Regulation of alphavbeta3 and alpha5beta1 integrin function plays a crucial role in atherosclerosis. Possible regulators of integrin-matrix interactions are integrin-binding ADAMs (proteins with a disintegrin- and metalloproteinase-domain), like ADAM-15 and ADAM-9. Molecular interactions between ADAM-15, alpha5beta1, and alphavbeta3 have been demonstrated. ADAM-9 and ADAM-15 were found to be interdependently regulated. This study, therefore, investigated whether the upregulation of integrins alpha5beta1 and alphavbeta3 was correlated with the expression of integrin-binding ADAMs in atherosclerotic processes. Human arterial and venous vascular smooth muscle cells (VSMCs) were incubated with PDGF over different time intervals up to a 3-day culture period. mRNA concentrations, quantified by real-time RT-PCR and normalized to PBGD, of integrins alphavbeta3 and alpha5beta1 were strongly increased after a 12-h PDGF-incubation in arterial and venous VSMC. ADAM-15 and ADAM-9 mRNA production showed a corresponding increase following integrin upregulation after a 24-h incubation period. Western blot anaylsis revealed an increased protein expression of integrins and ADAMs in PDGF-stimulated VSMC. Additionally, mRNA concentrations of atherosclerotic and normal human specimens were quantified by real-time RT-PCR. mRNA of ADAMs and integrins was significantly increased in atherosclerotic arteries compared to normal arteries. Immunohistochemistry of these specimens showed an increased expression and codistribution of both ADAMs and integrins in atherosclerosis. In conclusion, upregulation of ADAM-15 and ADAM-9 in atherosclerosis appears to follow an increase in alpha5beta1 and alphavbeta3 integrins. Since alpha5beta1 and alphavbeta3 are known to promote smooth muscle cell migration and proliferation, upregulation of ADAM-15 and ADAM-9 could balance integrin-matrix interactions and cell migration, thus modulating neointima progression.

Paraganglioma (PGL) is a rare disorder characterized by tumors of the head and neck region. Between 10% and 50% of cases of PGL are familial, and the disease is autosomal dominant and subject to age-dependent penetrance and imprinting. The paraganglioma gene (PGL1) has been mapped to 11q22.3-q23, and recently germline mutations in the SDHD gene have been identified. The SDHD region contains another gene, DPP2/TIMM8B, the homolog of which causes dystonia and deafness seen in Mohr-Tranebjaerg syndrome. Using four PGL pedigrees, two of which exhibit coinheritance of PGL and sensorineural hearing loss or tinnitus, analysis of 14 microsatellite markers provided support for linkage to the PGL1 locus. Sequence analysis identified novel mutations in exon 1 and exon 3 of the SDHD gene, including a novel two base pair deletion in exon 3 creating a premature stop codon at position 67; a novel three base pair deletion in exon 3 resulting in the loss of Tyr-93; a missense mutation in exon 3 resulting in the substitution of Leu-81 for Pro-81; and a novel G-to-C substitution in exon 1 resulting in the substitution of Met-1 for Ile-1. No base changes were detected in the DPP2/TIMM8B gene. There was no apparent loss of heterozygosity at the site of the SDHD mutations. However, RT-PCR analysis of tumor samples showed monoallelic expression of the mutant (paternal) allele as expected for imprinting. This has not previously been shown for this disorder. The inheritance and expression of the SDHD gene is consistent with the PGL1 gene being subject to genomic imprinting.

Plasminogen activator inhibitor-1 (PAI-1) and two-chain high molecular weight kininogen (HKa) exert anti-adhesive properties in vitronectin-dependent cell adhesion. Here, the hypothesis was tested that these anti-adhesive components promote apoptosis in vascular cells. PAI-1 or HKa induced a 2- to 3-fold increase in apoptosis of human umbilical-vein endothelial cells (HUVEC) and vascular smooth muscle cells (VSMC) adherent to vitronectin, as determined by annexin V-FACS assay, similar to alphav-integrin inhibitor cyclo-(Arg-Gly-Asp-D-Phe-Val)-peptide (cRGDfV). Apoptosis occurred after 12 h incubation and was attributable to caspase 3 activation that in turn induced DNA fragmentation. Induction of apoptosis strongly correlated with the anti-adhesive effect of PAI-1 and HKa on these cells. In contrast, PAI-1 and HKa did not affect fibronectin-dependent adhesion or cell survival. uPA did not influence apoptosis in vitronectin- or fibronectin-adherent cells. In atherosclerotic vessel sections, congruent distribution of vitronectin, PAI-1, HK, and of components of the urokinase plasminogen activator/receptor system with apoptotic cells lining foam cell lesions was demonstrated by immunostaining. These results indicate that inhibition of vitronectin-dependent cell adhesion through PAI-1 and HKa correlates with apoptosis induction in vascular cells mediated through the caspase 3 pathway. Co-distribution of apoptosis with plasminogen activation system components in atherosclerosis exemplifies the significance of anti-adhesive mechanisms and apoptosis for tissue remodeling, such as in neointima development.