Interaction between Brucella melitensis 16M and small ubiquitin-related modifier 1 and E2 conjugating enzyme 9 in mouse RAW264.7 macrophages

Brucella is an intracellular pathogen that invades a host and settles in its immune cells; however, the mechanism of its intracellular survival is unclear. Modification of small ubiquitin-related modifier (SUMO) occurs in many cellular activities. E2 conjugating enzyme 9 (Ubc9) is the only reported ubiquitin-conjugating enzyme that links the SUMO molecule with a target protein. Brucella's intracellular survival mechanism has not been studied with respect to SUMO-related proteins and Ubc9. Therefore, to investigate the relationship between Brucella melitensis 16M and SUMO, we constructed plasmids and cells lines suitable for overexpression and knockdown of SUMO1 and Ubc9 genes. Brucella 16M activated SUMO1/Ubc9 expression in a time-dependent manner, and Brucella 16M intracellular survival was inhibited by SUMO1/Ubc9 overexpression and promoted by SUMO1/Ubc9 depletion. In macrophages, Brucella 16M-dependent apoptosis and immune factors were induced by SUMO1/Ubc9 overexpression and restricted by SUMO1/Ubc9 depletion. We noted no effect on the expressions of SUMO1 and Ubc9 in B. melitensis 16M lipopolysaccharide-prestimulated mouse RAW264.7 macrophages. Additionally, intracellular survival of the 16M△VirB2 mutant was lower than that of Brucella 16M (p < 0.05). VirB2 can affect expression levels of Ubc9, thereby increasing intracellular survival of Brucella in macrophages at the late stage of infection. Collectively, our results demonstrate that B. melitensis 16M may use the VirB IV secretion system of Brucella to interact with SUMO-related proteins during infection of host cells, which interferes with SUMO function and promotes pathogen survival in host cells.

Expression and clinical significance of liver tissue HBVcccDNA and serum markers for chronic hepatitis B antiviral treatment / 中国基层医药

Objective To investigate the expression and clinical significance of liver tissue hepatitis B virus covalently closed circular DNA ( HBV cccDNA) and three serum markers ( including serum HBV DNA, HBsAg, HBeAg) for chronic hepatitis B antiviral treatment.Methods 80 patients with chronic hepatitis B were collected, they were treated with telbivudine tablets,and the liver tissue HBVcccDNA and serum HBV DNA expression were detected by fluorescence quantitative PCR( FQ-PCR) .Serum HBsAg,HBeAg expressions were detected by enzyme linked immunosorbent assay ( ELISA) before treatment,12 weeks,28 weeks,44 weeks,52 weeks after treatment. Then,the correlation of liver tissue HBVcccDNA with serum markers was analyzed.Results Liver tissue HBVcccD-NA,serum DNA HBV,serum HBsAg and serum HBeAg were significantly decreased with the time of treatment,the differences were statistically significant(F=3.786,4.785,3.806,3.452,P=0.034,0.009,0.031,0.042),and the liver tissue HBVcccDNA, serum DNA HBV and serum HBeAg before treatment compared with after treatment had statistically significant differences(all P<0.05).And the differences of serum HBsAg in the treatment of 44 and 52 weeks compared with before treatment were statistically significant(all P<0.05).The results of correlation analy-sis showed that the expression of HBVcccDNA was positively correlated with serum HBV DNA and HBeAg ( r =0.674,0.672,P=0.015,0.036),and had no significant correlation with serum HBsAg expression(r=0.125,P=0.142 ) .Conclusion The expressions of serum HBV DNA and HBeAg could reflect HBVcccDNA expression of liver tissue,and the detection method is simple and non-invasive,which is worthy of recommendation in clinical.