Induction of differentiation-dependent apoptosis in human esophageal squamous cell carcinoma by adenovirus-mediated p21(sdi1) gene transfer

When keratinocytes withdraw from the cell cycle, they migrate from the basa l to the superficial layers of the epidermis and undergo morphological and biochemical changes during the process of terminal differentiation. These d ifferentiation features of keratinocytes are known to be altered or reduced in esophageal cancer cells. Therefore, we examined the effects of transfer ring the cyclin-dependent kinase inhibitor p21(sdi1) gene into human esopha geal cancer cell lines as well as normal keratinocytes using an adenovirus vector system, Ectopic expression of p21(sdi1) protein resulted in cell cyc le arrest at the G(1) phase and produced morphological changes, such as enl arged nuclei and a flattened cellular shape, changes specific to the differ entiated phenotype. The human involucrin protein is a specific product of k eratinocyte differentiation, which is selectively expressed in the suprabas al epidermal layers. Western blot analysis and immunohistochemical staining demonstrated that involucrin expression was 3- to 5-fold enhanced by the f orced expression of p21(sdi1) esophageal cancer cells, whereas only a mild up-regulation up to 1.2-fold occurred in normal keratinocytes. We also foun d that exogenous introduction of the p21(sdi1) gene transcriptionally activ ated the upstream promoter function of the involucrin gene. These stimulato ry effects on involucrin expression were not observed when another cyclin d ependent kinase inhibitor gene, p16(INK4a), was transduced, Moreover, p21(s di1) expression in esophageal cancer cells transduced with p21(sdi1) led to a rapid apoptotic cell death after a transient dormant phase, although ker atinocytes transduced with p21(sdi1) survived longer by terminally withdraw ing from the cell cycle. These results may have an important implication fo r understanding the biology of differentiation-dependent apoptosis in human esophageal squamous cell carcinoma.