Multiplex PCR Assay for the Detection of Genetic Variants of α1-Antitrypsin

摘要

α1-Antitrypsin (A1AT), a highly polymorphic 52-kDa glycoprotein with at least 100 alleles, functions as the major inhibitor of neutrophil elastase (1) . Two allelic variants, PiZ and PiS , frequently lead to A1AT deficiency, which can manifest clinically as emphysema and, less frequently, as liver disease in neonates (2) . Recently, A1AT deficiency has been recognized to play a role in the development of bronchial asthma (3) . The inflammatory process in bronchial asthma involves various mechanisms by which the inflammatory response is perpetuated. These include, for example, macrophage-mediated or mast cell-mediated recruitment and activation of inflammatory cells as well as amplification of neutrophil recruitment and activation. The activated neutrophils release neutrophil elastase, which then can stimulate production of cytokines (4) .For the deficiency variants of A1AT, PiZ and PiS , reduced elastase inactivation capacity has been demonstrated. A diminished inhibitory activity has also been described for PiM2 and PiM1(ala) (5) . The resulting high elastase activity will stimulate cytokine production and can further amplify recruitment of activated neutrophils in the inflamed airway (3)(6)(7) . Recently, an association of the variants PiM2 and PiM1(ala) with bronchial asthma has been reported (5)(8)(9) . Thus, the measurement of the serum A1AT and the identification of the A1AT phenotype are clinically important.Traditionally, the A1AT phenotype has been analyzed by isoelectric focusing. This analysis is tedious and prone to difficulties in interpretation (10) . PiZ and PiS genotypes may be determined by DNA-based methods such as restriction fragment length polymorphism, allele-specific oligonucleotide hybridization, allele-specific amplification, direct sequencing, dual-color detection by ligase-mediated analysis, temperature or denaturing gradient gel electrophoresis, and PCR-mediated site-directed mutagenesis (11)(12)(13)(14)(15)(16) …