HCECs).


METHODS
The expressions of the mRNA and the protein of PLZF were determined by real-time PCR and western blot analysis, respectively. The changes in the expression of the PLZF gene of cultured HCECs were investigated at different times after cell-cell contacts were disrupted by incubation with EDTA. The cell proliferation rate was assessed with a real-time cell electronic sensing (RT-CES) system after cultured HCECs were infected with either PLZF or LacZ encoding adenovirus vector (Ad-PLZF or Ad-LacZ). The PLZF-regulating genes were analyzed by DNA microarray analysis in cultured HCECs infected with Ad-PLZF.


RESULTS
The expression of the mRNA of PLZF was first detected when the cultured HCECs became confluent, and the relative amount of PLZF mRNA continued to increase for up to 5 days as the cell-cell contacts were formed more firmly. On the other hand, the expression of the mRNA of PLZF decreased about 20 fold 3 h after EDTA exposure, and gradually returned to the original level as the cell-cell contacts were reformed at 72 h after the exposure. The assessment using the RT-CES system showed that the proliferation of cultured HCECs was inhibited for up to 72 h when infected by Ad-PLZF. DNA microarray analysis revealed that the transforming growth factor-beta stimulated clone 22 (TSC-22) gene was up-regulated by 2.32 fold when infected by Ad-PLZF.


CONCLUSIONS
These findings indicate that the expression of PLZF in HCECs is closely related to the formation of cell-cell contacts, and that PLZF may play a role in suppressing their proliferation.
Joko
Takeshi
T
Department of Ophthalmology, Ehime University Graduate School of Medicine, Shitsukawa, Toon, Ehime, Japan. takeshi@m.ehime-u.ac.jp <takeshi@m.ehime-u.ac.jp>
Nanba
Daisuke
D
Shiba
Fumio
F
Miyata
Kazunori
K
Shiraishi
Atsushi
A
Ohashi
Yuichi
Y
Higashiyama
Shigeki
S
D015820:Cadherins / Q000235:genetics; D002450:Cell Communication / Q000502:physiology; D049109:Cell Proliferation; D002478:Cells, Cultured; D003316:Corneal Diseases / Q000150:complications / Q000378:metabolism / Q000473:pathology; D004268:DNA-Binding Proteins / Q000235:genetics; D004728:Endothelium, Corneal / Q000166:cytology* / Q000378:metabolism* / Q000473:pathology; D015870:Gene Expression; D018014:Gene Transfer Techniques; D006801:Humans; D007499:Iris Diseases / Q000150:complications / Q000378:metabolism / Q000473:pathology; D007700:Kinetics; D051741:Kruppel-Like Transcription Factors / Q000235:genetics / Q000502:physiology*; D000075722:Promyelocytic Leukemia Zinc Finger Protein; D012333:RNA, Messenger / Q000378:metabolism; D013577:Syndrome; D014157:Transcription Factors / Q000235:genetics
D015820:Cadherins; D004268:DNA-Binding Proteins; D051741:Kruppel-Like Transcription Factors; D000075722:Promyelocytic Leukemia Zinc Finger Protein; D012333:RNA, Messenger; D014157:Transcription Factors; C092037:ZBTB16 protein, human