Ryuichiro Ohashi

Enhanced activity of cyclin A-associated kinase in immortalized human fibroblasts

Deregulation of cell cycle regulatory mechanisms leads to disruption of normal control of cell growth and may be associated with neoplastic transformation. To determine whether immortalized human cells have alterations in their cell cycle regulatory mechanisms, we analyzed cell cycle regulatory proteins in 2 immortalized human fibroblast cell lines, KMST‐6 and OUMS‐24/P6X, established by repeated irradiation with 60Co gamma rays alone or mutant p53 gene transfection plus X‐ray irradiation, respectively. Both the immortalized cell lines had markedly enhanced activity of cyclin A‐associated kinase as compared with their normal counterparts. The high activity of cyclin A‐associated kinase was well correlated with the increased expression of cyclin A mRNA and its protein. In addition, the immortalized cell lines showed significantly reduced amounts of p21Cip1/Waf1/Sdi1, a potent inhibitor of cyclin dependent kinases. Furthermore, among the pRb family of proteins, p107 and p130, were hyperphosphorylated in both the immortalized cell lines, suggesting possible participation in upregulation of cyclin A associated kinase activity. These changes represent some important characteristics of immortalized cells. Int. J. Cancer 82:754–758, 1999.

Pyruvate alleviates toxic effects of ethanol on cells in culture

Abstract Cytotoxic effects of ethanol on cultured human hepatocytes and fibroblasts differed with the type of culture medium used. When cytotoxic effects of ethanol were compared between culture systems using either RPMI-1640 or Dulbeccos modified Eagles medium (DMEM), the cytotoxicity was more prominent with the former medium than with the latter. A reduction of the cytotoxic effects appeared to be due to the pyruvate contained in DMEM. The protective effect of pyruvate against ethanol toxicity may be related to tricarboxylic acid (TCA) cycle activity because addition of malate to the medium also reduced the cytotoxic effects. Our results suggest that drug cytotoxicity testing in vitro must be done using various types of culture medium.