[The correlation between expression level of femB and resistance phenotype of methicillin-resistant Staphylococcus aureus (MRSA)].

OBJECTIVE: To investigate the correlation between the phenotype and expression level of femB of Staphylococcus aureus (MRSA), to discuss the mechanism of different phenotypes in methicillin-resistant Staphylococcus aureus. METHODS: The minimal inhibitory concentrations (MICs) of oxacillin against 71 clinical isolates of Staphylococcus aureus were determined by agar dilution method according to NCCLS. The production of beta-lactamase was identified by Cefinase paper strip method. The isolation rate of beta-lactamase-producing strains was counted and the correlation between the resistance phenotype and isolation rate of beta-lactamase was analysed by statistics. Real time fluorescent quantitative PCR was used to quantify the mRNA expression of femB of non-beta-lactamase-producing strains. RESULTS: The resistance rate of 71 Staphylococcus aureus to oxacillin was 66.20% (47/71), the isolation rate of beta-lactamase-producing MSSA strains was 58.3%,and that of strains of high- and low-level resistance to oxacillin were 63.15% and 55.56%. The standard curve was performed by series dilution of the heterogeneous resistant strain BB270, and the amount of femB-specific mRNA in strain BB270 was set to be 1. The calculated femB amounts in MSSA strains were from 0.4830-3.3636, while the amounts were from 0.4204-3.3636 in low-level MRSA strains, and 0.0718-16.0000 in high-level MRSA strains. There were no difference in the level of femB among MSSA, high-level MRSA and low-level MRSA. CONCLUSION: The expression level of femB may not be related to the resistance of non-beta-lactamase-producting Staphylococcus aureus to methicillin.

[Study on the correlation between resistance phenotype and expression level of femA of Staphylococcus aureus].

OBJECTIVE: To detect the expression level of femA of Staphylococcus aureus strains with different phenotype. METHODS: 15 strains of the non-beta-lactamase-producing clinical isolates with different phenotype by agar dilution and by nitrocephin paper strip method were chosen as the object of test, in addition to 4 donative strains (BB270, BB308, BB586, COL). Total RNA were extracted and analysed by agarose gel electrophoresis. Real time fluorescent quantitative PCR was performed to quantify the expression of femA gene. The expression level of femA of BB270 was set to be standard(100%). RESULTS: The expressions of femA were observed in all the tested strains. The amount of femA-specific mRNA in the mutant strain BB308 was approximately 37.82% and that of stain BB586 was 240.50%, homogeneous resistant strain COL was 862.61%. The amounts in MSSA strains were from 0.00353% to 29.92%, that in low-level MRSA strains were from 0.00554% to 310%, otherwise that in high-level MRSA strains were from 13.88% to 55000%, which were different among these groups. There was no significant difference in amount of femA-mRNA between MSSA and low-level MRSA strains (P1 = 0.83) but marked between high-level MRSA and low-level MRSA/MSSA strains (P2 = 0.006, P3 = 0.01)). CONCLUSION: Expression level of femA in high-level MRSA was significant higher than that in low-level MRSA and MSSA. femA was essential for the expression of high-level methicillin resistance in MRSA.

Inhibitory effect of silymarin on CCl4-induced liver fibrosis by reducing Ly6Chi monocytes infiltration.

It has been well established that silymarin has hepatoprotective and anti-fibrotic effects. But the mechanisms are poorly understood. In recent years, the role of Ly6Chi monocytes in liver fibrosis has been well demonstrated. Thus, in present study we aimed to investigate whether silymarin can relieve liver fibrosis by reducing Ly6Chi monocytes infiltration. The mouse model of liver fibrosis was established by injected with carbon tetrachloride (CCl4) via intraperitoneal repeatedly. Mice in silymarin group received silymarin treatment by gavage. Silymarin significantly reduced liver inflammation and fibrosis of the mice induced by CCl4 injection, as revealed by liver histological and pathological analysis. Mice administrated by silymarin exhibited less infiltration of Ly6Chi monocytes. But there was no difference on other tested leukocyte subsets between CCl4 group and silymarin group. Meanwhile, further study found that silymarin significantly reduced CCl4-induced increased expression of tumor necrosis factor (TNF)-α, transforming growth factor (TGF)-ß1 and monocyte chemoattractant protein 1 (MCP-1), which was in line with the decreased numbers of intrahepatic Ly6Chi monocytes. In conclusion, our study showed that the anti-inflammatory and anti-fibrotic effects of silymarin could be contributed to the prevention of Ly6Chi monocytes infiltration into the injured livers, which will give us a better understanding on the cellular mechanism of hepatoprotective and anti-fibrotic effect for silymarin.

[A study on the mechanism of drug resistance in Staphylococcus aureus].

OBJECTIVE: To inquire into the mechanism of drug resistance in Staphylococcus aureus. METHODS: A total of 198 strains of Staphylococcus aureus were isolated from the samples sent to the Clinical Laboratory of Microbiology,West China Hospital. The resistance of Staphylococcus aureus to Methicillin was assayed with agar dilution. Staphylococcus aureus mecA gene was measured by PCR assay and beta-lactamase was detected by Nitrocephin. RESULTS: The rate of resistance to methicillin was 64.65% in 198 strains of Staphylococcus aureus; 118 strains of methicillin-resistant staphylococcus aureus(MRSA) were found to have high level resistance in 128 MRSA;10 strains of MRSA were found to have low level resistance; 41(58.57%) strains of methicillin-sensitive Staphylococcus aureus (MSSA) expressed beta-lactamase; 2 Staphylococcus aureus had mecA among them; 67 Staphylococcus aureus expressed beta-lactamase in high level resistance, 63(53.39%)Staphylococcus aureus expressed beta-lactamase in high level resistance, among them, 5 Staphylococcus aureus had mecA; 40.00% MRSA expressed beta-lactamase in low level resistance, 55 MRSA did not express beta-lactamase in high level resistance, which had all mecA; 9 Staphylococcus aureus did not express beta-lactamase in low level resistance, among them, 5 Staphylococcus aureus had mecA. The difference in expression of beta-lactamase was statistically significant between MSSA and MRSA; MRSA(53.39%) was lower than MSSA (58.57%); the other differences were not significant. The difference in having mecA was statistically significant between MRSA(having high resistant level and no expression of beta-lactamase) and the others; MRSA had higher mecA than did the others. CONCLUSION: The resistance in Staphylococcus aureus mainly involved two mechanisms: the expression of beta-lactamase and the expression of mecA.

Protective effects of curcumin against liver fibrosis through modulating DNA methylation.

Recent research has demonstrated that advanced liver fibrosis in patients could be reversed, but no approved agents are available for the treatment and prevention of liver fibrosis in humans. Curcumin (CUR) is the principal curcuminoid of turmeric. Inhibitory effects of CUR and its underlying mechanisms in liver fibrogenesis have been explored. In the present study, we hypothesized that epigenetic mechanisms contribute to the protective effects of CUR against liver fibrosis. We used CCl4-induced liver injury in BALB/c mice and the rat hepatic stellate cell line HSC-T6 as experimental models. Genomic DNA methylation was analyzed by MeDIP-chip and verified by real-time PCR on MeDIP-enriched DNA. The mRNA and protein expressions of DNMT1, α-SMA, and Col1α1 were determined by real-time PCR and Western blotting, respectively. The methylation statuses of FGFR3, FZD10, Gpx4, and Hoxd3 were further confirmed by quantitative methylation-specific PCR (qMSP). Our results showed that CUR treatment reversed liver injury in vivo and in vitro, possibly through down regulation of DNMT1, α-SMA, and Col1α1 and by demethylation of the key genes. In conclusion, aberrant methylation is closely associated with liver fibrosis and CUR treatment may reverse liver fibrosis by epigenetic mechanisms.

Association of serum gamma-glutamyl transferase with treatment outcome in chronic hepatitis B patients.

AIM: To investigate the association of serum gamma-glutamyl transferase (GGT) levels with chronic hepatitis B infection and hepatitis B e antigen (HBeAg) seroconversion. METHODS: A retrospective study was performed on clinical data collected from patients who had been positive for hepatitis B surface antigen for > 6 mo and who were antiviral-treatment naïve (n = 215) attending the Hepatitis Clinic at Nanjing Drum Tower Hospital between August 2010 and December 2013. Healthy individuals without liver disease (n = 83) were included as controls. Patients were categorized into four groups based on disease status as recommended by the European Association for the Study of the Liver: immune tolerance (IT; n = 47), HBeAg-positive hepatitis (EPH; n = 93), HBeAg-negative hepatitis (ENH; n = 20), and inactive carrier (IC; n = 55). Prediction of complete response (CR) based on serum GGT was also examined in EPH patients (n = 33) treated for 48 wk with nucleos(t)ide analogue (NA) therapy, including lamivudine plus adefovir combination therapy (n = 20) or entecavir monotherapy (n = 13). CR was defined as a serum hepatitis B virus DNA level < 500 copies/mL and HBeAg seroconversion by 48 wk of treatment. RESULTS: Serum GGT levels were significantly increased in EPH and ENH patients relative to the IT, IC, and healthy control groups (P < 0.01 for all). However, no significant difference in serum GGT levels was found between the EPH and ENH groups. Baseline serum GGT levels were significantly higher in patients who achieved CR (7/33; 21.2%) compared to patients in the non-CR group (26/33; 78.8%; P = 0.011). In addition, the decline in serum GGT was greater in CR patients compared to non-CR patients after 24 wk and 48 wk of treatment (P = 0.012 and P = 0.008, respectively). The receiver operating characteristic curve yielded a sensitivity of 85.71% and a specificity of 61.54% at a threshold value of 0.89 times the upper limit of normal for baseline serum GGT in the prediction of CR following NA therapy. CONCLUSION: Serum GGT is significantly elevated in EPH and ENH patients and is a potential biomarker for the prediction of HBeAg seroconversion following NA therapy.