MR analysis
We performed MR20 analyses by using genetic
instrumental variables (IVs) to proxy for candidate obesity-related
factors. The use of MR enables unbiased causal effects of risk factors
on respiratory outcomes to be estimated if the following assumptions are
met: (1) the genetic IVs are robustly associated with the risk factor of
interest; (2) the genetic IVs are not associated with any confounding
factors; (3) the pleiotropic pathways through which the IVs influence
the outcome are all due to the risk factors. GRS composed of risk
factors associated-single-nucleotide polymorphism (SNP) was used as IVs.
The dosage of the effect allele was multiplied by the regression
coefficients of each SNP on associated traits divided by the mean value
of all regression coefficients.
By referencing candidate SNPs from large genome-wide association
studies, we collected well-known genetic variants associated with the
five obesity-related factors. To avoid the pleiotropic effects of
genetic variants, SNPs previously reported to be associated with asthma
and confounders were excluded. After we selected SNPs for the
obesity-related factors, some criteria were applied to filter the
selected SNPs for quality control: (1) minor allele frequency of ≥5%,
(2) genotyping call rate of >98% for all children, and (3)
no linkage disequilibrium with candidate SNPs. The primers were designed
using the National Genotyping Center of the Academia Sinica platform
(http://lims.ngc.sinica.edu.tw/service/)21.
Genotyping was performed using Sequenom iPLEX matrix-assisted laser
desorption/ionization–time of flight mass spectrometry at the National
Center for Genome Medicine, Taiwan. The online supplementary file
provides details on how candidate SNPs were selected.