Anti-LOXL1 (aa 570-590) polyclonal antibody

Lysyl oxidase-like 1 (LOXL1) is an amino-oxidase involved in maturation of elastic fibers. Its downregulation has been associated with elastic fibers repair loss in aging aorta, lung, ligament, and skin. Several evidences of LOXL1 epigenetic silencing by promoter methylation were reported in cancer and cutis laxa syndrome. We hypothesized that this mechanism could be implicated in skin aging process, as far as elastic fibers are also concerned. Anti-DNMT3A chromatin immunoprecipitation was conducted with nuclear extracts from skin fibroblasts isolated from young and elderly individuals, and showed a higher level of DNMT3A protein binding to the LOXL1 promoter in older cells concomitantly to the decrease of LOXL1 mRNA expression and the increase of LOXL1 promoter methylation. Using luciferase reporter assay driven by LOXL1 promoter in HEK293 cells, we demonstrated that LOXL1 transcriptional activity was dramatically reduced when a recombinant DNMT3A was concomitantly overexpressed. LOXL1 promoter transcriptional activity was restored in the presence of a broad-spectrum inhibitor of DNMT activity, 5-aza-2'-deoxycytidine. Finally, to assess whether the interplay between DNMT3A and LOXL1 promoter could be targeted to increase LOXL1 mRNA expression level, an Origanum majorana extract was selected among 43 plant extracts as a new inhibitor of human DNMT3A activity to restore LOXL1 secretion without cytotoxicity in aged skin fibroblasts.

Genetic and nongenetic factors contribute to development of pseudoexfoliation (PEX) syndrome, a complex, age-related, generalized matrix process frequently associated with glaucoma. To identify specific genetic variants underlying its etiology, we performed a genome-wide association study (GWAS) using a DNA-pooling approach. Therefore, equimolar amounts of DNA samples of 80 subjects with PEX syndrome, 80 with PEX glaucoma (PEXG) and 80 controls were combined into separate pools and hybridized to 500K SNP arrays (Affymetrix). Array probe intensity data were analyzed and visualized with expressly developed software tools GPFrontend and GPGraphics in combination with GenePool software. For replication, independent German cohorts of 610 unrelated patients with PEX/PEXG and 364 controls as well as Italian cohorts of 249 patients and 190 controls were used. Of 19, 17 SNPs showing significant allele frequency difference in DNA pools were confirmed by individual genotyping. Further single genotyping at CNTNAP2 locus revealed association between PEX/PEXG for two SNPs, which was confirmed in an independent German but not the Italian cohort. Both SNPs remained significant in the combined German cohorts even after Bonferroni correction (rs2107856: P-c=0.0108, rs2141388: P-c=0.0072). CNTNAP2 was found to be ubiquitously expressed in all human ocular tissues, particularly in retina, and localized to cell membranes of epithelial, endothelial, smooth muscle, glial and neuronal cells. Confirming efficiency of GWAS with DNA-pooling approach by detection of the known LOXL1 locus, our study data show evidence for association of CNTNAP2 with PEX syndrome and PEXG in German patients. European Journal of Human Genetics (2011) 19, 186-193; doi:10.1038/ejhg.2010.144; published online 1 September 2010