Objective To investigate the mechanism of fluoroquinolone resistance in clinical isolates of Enterococcus faecium. Methods The MICs of six fluoroquinolones(norfloxacin,ciprofloxacin,ofloxacin,levofloxacin,gatifloxacin and moxifloxacin) against 35 clinical isolates of E.faecium from eight hospitals in Tianjin were determined by agar dilution method in the absence or presence of multidrug resistance efflux pump inhibitor reserpine.The quinolone-resistance determining region(QRDR)of parC and gyrA were amplified and sequenced.Results No less than twofold decrease in MIC values of the six fluoroquinolones in the presence of reserpine was observed in 35,29,1,0,6 and 2 of the 35 strains of E.faecium respectively.One fluoro- quinolone-susceptible isolate and five fluoroquinolone-resistant isolates were selected randomly to analyze the QRDR of parC and gyrA.All five fluoroquinolone-resistant isolates had single amino acid alteration in both GyrA and ParC.Ser-80 in ParC was substituted by lie(4 isolates)or Arg(1 isolates).Glu-87 in GyrA was replaced by Lys(2 isolates)or Gly(2 isolates). The other one had an Ser-83-to-Ile substitution.The one fluoroquinolone-suseeptible isolate had no alteration in the QRDR of either ParC or GyrA.Conclusions Both target alteration and active efflux are responsible for the resistance to fluoroquinolone in clinical isolates of E.faecium.

Objective To compare the difference of transformation profile and transformation rate of tecomin by using two in vitro liver metabolism models. Methods Liver microsomes and liver S9 fraction models were employed to transform tecomin. HPLC was used to determine the contents of tecomin and its metabolites at the detecting wavelength of 254 nm. The gradient elution (0–6 min, 5％–40％ A; 6–9 min, 40％–50％ A; 9–11 min, 50％–5％ A) was carried out by using mobile phase of acetonitrile (A) - 1％ acetic acid (B) at a flow rate of 1 mL/min. Results Both models could transform tecomin into veratric acid; however, the metabolites obtained with liver S9 were more than those obtained with liver microsomes, and the transformation rate of the former was higher than that of the latter. Conclusion The liver S9 fraction can more efficiently transform esters than liver microsomes.

OBJECTIVETo observe the hepatitis B virus (HBV) replication in the tree shrews that were inoculated with HBV at neonatal period.

METHODSSix new-born tree shrews were inoculated with human HBV positive serum. Blood samples and liver biopsies were collected at different time points after inoculation. The HBV infection markers were tested by nested polymerase chain reaction (nPCR), fluorescence quantitative polymerase chain reaction (FQ-PCR), Southern blot, ELISA and immunohistochemistry staining. The liver tissues were observed under electron and light microscope.

RESULTS48 weeks after inoculation, HBV DNA and HBV cccDNA were detected in the serum and liver samples of three animals (number 1, 2 and 6) by nPCR. The copy-numbers of HBV DNA detected by FQ-PCR in their serum and liver samples were 103 and-104/ml respectively,and the total DNA in 1microg liver tissue was 107-108. Southern blot indicated that HBV replication intermediates such as HBV cccDNA and HBV ssDNA was detectable in liver tissues. HBsAg was detected by ELISA, and immunohistochemical staining showed a gradual increase of HBsAg-positive liver cells. High copy number of HBV DNA and suspected HBV EM particles could be detected in the liver samples from one of the three animals that have survived more than 2 years after inoculation. The other three animals showed low HBV DNA copy number, and the rest of the signs of HBV infection were negative or transiently positive.

CONCLUSIONSNeonatal tree shrews can be infected with human HBV. HBV can replicate inside the liver cells of tree shrew.

Animals ; Animals, Newborn ; Biopsy ; DNA, Circular ; analysis ; blood ; DNA, Viral ; analysis ; blood ; Disease Models, Animal ; Hepatitis B ; etiology ; pathology ; virology ; Hepatitis B Surface Antigens ; blood ; Hepatitis B virus ; genetics ; physiology ; Humans ; Immunohistochemistry ; Liver ; pathology ; virology ; Polymerase Chain Reaction ; methods ; Tupaiidae ; Virus Replication

Objective To evaluate the effects of small interfering RNA(siRNA)against Ki67 gene on the proliferation and apoptosis of human renal carcinoma cell line 786-0 cells.Methods The human renal carcinoma 786-0 cells were treated with Ki67-siRNA(100 nmol/L).The mRNA expression of Ki67 was detected by RT-PCR.The protein expression of Ki67 was detected by Western blot and immunohisto- chemical technique,respectively.The proliferation of 786-0 cells was detected by MTT assay.The apoptosis of 786-0 cells was detected by TUNEL assay.Results RT-PCR and Western blot analysis showed that the Ki67 mRNA and Ki67 protein expression levels of the 786-0 cells treated with Ki67-siRNA were(37.6?1.9)% and(46.4?0.9)% ,respectively,which were significantly lower than those of controls [(97.3?0.9)% and(95.3?0.9)%,P＜0.01],The Ki67 positive expression rate of 786-0 cells treated with Ki67-siRNA by immunohistochemical technique was 52.5?2.3,which was significantly lower than that of controls(114.5?4.9 ,P＜0.01).The proliferation-inhibiting rate and apoptosis rate of the 786-0 cells trea- ted with Ki67-siRNA were( 63.6?1.6)% and(41.7?0.6)% ,respectively,which were significantly higher than those of controls [(2.8?0.2)% and(10.3?1.4)%,P＜0.01].Conclusions siRNA against Ki67 gene can inhibit the proliferation and induce the apoptosis by blocking Ki67 expression of hu- man renal carcinoma 786-0 cells.The inhibition of Ki67 expression by siRNA may be a promising approach in gene therapy for renal cancer.

OBJECTIVETo investigate the impact of staghorn stone branch number on outcomes of percutaneous nephrolithotomy (PNL).

METHODSFrom January 2009 to January 2013, the 371 patients with staghorn stones who were referred to our hospital for PNL were considered for this study. All calculi were showed with CT 3-dimentional reconstruction (3-DR) imaging. The computerized database of the patients had been reviewed. Our exclusion criterion was patients with congenital renal anomalies, such as horse-shoe and ectopic kidneys. And borderline stones that branched to one major calyx only were also not included. From 3-DR images, the number of stone branching into minor renal calices was recorded. We made "3" as the branch breakdown between groups. And the patients were divided into four groups. The number of percutaneous tract, operative time, staged PNL, intra-operative blood loss, complications, stone clearance rate, and postoperative hospital day were compared.

RESULTSThe 371 patients (386 renal units) underwent PNL successfully, included 144 single-tract PNL, 242 multi-tract PNL, 97 staged PNL. The average operative time was (100 ± 50) minutes; the average intra-operative blood loss was (83 ± 67) ml. The stone clearance rate were 61.7% (3 days) and 79.5% (3 months). The postoperative hospital stay was (6.9 ± 3.4) days. A significantly higher ratio of multi-tract (χ(2) = 212.220, P < 0.01) and staged PNL (χ(2) = 49.679, P < 0.01), longer operative time (F = 4.652, P < 0.01) and postoperative hospital day (F = 2.067, P = 0.043) and lower rate of stone clearance (χ(2) = 10.691 and 47.369, P < 0.05) were found in PNL for calculi with stone branch number ≥ 5. There was no statistically meaningful difference among the 4 groups based on Clavien complication system (P = 0.460).

CONCLUSIONThe possibility of multi-tract and staged PNL, lower rate of stone clearance and longer postoperative hospital day increase for staghorn calculi with stone branch number more than 5.

Adult ; Aged ; Female ; Humans ; Kidney Calculi ; pathology ; surgery ; Length of Stay ; Male ; Middle Aged ; Nephrostomy, Percutaneous ; Operative Time ; Retrospective Studies ; Treatment Outcome

The present study was designed to determine the intestinal bacterial metabolites of trollioside and isoquercetin and their antibacterial activities. A systematic in vitro biotransformation investigation on trollioside and isoquercetin, including metabolite identification, metabolic pathway deduction, and time course, was accomplished using a human intestinal bacterial model. The metabolites were analyzed and identified by HPLC and HPLC-MS. The antibacterial activities of trollioside, isoquercetin, and their metabolites were evaluated using the broth microdilution method with berberine as a positive control, and their potency was measured as minimal inhibitory concentration (MIC). Our results indicated that trollioside and isoquercetin were metabolized by human intestinal flora through O-deglycosylation, yielding aglycones proglobeflowery acid and quercetin, respectively The antibacterial activities of both metabolites were more potent than that of their parent compounds. In conclusion, trollioside and isoquercetin are totally and rapidly transformed by human intestinal bacteria in vitro and the transformation favors the improvement of the antibacterial activities of the parent compounds.
Activation, Metabolic ; Anti-Bacterial Agents ; metabolism ; Bacteria ; metabolism ; Benzoates ; metabolism ; Biotransformation ; Gastrointestinal Microbiome ; Glucosides ; metabolism ; Humans ; Intestines ; microbiology ; Microbial Sensitivity Tests ; Models, Biological ; Quercetin ; analogs & derivatives ; metabolism

This study was performed to determine the metabolic profile of a new illicit drug, PX-2, in human liver microsomes. Q Exactive™ HF Quadrupole-Orbitrap LC-MS (LC-QE-HF-Orbitrap-MS) was employed to determine the metabolic sites and pathways of phase Ⅰ and phase II metabolism. PX-2 was added to a microsomal incubation model to simulate human hepatic metabolism. The results showed that a total of 18 phase Ⅰ metabolites and 3 glucuronidated phase II metabolites were generated, with the main metabolic pathways of phase Ⅰ metabolism including amide hydrolysis, fluoropentyl oxidative defluorination, benzyl hydroxylation, and carbazole ring hydroxylation. Based on the type and sites of metabolism, phase Ⅰ metabolites M1.1 (amide hydrolysis), M4.1 (carbazole cyclic hydroxylation), and M3.1 (oxidative defluorinative hydroxylation) are proposed to be potential poisoning markers. The results of this study provide a basis for identification of related drugs and establishment of testing methods in biological samples.

ObjectiveTo explore the therapeutic effect and possible mechanism of Banxia Xiexintang and its disassembled prescriptions in regulating the flora disorder induced by mixed antibiotics in young rats. MethodSeventy male BALB/C young rats were randomly assigned into 7 groups： blank group， model group， Bifidobacterium tetralogy viable tablets （0.68 g·kg^-1） group， Banxia Xiexintang （9.1 g·kg^-1） group， Xinkai （3.19 g·kg^-1） group， Kujiang （1.82 g·kg^-1） group， and Ganbu （4.1 g·kg^-1） group， with 10 rats in each group. Except the blank group， the other groups were given mixed antibiotics by gavage to induce intestinal flora disorder. After 14 days， the rats in different drug groups were administrated with corresponding drugs by gavage， and those in the blank group and model group with the same amount of normal saline once a day for 14 days. After that， fecal samples were collected aseptically for 16S rDNA sequencing of intestinal flora， and lipopolysaccharide （LPS， 10 mg·kg^-1） was injected intraperitoneally to induce inflammatory reaction. The tissue morphology of colonic mucosa was observed via hematoxylin-eosin （HE） staining， and the macrophage infiltration of colonic mucosa was observed via toluidine blue staining and immunohistochemistry. The expression of interleukin-1β （IL-1β）， interleukin-6 （IL-6）， interleukin-8 （IL-8）， tumor necrosis factor-α （TNF-α）， and interleukin-10 （IL-10） mRNA were determined by real-time fluorescence quantitative polymerase chain reaction （Real-time PCR）. ResultCompared with the blank group， the modeling changed the intestinal flora structure of the young rats （P<0.01）， damaged the colonic mucosa， reduced the macrophage infiltration， and down-regulated the mRNA levels of IL-1β， IL-6， IL-8， TNF-α， and IL-10 （P<0.01）. Compared with the model group， bifidobacterium quadruple viable tablets， Banxia Xiexintang and its disassembled prescriptions increased the diversity of intestinal flora and the relative abundance of beneficial bacteria such as Bacteroidetes and Firmicutes （P<0.01）. At the same time， they ameliorated colonic mucosal injury （P<0.05， P<0.01）， increased macrophage infiltration （P<0.05， P<0.01）， and up-regulated the mRNA levels of IL-6， IL-8， and TNF-α （P<0.01）. The mRNA level of IL-1β was up-regulated in Bifidobacterium tetralogy viable tablets， Banxia Xiexintang， Kujiang， and Ganbu groups （P<0.01）， and that of IL-10 was up-regulated in Bifidobacterium tetralogy viable tablets， Banxia Xiexintang， Xinkai， and Ganbu groups （P<0.01）. ConclusionBanxia Xiexintang and the disassembled prescriptions can adjust the intestinal flora of young rats exposed to antibiotics and protect the immune barrier of colonic mucosa after intestinal flora disorder. In particularly， the whole prescription of Banxia Xiexintang demonstrates the best performance.

Objective: To investigate the association between the urinary arsenic level and serum total testosterone in Chinese men aged 18 to 79 years. Methods: A total of 5 048 male participants aged 18 to 79 years were recruited from the China National Human Biomonitoring (CNHBM) from 2017 to 2018. Questionnaires and physical examinations were used to collect information on demographic characteristics, lifestyle, food intake frequency and health status. Venous blood and urine samples were collected to detect the level of serum total testosterone, urinary arsenic and urinary creatinine. Participants were divided into three groups (low, middle, and high) based on the tertiles of creatinine-adjusted urinary arsenic concentration. Weighted multiple linear regression was fitted to analyze the association of urinary arsenic with serum total testosterone. Results: The weighted average age of 5 048 Chinese men was (46.72±0.40) years. Geometric mean concentration (95%CI) of urinary arsenic, creatinine-adjusted urinary arsenic and serum testosterone was 22.46 (20.08, 25.12) μg/L, 19.36 (16.92, 22.15) μg/g·Cr and 18.13 (17.42, 18.85) nmol/L, respectively. After controlling for covariates, compared with the low-level urinary arsenic group, the testosterone level of the participants in the middle-level group and the high-level group decreased gradually. The percentile ratio (95%CI) was -5.17% (-13.14%, 3.54%) and -10.33% (-15.68%, -4.63). The subgroup analysis showed that the association between the urinary arsenic level and testosterone level was more obvious in the group with BMI<24 kg/m² group (P_interaction=0.023). Conclusion: There is a negative association between the urinary arsenic level and serum total testosterone in Chinese men aged 18 to 79 years.
Humans ; Male ; Arsenic/urine* ; Creatinine ; East Asian People ; Testosterone/blood* ; Urinalysis ; Adolescent ; Young Adult ; Adult ; Middle Aged ; Aged