Real-time PCR for quantitative detection of mitochondrial DNA from peripheral blood mononuclear cell in patients with HBV-related hepatocellular carcinoma

The alteration of mitochondrial DNA (mtDNA) content could affect the expression of genes which causes many tumor diseases. However, the association between mtDNA content in peripheral blood mononuclear cell (PBMC) and hepatitis B virus (HBV)-related hepatocellular carcinoma (HCC) remains undetermined. First of all, establishing a reliable assay to detect mtDNA content is of great clinical significance. In this study, the method of real-time quantitative polymerase chain reaction (RT-qPCR) with SYBR Green I was established to evaluate mtDNA content in PBMC of healthy controls (n=23) and non-surgical HBV-related HCC cases (n=46). Receiver operating characteristic (ROC) curve analysis was carried out to assess the clinical significance of mtDNA content for diagnosing HCC. Consequently, linear range of the assay was between 1×1010 copies/μl and 1×103 copies/μl. Sensitivity was 800 copies/μl. Besides, HCC cases had a significantly lower mtDNA content than healthy controls (378.55 [58.20-784.85] vs 715.48 [292.00-1280.00]; P<0.001). When 489.90 copies/μl was set as the cut-off point, the sensitivity and specificity of mtDNA content to diagnose HCC were 82.6% and 71.7%, respectively. In conclusion, a simple, cost-effective, highly sensitive and specific method to detect mtDNA content is established. This method can be applied to clinical laboratory for detecting mtDNA content. Moreover, our study provides the first epidemiological evidence that mtDNA content in PBMC is significantly associated with HCC. mtDNA content in PBMC could serve as a novel clinical diagnostic indicator for HCC which may need more researches to validate.