The Clinical Significance Of Serum And Peripheral Blood Mononuclear Cells HBVcccDNA Detection In Patients With Chronic Hepatitis B

Objective: 1.To determine cccDNA whether presence in the serum of patients with chronic Hepatitis B and investigate the relative factors and clinical significance 2. To determine cccDNA whether exists in peripheral blood monouclear cells of patients with chronic Hepatitis B and to discussion of potential clinical significance .Methods:103 patients chronically infected with B virus (including 8 transplantation of liver patients)were investigated ,HBV-DNA in the serum and peripheral blood monouclear cells was assessed by Real time PCR. To reduce miscellaneous rcDNA and line double strands DNA in the sample, cccDNA was extracted by an alkaline lysis method; To purify cccDNA further,the extraction product was digested by mung-bean nuclease enzyme to remove the residual rcDNA . Designing a couple of two span nick primer to avoid line double strands DNA amplification spell false positive; Using synergy PCR technique to improve PCR assay sensitivity.Results:103 patients serum cccDNA positive ratio was 41.7%. The groups with different HBV-DNA load or ALT level in serum were compared, positive ratio of cccDNA had a significant difference (P﹤0.05);The groups onset in different period were compared, potive ratio of cccDNA also had a significant difference (P﹤0.05). Positive ratio of HBV-DNA in the peripheral blood monouclear cells was 45.6% . Groups with different HBV-DNA load in serum were compared, potive ratio of HBV-DNA in the peripheral blood monouclear cells had a significant difference (P﹤0.05). Potive patients in different groups,the HBV-DNA load in the serum and peripheral blood monouclear cells had a significant difference (P﹤0.05). cccDNA was not detected in the peripheral blood monouclear cells of 43 chronic HBV patients (including 8 transplantation of liver patients)Conclusions: 1.We have found cccDNA in serum , cccDNA in serum maybe derived from hepatic cell necrosis. 2.cccDNA positive ratio in serum is increasing with high HBV-DNA load or high level ALT; cccDNA in serum positive ratio is reduced with onset time 3. We have detected HBV-DNA in the peripheral blood monouclear cells of patient with chronic Hepatitis B ,but haven't detected cccDNA in it 4. The more HBV-DNA load in serum accompany with the more HBV-DNA load in the peripheral blood monouclear cells,HBV in the peripheral blood monouclear cells maybe derived from serum. 5.The study can not confirm HBV replication in the peripheral blood monouclear cells.