Detection of APRIL mRNA in peripheral blood of auto-immune diseases using a quantitative real-time PCR method / 临床检验杂志

ABSTRACT

Objective To establish a SYBR Green Ⅰ quantitative real-time PCR method for detecting the expression of APRIL gene(a proliferation-inducing ligand,APRIL) in peripheral blood of patients with auto-immune diseases,e.g.,systemic lupus erythematosus(SLE) and rheumatoid arthritis(RA),and investigate the relationship of APRIL mRNA expression with pathogenesis and prognosis of auto-immune diseases.Methods Plasmid PGEM-T easy-APRIL was cloned as the standard template.SYBR Green Ⅰ quantitative real-time PCR was set up to examine the expression of APRIL mRNA in peripheral blood of 58 patients with auto-immune diseases and 20 healthy controls using Line Gene FQD-33A Detection System.Results The obtained data were normalized by dividing the copy number of target cDNA by those of GAPDH(glyceraldehycle-3-phosphate dehydrogenase,GAPDH).APRIL expression levels ranges from 3.95 to 192 and mean value was 29.68?4.5.APRIL expression of twenty healthy controls showed range from 3.1 to 18.7 and mean value of 10.56?2.0.APRIL expression levels in patients with auto-immune diseases were higher than those in healthy controls.In auto-immune diseases group APRIL expression levels of untreated patients were higher than those of the other patients,and a statistical significance was found.Conclusions APRIL mRNA was successfully detected by SYBR Green Ⅰ quantitative real-time PCR and the method was accurate and reliable.The expression of APRIL mRNA of auto-immune disease patients was higher than those of healthy controls,and the expression of untreated patients was the highest.This method may be used for further study on the high-level expression of APRIL mRNA in mechanism of auto-immune diseases as well as the development and prognosis of diseases.