Rabbit Anti-Mouse Connexin 46 monoclonal antibody, clone 22I27M44

Background and Aims: Connexins and their cell membrane channels contribute to the control of cell proliferation and compartmental functions in breast glands and their deregulation is linked to breast carcinogenesis. Our aim was to correlate connexin expression with tumor progression and prognosis in primary breast cancers. Materials and Methods: Meta-analysis of connexin isotype expression data of 1809 and 1899 breast cancers from the Affymetrix and Illumina array platforms, respectively, was performed. Expressed connexins were also monitored at the protein level in tissue microarrays of 127 patients equally representing all tumor grades, using immunofluorescence and multilayer, multichannel digital microscopy. Prognostic correlations were plotted in Kaplan-Meier curves and tested using the log-rank test and cox-regression analysis in univariate and multivariate models. Results: The expression of GJA1/Cx43, GJA3/Cx46 and GJB2/Cx26 and, for the first time, GJA6/Cx30 and GJB1/Cx32 was revealed both in normal human mammary glands and breast carcinomas. Within their subfamilies these connexins can form homo- and heterocellular epithelial channels. In cancer, the array datasets cross-validated each other's prognostic results. In line with the significant correlations found at mRNA level, elevated Cx43 protein levels were linked with significantly improved breast cancer outcome, offering Cx43 protein detection as an independent prognostic marker stronger than vascular invasion or necrosis. As a contrary, elevated Cx30 mRNA and protein levels were associated with a reduced disease outcome offering Cx30 protein detection as an independent prognostic marker outperforming mitotic index and necrosis. Elevated versus low Cx43 protein levels allowed the stratification of grade 2 tumors into good and poor relapse free survival subgroups, respectively. Also, elevated versus low Cx30 levels stratified grade 3 patients into poor and good overall survival subgroups, respectively. Conclusion: Differential expression of Cx43 and Cx30 may serve as potential positive and negative prognostic markers, respectively, for a clinically relevant stratification of breast cancers.

Connexin 46 (Cx46) is important for gap junction channels formation which plays crucial role in the preservation of lens homeostasis and transparency. Previously, we have identified a missense mutation (p.V44M) of Cx46 in a congenital cataract family. This study aims at dissecting the potential pathogenesis of the causative mutant of cataract. Plasmids carrying wild-type (wt) and mutant (V44M) of Cx46 were constructed and expressed in Hela cells respectively. Western blotting and fluorescence microscopy were applied to analyse the expression and subcellular localization of recombinant proteins, respectively. Scrape loading dye transfer experiment was performed to detect the transfer capability of gap junction channels among cells expressed V44M mutant. The results demonstrated that in transfected Hela cells, both wt-Cx46 and Cx46 V44M were localized abundantly in the plasma membrane. No significant difference was found between the protein expressions of the two types of Cx46. The fluorescent localization assay revealed the plaque formation, significantly reduced in the cells expressing Cx46 V44M. Immunoblotting analysis demonstrated that formation of Triton X-100 insoluble complex decreased obviously in mutant Cx46. Additionally, the scrape-loading dye-transfer experiment showed a lower dye diffusion distance of Cx46 V44M cells, which indicates that the gap junction intercellular communication activity was aberrant. Human Cx46 V44M mutant causing cataracts result in abnormally decreased formation of gap junction plaques and impaired gap junction channel function.