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Gap junctions are specialized plasma membrane domains consisting of channels formed by members of the connexin protein family. Gap junctional intercellular communication is often lost in cancers due to aberrant localization or downregulation of connexins, and connexins are therefore suggested to act as tumor suppressor genes in various tissues. The aim of this study was to investigate the expression pattern and DNA promoter methylation status of connexins in colorectal cancer. Expression of six (GJA1, GJA9, GJB1, GJB2, GJC1 and GJD3) connexin genes was detected in normal colonic tissue samples. GJC1 expression was reduced in colorectal carcinomas compared to normal tissue samples. All analyzed connexins were hypermethylated in colon cancer cell lines, although at various frequencies. GJA4, GJB6 and GJD2 were hypermethylated in 60% (29/48), 25% (12/48) and 96% (46/48) of primary colorectal carcinomas, respectively. However, the methylation status was not associated with gene expression. GJC1 has two alternative promoters, which were methylated in 42% (32/76) and 38% (25/65) of colorectal tumors, and in none of the normal mucosa samples. Expression of GJC1 was significantly lower in methylated compared with unmethylated samples (p < 0.01) and was restored in cell lines treated with the demethylating drug 5-aza-2'deoxycytidine. Taken together, DNA hypermethylation of the promoter region of GJC1, encoding connexin45, is an important mechanism in silencing gene expression in colorectal cancer.

Mutations in the KCNQ1, HERG, SCN5A, minK and MiRP1 genes cause long QT syndrome (LQTS), of which there are two forms: the Romano Ward syndrome and the Jervell and Lange-Nielsen syndrome. We have performed DNA sequencing of the LQTS-associated genes in 169 unrelated patients referred for genetic testing with respect to Romano Ward syndrome and in 13 unrelated patients referred for genetic testing with respect to Jervell and Lange-Nielsen syndrome. A total of 37 different mutations in the 5 genes, of which 20 were novel, were identified. Among patients with the most stringent clinical criteria of Romano Ward syndrome, a mutation was identified in 71%. Twelve of the 13 unrelated patients referred for genetic testing with respect to Jervell and Lange-Nielsen syndrome were provided with a molecular genetic diagnosis. Cascade genetic screening of 505 relatives of index patients with molecularly defined LQTS identified 251 mutation carriers. The observed penetrance was 41%. Although caution must be exerted, the prevalence of heterozygotes for mutations in the LQTS-associated genes in Norway could be in the range 1/100-1/300, based on the prevalence of patients with Jervell and Lange-Nielsen syndrome.

Background: Adenomatous polyposis coli (APC) and b-catenin (encoded by CTNNB1) are important components in the WNT signalling pathway, a pathway altered in nearly all colorectal tumours. Cone icting results are reported on whether APC mutations are less common in tumours with a high degree ofmicrosatellite instability (MSI-H)than in microsatellite stable (MSS) ones,and whether mutations in the regulatory domain of CTNNB1 substitute for APC mutations in the MSI-H tumours. Methods: A consecutive series of 218 primary colorectal carcinomas, stratie ed by MSI status, were analysed for mutations in the APC gene (by the protein truncation test) and in the CTNNB1 gene (by single-strand conformation polymorphism). Results: APC mutations detected in 66% of the patients were signie cantly more frequent in the MSS and MSI-L (low) tumours than in the MSI-H tumour group (P < 0.001). The MSI-H tumours tended to have more frameshift mutations than the MSS/MSI-L tumours. The majority of the APC mutations were located in the mutation cluster region (MCR). Patients that had lost all b-catenin binding sites of the APC gene showed a shorter survival time than patients who retained some or all of these binding sites (P = 0.045). Two mutations were found in the CTNNB1 gene, but neither of them was located in the regulatory domain in exon 3. Conclusion: This study conerms that APC mutations are less frequent in MSI-H tumours than in MSS and MSI-L tumours. However, CTNNB1 mutations do not substitute for APC mutations in MSI-H tumours in these Norwegian patients.

Colorectal carcinogenesis is regarded as a multistep process involving several genetic alterations, with mutation in the K-ras gene in about half of the tumours. We aimed at clarifying the role of this genetic alteration related to survival and clinicopathologic variables.One hundred large-bowel carcinomas operated on between 1978 and 1982 were studied for the presence of point mutations in codons 12 and 13 of the K-ras gene, using enriched polymerase chain reaction amplification, restriction fragment length polymorphism analysis, and direct sequencing.Forty mutations were found (40%): 31 in codon 12 and 9 in codon 13, 7 different types. There was no relationship between tumours with and without K-ras mutations with regard to Dukes' stages, age or sex of the patient, tumour localization, histologic grade, DNA ploidy pattern, HLA-DR staining pattern, or survival. Samples from 5 different localizations in 7 carcinomas showed identical K-ras mutation pattern, as did 19 recurrences/ metastases originating from 11 carcinomas.When present, the primary tumour shows homogeneous distribution of K-ras mutation, and the mutation follows the carcinoma in the secondary deposit, regardless of lymphogenous or hematogenous spread. The presence of K-ras mutation does not seem to have prognostic significance for the patient, and the precise nucleotide change is furthermore not predictive of tumour behaviour.

Serum carcinoembryonic antigen (CEA) levels in relation to survival, flow cytometric DNA ploidy pattern, Dukes stage, and recurrent disease was prospectively evaluated in 406 patients with colorectal carcinoma. In 246 patients (61%) the carcinomas were DNA aneuploid. Increased preoperative CEA levels (> 5 micrograms/l) were found in 151 of 363 evaluable patients (42%). Dukes stage-B patients with preoperative CEA elevation showed significantly poorer prognosis than those with normal CEA values (p = 0.001). A weak but significant correlation was found between preoperative CEA level and Dukes stage (Kendall's tau = 0.25, p < 0.01). Of 50 evaluable patients with clinical recurrence and postoperative normal or normalized CEA levels, 28 (56%) had a rise in CEA before or at the time of clinical recurrence. The sensitivity of the CEA test for primary and for recurrent disease was not significantly different in the DNA aneuploid and the DNA near-diploid groups.

In a material of 513 consecutively collected large-bowel polyps, tubular and villous adenomas with corresponding histologic grades of epithelial dysplasia were selected. Fifty-two samples of the former and 27 of the latter category were evaluated semiquantitatively for immunohistochemical staining of the epithelial marker proteins carcinoembryonic antigen (CEA), secretory component (SC), and IgA. No consistent differences in the overall staining properties were found between the two types of tumours. In both categories the staining intensity for SC and epithelial IgA decreased with increasing degree of dysplasia, most strikingly so for IgA. This observation shows that the malignant alterations impair the capacity of the columnar epithelium to produce SC and to express it as a functional receptor mediating uptake of dimeric IgA. Thus, the absence of epithelial IgA in large-bowel adenomas may be a useful adjunct in grading of dysplasia. Conversely, the presence of CEA did not seem to be related to dysplastic alterations in the epithelium.

The main function of secretory IgA is to exert immune exclusion; that is, by intimate cooperation with innate non-specific defence mechanisms, it dampens down penetration of soluble antigens and inhibits epithelial colonisation of bacteria and viruses. Secretory IgM may exert a similar protective function in the gut as its local synthesis sometimes is markedly increased, especially in selective IgA deficiency. IgG should not be considered a secretory immunoglobulin because its external translocation depends on passive intercellular diffusion. By activating complement, antibodies of this isotype may cause increased mucosal permeability and tissue damage. IgG may thus contribute to persistent immunopathology in mucosal lesions. The same is true for IgE antibodies which, in atopic individuals, may be carried into the gut mucosa by mast cells and cause their degranulation with histamine release. Secretory IgA and secretory IgM are the products of two cell types: plasma cells synthesise IgA dimers and IgM pentamers which, by non-covalent association, become complexed with the secretory component (SC) which is synthesized by serous-type glandular cells. The adsorption of the Ig polymers to the SC-expressing epithelial cells depends on J chain-determined binding sites. This fact gives biological significance to the striking J chain expression shown by mucosal immunocytes regardless of the Ig class they produce. The immunocytes populating the gut mucosa apparently belong to relatively early memory B cell clones. The obvious functional goal of J chain expression at this stage of clonal differentiation is local generation of SC-binding IgA and IgM polymers. In various gut diseases, altered immune regulation results in a disproportionately increased number of J chain-negative IgG-producing cells in the mucosa. Such altered immunological homeostasis may contribute to perpetuation of inflammatory bowel diseases.