CHROMOSOMAL ALTERATIONS IN PERMANENTLY DIFFERENTIATED HT29 COLON-CARCINOMA CELLS INDUCED WITH SODIUM-BUTYRATE

We have established differentiated subclones from sodium butyrate-treated HT29 colon carcinoma cells. Two subclones, CIII and CVII, grew as tightly-compacted colonies and showed polarized confluent monolayers in vitro. CIII cells formed domes in vitro, a morphological characteristic of columnar absorptive epithelium. Besides these differentiated characteristics, CIII and CVII cells showed lower tumorigenicity than parental HT29 cells. Karyotypic analysis indicated that all cells retained chromosomes in the triploid range. However, modal chromosome numbers among 30 metaphases of each clone were concomitantly reduced from 71 (in HT29) to 68 (in CVII) or 67 (in CHI) with the differentiated phenotypes. Our preliminary karyotype analysis has suggested that the number of chromosomes 10q21-qter, 14q22-qter and 16q were decreased in both subclones. To confirm this, DNAs from these cells and another butyrate-induced subclone, CI-1, were analyzed with several DNA markers. Southern blot analysis indicated that parental HT29 cells were polymorphic for the BamHI and TaqI RFLPs detected by urokinase (7 kbp and 1.6 kbp) and D10S25 (2.5 kbp and 2.1 kbp) probes, respectively, both of which are located on chromosome band 10q. In CI-1 and CVII cells, allelic losses of urokinase (1.6 kbp) and D10S25 (2.5 kbp) were observed, respectively. Although both alleles of these markers were conserved in CIII cells, one allele of urokinase (7 kbp) and D10S25 (2.1 kbp) was less representative. Southern blot analysis using probes for Fos proto-oncogene, D14S20 marker type IV collagenase gene, haptoglobin gene located on chromosome bands 14q and 16q did not show any loss or rearrangement occurred in these subclones. Similarly, no loss or rearrangement was observed for the krev-1 gene on chromosome 1, whose number was not changed in either subclone. These findings indicate that in CI- 1, CIII, and CVII cells, the number of chromosome band 10q is decreased relative to HT29 cells. These deletions may be implicated in the morphological differentiation of HT29 cells treated with sodium butyrate.