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Regulation of Limbal Keratinocyte Proliferation and Differentiation by TAp63 and Np63 Transcription Factors

¹From the Department of Physiology, College of Medicine, Chang Gung University Kweishan, Taoyuan, Taiwan; the ²Section of Biologics, Division of Drug Biology, Bureau of Food and Drug Analysis, Department of Health, Taiwan; and the ³Department of Ophthalmology, Chang Gung Memorial Hospital, Linkou, Taipei, Taiwan.

PURPOSE. To examine the effects of TAp63 and Np63 on the proliferation and differentiation of rabbit limbal keratinocytes cultured on human amniotic membrane.

METHOD. Real-time Q-RT-PCR was used to quantify the relative abundance of TAp63 and Np63 transcripts in limbal, peripheral corneal, and central corneal epithelia. Antisense oligonucleotides were designed specifically to suppress the expression of TAp63 or Np63 in limbal keratinocytes, and their effects on cell proliferation and differentiation were examined. Immunofluorescence was used to examine the expressions of p63 and keratin-3 and -14.

RESULTS. The expressions of TAp63 and Np63 transcripts appeared to be site specific. TAp63 was expressed at the highest level in limbus, decreased by approximately 10-fold in peripheral cornea and was undetectable in the central cornea. Np63 was also expressed at the highest level in limbus, decreased by approximately 35% in peripheral cornea, and was undetectable in the central cornea. Suppression of TAp63 expression inhibited limbal keratinocyte proliferation but promoted differentiation. Suppression of Np63 expression also inhibited cell proliferation but had no obvious effect on cell differentiation.

CONCLUSIONS. TAp63 and Np63 affect the proliferation of limbal keratinocytes by a different mechanism. The inhibition by TAp63 antisense oligos appeared to be secondary to the promotion of cell differentiation. In contrast, the inhibition by Np63 antisense oligos appeared to be independent of cell differentiation.

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