Methods:
Asthmatics (n=1277) were categorized into Aspirin exacerbated respiratory disease (AERD) and aspirin-tolerant asthma (ATA). 8 SNPs were genotyped. Messenger RNA expression of the RAB1A gene by peripheral blood mononuclear cell is measured by Real time PCR and reverse transcriptase polymerase chain reaction (RT-PCR). Human PBMC culture supernatant expression of 11-dehydrothromboxane B2. Protein expression of the RAB1A gene by PBMC is measured by RNAi (Knock down) analysis.
Results: The logistic regression analysis showed that the rare allele frequency of +41170 C>G on intron 5 was significantly lower in the AERD group (n=261) than in the ATA group (n=1016) (P=0.002). The linear regression analysis revealed a strong association of +41170 C>G with the aspirin challenge induced-FEV1 fall (P=0.00008). RT-PCR and real time PCR revealed an exon-4-deleted variants. The level of full-length RAB1A mRNA did not differ, but the variants was significantly higher in +41170 G homozygotes than in +41170 C homozygotes (P= 0.002). Intron based PCR was was used to amplify transcripts of PBMC pre-mRNA in which intron-5 had been removed (mRNA) while another set of primers was used to amplify intron 5-containing pre-transcripts (pre-mRNA). Knock down analysis of RAB1A Transcripts level in hPBMC. Thromboxane B2 were increased in the siRNA treated when compared those of scramble and control. After knock down analysis, the levels of thromboxane B2 were significantly decreased in PBMC culture supernatant .
Conclusions:
The rare allele of +41170 C>G may play a protective role against aspirin hypersensitivity via a lower catalytic activity of the RAB1A gene attributed to the increase of a non-functioning variants of RAB1A.