Establishment and application of detection method for interferon stimulated genes expression in children
10.3760/cma.j.cn114452-20210927-00608
- VernacularTitle:儿童干扰素刺激基因表达检测方法的建立与应用
- Author:
Wendao LI
1
;
Wei WANG
;
Wei WANG
;
Mingsheng MA
;
Hongmei SONG
Author Information
1. 中国医学科学院北京协和医学院北京协和医院儿科，北京100730
- Keywords:
Interferon type Ⅰ;
Interferon stimulated genes;
Interferon score;
Type Ⅰ interferonopathies;
Real time fluorescence quantitative PCR
- From:
Chinese Journal of Laboratory Medicine
2022;45(6):603-609
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To establish the detection method for the interferon stimulated genes(ISGs), calculate the cut-off value and test it in clinical practice.Methods:Patients with type I interferonopathies who were admitted to Peking Union Medical College Hospital from November 2017 to September 2021 were chosen as the disease group, and healthy children were included as the control group. A total of 18 children were in the disease group, including 8 males and 10 females, with a median age of 8.5 years for the first test. From them 25 blood specimens were collected. A total of 28 healthy children, aged 1 to 18 years, with a median age of 10.5 years, including 15 males and 13 females, were included in the control group. Blood samples of 34 controls and 18 interferonopathies patients were collected, then total RNA extraction and cDNA synthesis were performed. Real-time quantitative polymerase chain reaction assays were run in duplicate to measure the expression of six ISGs: interferon induced protein with tetratricopeptide repeats 1 (IFIT1), interferon α inducible protein 27 (IFI27), interferon induced protein 44 like (IFI44L), interferon stimulated genes 15 (ISG15), sialic acid binding Ig like lectin 1 (SIGLEC1), and radical S-adenosyl methionine domain containing 2 (RSAD2). The relative abundances of each target transcript was normalized to the expression level of β-Actin and OAZ. The median fold change of the six ISGs was used to create an interferon score (IS) for each individual. Samples with abnormal expressions were removed and the cDNA mix of the remaining samples was used as a calibrator to calculate the IS. We define an abnormal IS as being greater than+2 standard deviations above the mean of controls. Differences in IS between groups were compared using t-test or Mann-Whitney U-test. Results:The mean IS of controls was 1.046, standard 0.755, and the cut-off value was 2.556. A total of 25 samples from 18 interferonopathies patients were tested. The mean value was 27.010 with a 15/18 abnormality rate. Compared with the control group, IS in patients was significantly higher, t=4.247( P=0.000 1). The accuracy, precision, sensitivity, and specificity were 91.30% (42/46), 7.47%(0.084/1.124), 15/18, and 96.43% (27/28), respectively. Conclusion:This study provides a new and reliable method for clinical screening and dynamic monitoring of type Ⅰ interferonopathies by detecting ISGs expression and creating an IS.
