OBJECTIVETo evaluate the effects of copper-phenanthroline (CuOP) on pentachlorophenol (PCP)-induced adaptation and cell death of Escherichia coli.

METHODSBacterial growth and adaptation to PCP were monitored spectrophotometrically at 600 nm. Inactivation of bacterial cells was determined from colony count on agar dishes. Cellular ATP content and accumulation of PCP were assessed by chemiluminescence and HPLC analysis respectively. The formation of PCP-Cu-OP complex was shown by UV-visible spectra.

RESULTSEscherichia coli (E. coli) could adapt to PCP, a wood preservative and insecticide used in agriculture. The adaptation of E. coli to PCP prevented its death to the synergistic cytotoxicity of CuOP plus PCP and declined cellular accumulation and uncoupling of oxidative phosphorylation of PCP. Furthermore, CuOP and PCP neither produced reactive oxygen species (ROS) nor had a synergistic effect on uncoupling of oxidative phosphorylation in E. coli. The synergistic cytotoxicity of CuOP and PCP in E. coli might be due to the formation of lipophilic PCP-Cu-OP complex.

CONCLUSIONOur data suggested that adaptation of E. coli to PCP decreased the synergistic effects of CuOP and PCP on prokaryotic cell death due to the formation of lipophilic PCP-Cu-OP complex, but it had no effect on the uncoupling of oxidative phosphorylation and production of reactive oxygen species in E. coli.

Adaptation, Physiological ; Adenosine Triphosphate ; metabolism ; Antioxidants ; metabolism ; Apoptosis ; drug effects ; Copper ; pharmacology ; Cytotoxins ; pharmacology ; Drug Resistance, Bacterial ; Drug Synergism ; Escherichia coli ; drug effects ; metabolism ; Pentachlorophenol ; pharmacology ; Phenanthrolines ; pharmacology

Objective:To explore whether CD5 + CD19 + B cells has the function of secreteing interleukin-10 (IL-10) in vitro, and to further investigate its possible effects and mechanisms on CD8 + cells in the process of hepatitis B virus (HBV) infection. Methods:From July 2017 to June 2018, at Wuxi Second People′s Hospital Affiliated to Nanjing Medical University, 23 patients with chronic hepatitis B (chronic hepatitis B group), 18 patients with liver cirrhosis (liver cirrhosis group) and 19 healthy individuals in the same period as healthy controls (healthy control group) were enrolled. Peripheral blood mononuclear cell (PBMC) were isolated and cultured. CD5 + CD19 + B cells were isolated. The cells were analyzed by flow cytometry. The ratio of high CD5 + CD19 + B cells content (>6 % of lymphocytes), the secretion of IL-10 by CD5 + CD19 + B and the ratio of high IL-10 + cells content (>4 % of lymphocytes) of three groups were compared. The effects and possible mechanisms of CD5 + CD19 + B cells on the secreting of interferon-γ (IFN-γ) by CD8 + cells were analyzed. Liver biopsy and immunohistochemistry examination were conducted in 18 patients (13 patients with chronic hepatitis B and 5 patients with liver cirrhosis) and the expression of CD5 + CD19 + B cells in human liver tissues was analyzed. Chi square test and Fisher exact probability test were used for statistical analysis. Results:The ratio of high CD5 + CD19 + B cells content of liver cirrhosis group was higher than that of healthy control group (8/18 vs. 2/19) and the difference was statistically significant (Fisher exact probability test, P=0.029). The precentage of CD5 + CD19 + B cells in healthy control group ( n=10), chronic hepatitis B group ( n=23) and liver cirrhosis group ( n=18) accounted for 81.6%, 82.3% and 70.1%of IL-10 + cells, respectively, and the number of patients with high IL-10 + cells precentage was 2, 7 and 2, respectively. There were no statistically significant differences among three groups (all P>0.05). After stimulated with lipopolysaccharide and cultured for 48 hours, the precentage of CD8 + IFN-γ + cells in lymphocytes of healthy control group ( n=10), chronic hepatitis B group ( n=10) and liver cirrhosis group ( n=10) were compared, and the differences were not statistically significant (all P>0.05). After CD5 + CD19 + B cells were eliminated, the precentage of CD8 + IFN-γ + cells in lymphocytes increased in 5, 4 and 4 patients of healthy control group ( n=10), chronic hepatitis B group ( n=10) and liver cirrhosis group ( n=10). After adding IL-10 receptor blocker, the precentage of CD8 + IFN-γ + cells in lymphocytes in PBMC increased compared with that before the addition of IL-10 receptor blocker (7.23% vs. 6.87%). The results of immunohistochemistry examination of liver biopsy indicated that CD4 + and CD8 + cells were strong expressed in portal area of liver tissue of patients, while CD5 + and CD19 + were less expressed. Conclusions:CD5 + CD19 + B cells do not show obvious quantitative and functional differences in the process of chronic HBV infection, however the ability of CD8 + cells to secrete IFN-γ, which may be achieved by secreting IL-10 rather than by direct contact between cells.