Abstract
The effects of growth factors and calcium concentrations present in different culture media on induction of terminal differentiation were investigated for four different epidermoid carcinoma cell lines, Hela, KB, A431, and SCC-25, and their responses determined relative to those elicited by normal human keratinocytes subjected to these culture conditions. Differentiation status was determined cyto-chemically by a validated keratin protein staining method, and by autoradiographic analyses. Growth and differentiation promoting factors that influenced the direction of integrated control of growth and differentiation in normal human keratinocytes were found to be effective for some cell lines but not others. The factors examined were 1) high density arrest in serum-free and serum-containing media, 2) media shifts from high density culture in serum-containing media to low density growth factor-depleted or supplemented serum-free medium, and 3) the concentration of calcium in the media. The extent and degree of differentiation achieved varied among different cell lines depend on the presence or absence of serum, EGF and insulin protein growth factors. Certain growth media appear to sponsor keratin protein, cyto-chemically-detected differentiation, and evidence of quantal mitotic division in low density HeLa cell and SCC25 cell cultures. Epidermoid carcinoma cell lines retain limited capacity to commit to early stages of cell differentiation.
Highlights
The effects of growth factors and calcium concentrations present in different culture media on induction of terminal differentiation were investigated for four different epidermoid carcinoma cell lines, Hela, KB, A431, and SCC-25, and their responses determined relative to those elicited by normal human keratinocytes subjected to these culture conditions
The results show that KB cultures have a greater propensity to spontaneous undergo commitment to keratinizing stratified squamous differentiation relative to normal human keratinocytes (NHK) grown under high calcium (2 mM) SFM MCDB 153 supplemented with EGF and insulin
Both HeLa and KB cells differs from NHK in failing to differentiate in high calcium MCDB153 medium lacking protein growth factors, but are induced to differentiate in low calcium serum-free MCDB153 medium supplemented with EGF and insulin
Summary
Earlier experimental studies reported that the architecture of normally developing keratinocyte colonies cultured in vitro assumed a structure with the dividing cells located at the colony periphery and histologically-differentiated cells located in the colony interior [2] They reported that kereatinocytes infected with simian virus 40 (SV-40) displayed a drastic restructuring of developing keratinocyte colonies with an profound increase in the proliferative monolayer cells at the periphery and a much reduced multilayered layer differentiation cells present in the colony interior. This demonstration was achieved using Ayoub and Shklar’s (A & S) modification of Mallory’s Trichrome staining method [3], whereby keratin proteins (KP) stained a red color and by autoradiography. Fusenig et al [5] reported altered morphology of transformed human keratinocytes and related them to different degrees of differentiation rather than different stages of malignancy
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