α1-Antitrypsin TAQ I polymorphism and α1-antichymotrypsin mutations in patients with obstructive pulmonary disease

Abstract

Obstructive pulmonary disease is a multifactorial condition deriving from the interaction of environmental and genetic factors. From biochemical knowledge of the basis of the disease, alpha 1-antitrypsin and alpha 1-antichymotrypsin are considered two likely candidate genes. We therefore designed an association study comprising 232 unrelated Italian individuals divided as follows: 89 individuals with obstructive lung disease (66 with COPD and 23 with disseminated bronchiectasis) and 143 controls (45 patients with non-obstructive lung disease and 98 healthy individuals). We screened for Taq I (G1237A) polymorphism of the alpha 1-antitrypsin gene as well as the rare variants Bonn-1 (Pro229Ala), Bochum-1 (Leu55Pro), Isehara-1 (Met389Val) and Isehara-2 (1258delAA), and the common signal peptide polymorphism Thr-15Ala of the alpha 1-antichymotrypsin gene. The frequencies of Taq I G1237A alleles were 11.7 and 10.8% in obstructed patients and controls, respectively (P = 0.43), while those of signal peptide Thr-15Ala alleles were 51.6 and 50.3% in obstructed patients and controls, respectively (P = 0.42). We conclude that alpha 1-antitrypsin Taq I polymorphism and alpha 1-antichymotrypsin Thr-15Ala mutation are not major genetic risk factors for the development of obstructive lung disease in Italian patients. The alpha 1-antichymotrypsin rare variants were not detected: our results do not exclude the possibility that other alpha 1-antichymotrypsin gene mutations might be present in Italian obstructed patients but, if so, these genetic defects must be rare.

Diagnosis of alpha-1 antitrypsin deficiency: Modalities, indications and diagnosis strategy

“…These techniques include RFLP (Restriction of Fragment Length Polymorphism) [21][22][23], SSCP (Single Strand Conformation Polymorphism) [24] and DGGE (Denaturing Gradient Gel Electrophoresis) [25], based on the comparative analysis of electrophoresis profiles of DNA fragments, which have different sizes depending on the alleles present. The allelespecific PCR is also commonly used, possibly in multiplex [26]; amplification and hybridization kits with PI S and PI Z allele-specific labelled probes are also marketed [27].…”

“…Il peut aussi s'agir de techniques de RFLP (restriction of fragment lenght polymorphism) [22,23], de SSCP (single strand conformation polymorphism) [24] ou de DGGE (denaturing gradient gel electrophoresis) [25], basées sur l'analyse comparative de profils électrophorétiques de fragments d'ADN qui présentent des tailles différentes selon les allèles présents. La PCR allèle-spécifique est également couramment utilisée, éventuellement en multiplex [26] ; des kits d'amplification puis hybridation avec des sondes marquées spécifiques des allèles PI S et PI Z sont aussi commercialisés [27].…”

A Fast Amplification-Reverse Hybridization Assay Kit to Detect the Most Frequent Deficient Variants in the Alpha-1-Antitrypsin Gene

“…PCR amplification was performed using the same equipment as above. Five picomoles of each primer, as previously described [11]and 0.2 U Taq I DNA polymerase (Bioline) were added to 100 ng of DNA in a 25-µl final volume of a solution containing 50 m M KCl, 10 m M Tris-HCl pH 8.3, 200 µ M of each dNTP and 1.5 m M MgCl 2 . Temperature cycling conditions were as follows: initial 5-min denaturation at 94°C, 30 cycles of 30 s at 94°C, 30 s at 51°C, 30 s at 72°C, and a final extension for 3 min at 72°C.…”

A Novel Method for Rapid Genotypic Identification of Alpha 1-Antitrypsin Variants

“…To check the results obtained by the SexAI/Hpy99I method, all samples were also submitted to genotyping by reported methodologies such as directed sequencing of exons III and V, TaqI RFLP, 17 and an amplification-reverse hybridization commercial kit (Symbiosis, Cocconato, Italy). 18…”