Objective To investigate the association of AcrAB-tolC efflux pump and marA-soxS-rob regulon with ciprofloxacin-resistance in Shigella flexneri isolates.Methods Forty five strains of Shigella flexneri were isolated in stool samples collected from outpatient diarrhea clinics in Tianjin from 2009 to 2013.The antimicrobial susceptibility of bacteria was analyzed by Kirby-Bauer disk diffusion method.Ten ciprofloxacin-resistant Shigella flexneri isolates and 10 ciprofloxacin-sensitive isolates were randomly selected.The gyrA and parC genes,plasmid mediated quinolone resistant (PMQR) determinants,efflux pump genes (acrA,acrB) and regulation genes (marA,soxS,rob) were screened and sequenced.Minimum inhibition concentrations (MICs) of the strains were determined before and after efflux pump inhibitor carbonyl cyanide m-chlorophenylhydrazone (CCCP) was added.The expression of acrA,acrB,marA,soxS and rob genes in ciprofloxacin-resistant and ciprofloxacin-sensitive strains was detected by realtime fluorescence quantitative RCR (RT-PCR),and the differences of expression were evaluated using t test.Results Both gyrA and parC mutations were detected in all ciprofloxacin-resistant strains; qnrS1 was positive in ciprofloxacin-resistant strain CR2 and aac(6')-Ib-cr was positive in ciprofloxacin-resistant strain CR5.Efflux pump genes and regulation genes were not detected in ciprofloxacin-sensitive strains,while soxRS mutation was detected in all ciprofloxacin-resistant strains except CR10.MICs of quinolones in ciprofloxacin-resistant strains decreased to one-fourth or one-eighth when CCCP added,while not changed or only decreased to one-half in ciprofloxacin-sensitive strains.Expressions of acrA,acrB,marA and soxS were significantly higher in ciprofloxacin-resistant strains than those in ciprofloxacin-sensitive strains (P ＜ 0.05 or P ＜ 0.01).Conclusion Efflux pump may involve in the high-level ciprofloxacin resistance in Shigella flexneri through activating transcription of efflux pump genes by marA-soxS-rob regulon,in which soxRS mutation may play an important role.

Objective To study the biological exposure limit in bone metabolism damage caused by coexposure to fluorine and arsenic from coal burning in exposed population.Methods One hundred and ninety-eight cases of fluoride and arsenic co-exposed people from Liuchang village,Qinzhen city,Guizhou province were enrolled in the study.Urinary fluorine (UF),urinary arsenic (UAs),urinary hydroxyproline (UHYP),ross-linked Ntelopeptides of type Ⅰ collagen(UNTX) and bone strength index(STI) were detected.BMDS Version 2.1 software was used to calculate UF,UAs benchmark dose (BMD) and its lower confidence limit (BMDL) on the damage of bone metabolism caused by co-exposure to fluorine and arsenic from coal burning.Results The BMD and BMDL range of UF caused by co-exposure to fluorine and arsenic from coal burning were 0.68-1.35 mg/g Cr,0.57-1.11 mg/g Cr.The BMD and BMDL range of UAs caused by co-exposure to fluorine and arsenic from coal burning were 8.36-18.77 μg/g Cr,7.12-15.40 μg/g Cr.Conclusion The biological exposure limits of UF and UAs for bone metabolism toxicity are proposed as 0.57 mg/g Cr and 7.12 μg/g Cr in co-exposure to fluoride and arsenic from coal burning,respectively.

N-Benzyl matrinol was obtained by hydrolysis, benzylation and reduction reaction from matrine. A series of hybrids (8a-8n) from (phenylsulfonyl)furoxan and N-benzyl matrinol were synthesized and biologically evaluated as anti-hepatocellular carcinoma agents. All target compounds were evaluated for anti-proliferative activity against human hepatocellular Bel-7402, SMMC-7721, Bel-7404, and HepG2 cells in vitro by MTT method. The results indicated that all of these compounds had potent anti-proliferative activity which were more potent than their parent compound and 5-FU, especially 8a-8h and 8j showed the strongest anti-HCC HepG2 cell activity with IC50 values of 0.12-0.93 μmol x L(-1).
Antineoplastic Agents ; pharmacology ; Carcinoma, Hepatocellular ; Fluorouracil ; Hep G2 Cells ; Humans ; Liver Neoplasms ; Oxadiazoles ; pharmacology

Objective To investigate the role of AcrAB-TolC efflux pump in fluoroquinolones resistance by Shigella. spp and to explore the significance of AcrAB-TolC efflux pump on mutation of acrR, soxS and marOR as well as on drug re?sistence. Methods Drug resistant bacteria were selected by Kirby-Bauer disk diffusion test. After addition of efflux pump inhibitor carbonylcyanide-m-chlorophenylhydrazone (CCCP), change of minimal inhibitory concentration (MIC)s of nilidixic acid, Levofloxacin, ofloxacin, ciprofloxacin and Norfloxacin were examined. The DNA binding region of acrA, acrB, soxS, acrR and marOR gene in these mutants were amplified by PCR and sequenced. Results Among the 159 clinical isolates of Shigella,11 strains are resistant to fluoroquinolone. After the addition of CCCP, MICs of 2 fluoroquinolone resistant strains decreased; the MICs of 7 fluoroquinolone resistant strains did not change; MICs of 2 fluoroquinolone resistant strains in?creased. The corresponding nucleotides C, A, T, T on the 36th to 39th of marOR gene were missing, showing by sequencing, in fluoroquinolone resistent strains which might be regulated by the efflux pump gene AcrAB-TolC. Conclusion Efflux pump inhibitor could restrain the activity of efflux partially. The mutations of marOR might play an important role in fluoroquino?lone resistent by shigella.

Objective To explore carrying rates of class 1, class 2 integrons as well as ISCR1 in Shigella isolates and their connection with drug resistance. Methods Antibiotic sensitivities were detected by K-B disk diffusion in 159 clinical isolates. Total bacteria DNA was prepared through boiling the isolates and the DNA was then used as template for PCR am?plification. PCR, ZSCR1 and sequencing analyse of integrons were applied to all of them. Results were compared by Blast and GenBank. Results Antibiotic sensitivity results showed that in the S. flexneri strains the incidence of resistance to tet?racycline and streptomycin were 88.68%and 81.13%in the S. flexneri strains while the incidence of resistant to chloram?phenicol and trimethoprim-sulfamethoxazol were both 56.60%, and the incidence of multidrug drug resistance was 77.36%. In the sonnei strains, the incidence of resistance to ampicillin, trimethoprim-sulfamethoxazo were 97.17% and 95.28%, 83.96%and 76.42%respectively, and the incidence of multidrug resistance was 98.11%. Among all isolates, 118 were class 1 integron positive , 70 were class 2 integron positive and 89 were double positives. For those 118 isolates that are positive of class 1 integron, 23 were typical while 95 weres atypical. The gene cassettes of typical class 1 integrons contains aadA2, aa?dA1, dfrⅠ, blaoxa-10 and blaoxa-1. IntI1, aadA, blaoxa-1 and IS1 were included in the gene cassetes of the atypical class 1 integrons. Class 2 integrons positive isolates carried gene cassttes which include dfrA1, satl and aadA1. No ISCR1 was found in any isolate. Integron carriage strains were closely associated with higher rate of multiple antibiobic resistance com?pared with the organisms without integrons (90.65%,50%, P<0.05). Conlusion Class 1 and class 2 integrons were widely existence in Shigella isolates and were related to the multidrug resistance.

AIM:To investigate the ability of a metal complex ammonium tetrathiomolybdate (ATTM) to re-lease H2 S and its cytoprotective effect on an oxidative injury model .METHODS:Released H2 S was absorbed in a reaction flask from ATTM dissolved in the cell medium .Staining with dichlorodihydrofluorescein diacetate or rhodamine 123 fol-lowed by photofluorography was conducted for the observation of reactive oxygen species ( ROS) and mitochondrial mem-brane potential (ΔΨm) levels, respectively.Cell viability and release of lactate dehydrogenase (LDH) from the cells were measured with commercial kits.RESULTS:Similar to another H2S donor GYY4137, ATTM had an ability to release H2S in the cell medium in a dose-dependent manner .Treatment of human skin HaCaT cells with ATTM at concentrations of 25~400 μmol/L didn’ t significantly alter cell viability .Exposure of the cells to ultraviolet rays or a ROS donor H 2 O2 in-creased the intracellular ROS levels .Treatment with 400 μmol/L H2 O2 significantly reduced the viability of HaCaT cells (P<0.01).However, before the treatment with H2O2, pretreatment with ATTM at 100 and 200 μmol/L markedly pre-vented the H2O2-induced cell injury (P<0.01).In addition, the treatment with H2O2 triggeredΔΨm loss (P<0.01) and LDH release from the cells (P<0.01).Prior to suffering from H2O2 injury, the preconditioning with 200 μmol/L ATTM significantly improved ΔΨm levels ( P<0.05 ) and attenuated LDH release from the cells ( P<0.01 ) .CONCLUSION:ATTM is capable of releasing H 2 S and protecting human skin cells against oxidative injury .

OBJECTIVETo observe mainfestations of syndrome and biochemical indices of hypertensive model rats with excessive accumulation of phlegm-dampness syndrome (EAPDS), and to explore its possible pathological mechanism.

METHODSEAPDS rat model was prepared in 50 Wistar rats by feeding with high fat forage. Meanwhile, a normal control group consisting of 10 Wistar rats was set up by feeding with normal forage. After 25-week continuous feeding, 22 rats with body weight (BW) and blood pressure (BP) exceeding 25% those of the control group were selected as a model group. BW, BP, blood lipids, and related serological indicators were detected in all rats. Morphological changes of target organs were observed. mRNA expression levels of leptin receptor (LepR), Janus kinase2 (Jak2), signal transducer and activator of transcription 3 (Stat3), suppressor of cytokine signaling-3 (Socs3), angiotensin II receptor type 1 (AT1), angiotensin II receptor type 2 (AT2), phosphatidylinositol 3 kinase (P13K), serine threonine kinase (Akt), nuclear factor of kappa B (NF-κBp65), inhibitor of nuclear factor kappa-B kinase α (IKKα), NF-kappa-B inhibitor β (lKKβ), NF-kappa-B inhibitor α (IKBα), and AMP-activated protein kinase (AMPK) were detected by quantitative real-time PCR (qPCR). Expression levels of AT1 and LepR in aorta were detected by immunohistochemical assay and Western blot respectively.

RESULTSCompared with the control group, BW, BP, and blood lipids increased; serum levels of leptin (Lep) , Ang II, Hcy, ET-1, TNF-α, IL-6, and p2-MG increased, but NO decreased in the model group (P < 0.05, P < 0.01). Aortal endothelial injury and smooth muscle cell proliferation occurred in the model group, accompanied with heart and renal injury. Compared with the control group, mRNA expression levels of LepR, Jak2, Stat3, Socs3, AT1 , PI3K, Akt, NF-κB p65, IKKβ, IKBα, and AMPK in aorta were up-regulated significantly (P < 0.05), while the expression of IKKa decreased (P < 0.05). Immunohistochem- ical staining showed, brownish yellow deposit of AT1 and LepR was obviously increased, with more extensively positive distribution. Western blot results showed, as compared with the control group, protein expression levels of AT1 and LepR obviously increased in the model group (P < 0.05).

CONCLUSIONSModel rats exhibited typical syndromes of EAPDS. They put up weight with fat abdomen, gloomy hair, poor appetite, hypersomnia, lowered activities , reduced food intake, loose stool, dark red tongue, white tongue with white, thick, greasy fur. Lep could be taken as one of objective indicators for evaluating hypertension rat model with EAPDS.

Animals ; Aorta ; Cell Proliferation ; Disease Models, Animal ; Hypertension ; physiopathology ; I-kappa B Proteins ; Interleukin-6 ; Leptin ; blood ; NF-KappaB Inhibitor alpha ; NF-kappa B ; Phosphatidylinositol 3-Kinases ; Rats ; Rats, Wistar ; Suppressor of Cytokine Signaling Proteins ; Transcription Factor RelA ; Tumor Necrosis Factor-alpha

Background:Pulmonary arterial hypertension (PH) is a progressive disease with limited therapeutic options,ultimately leading to right heart failure and death.Recent findings indicate the role of the Warburg effect (aerobic glycolysis) in the development of PH.However,the effect of the glycolysis inhibitor 3-bromopyruvate (3-BrPA) on the pathogenesis of PH has not been well investigated.This study aimed to determine whether 3-BrPA inhibits PH and its possible mechanism.Methods:PH was induced in adult Sprague-Dawley rats by a single intraperitoneal injection of monocrotaline (MCT).3-BrPA,or phosphate-buffered saline (PBS) was administered via intraperitoneal injection every other day from the first day of MCT-injection to 4 weeks of follow-up,and indices such as right ventricular systolic pressure (RVSP),right ventricular hypertrophy index (RVHI),pulmonary arteriolar remodeling indicated by percent media thickness (％ MT),lactate levels and glucose consumption,were evaluated.Pulmonary arteriolar remodeling and right ventricular hypertrophy were observed in hematoxylin-eosin-stained lung sections.Western blotting,immunohistochemistry,and/or immunofluorescence analyses were used to measure the expression of relevant proteins.A cytochrome C release apoptosis assay and terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling staining were used to measure cell apoptosis.Results:MCT-induced PH showed a significant increase in glucose consumption (0 vs.4 weeks:0.87 ± 0.23 vs.2.94 ± 0.47,P =0.0042) and lactate production (0 vs.4 weeks:4.19 ± 0.34 vs.8.06 ± 0.67,P =0.0004).Treatment with 3-BrPA resulted in a concomitant reduction in glucose consumption (1.10 ± 0.35 vs.3.25 ± 0.47,P =0.0063),lactate production (5.09 ± 0.55 vs.8.06 ± 0.67,P =0.0065),MCT-induced increase in RVSP (39.70 ± 2.94 vs.58.85 ± 2.32,P =0.0004),pulmonary vascular remodeling (％ MT,43.45％±1.41％ vs.63.66％±1.78％,P＜0.0001),and right ventricular hypertrophy (RVHI,38.57％ ± 2.69％ vs.62.61％ ± 1.57％,P ＜ 0.0001) when compared with those of the PBS-treated group.3-BrPA,a hexokinase 2 inhibitor,exerted its beneficial effect on PH by decreasing aerobic glycolysis and was also associated with inhibiting the expression of glucose transporter protein-1,inducing apoptosis,and suppressing inflammation.Conclusions:3-BrPA might have a potential beneficial effect on the PH treatment.

Objective To investigate changes in the healthcare and support needs during the diagnostic period,and factors that affect these needs in women with suspected breast cancer.Methods This study used an investigator-developed,self-administered questionnaire to collect data from 283 women on three occasions:notification cation of need for breast biopsy,before biopsy and after diagnosis.Results The total score of need before the patients told breast biopsy was (27.68 ±0.53 ),and was higher than that after diagnosis ( 26.80 ±0.47) and the highest score was that before biopsy,which was ( 27.93 ± 0.49),and the difference among the three groups was significant ( F=6.48,P ＜ 0.01 ) ; needs score before diagnosis was ( 28.83 ± 0.31 ) and (27.06 ± 0.46) after diagnosis in participants whose education background was senior middle school or above;needs score before diagnosis was (27.04 ± 0.34) and ( 26.92 ± 0.48) after diagnosis in participants whose education background was junior high school or below; the differences was significant ( t=- 44.09,- 2.40,respectively; P ＜ 0.05 ).Conclusions Need levels of women with suspected breast cancer vary during the diagnostic period,and are highest before breast biopsy,and related to personal characteristics and cultural context.Therefore,during this period,nursing staff should provide patients and families with cuhurally sensitive,individualized,supportive care.

Objective To investigate the role and molecular mechanism of exosomal miR-224-5p in colorectal cancer (CRC).Methods The miR-224-5p expression in CRC patient tissues and cell-derived exosomes was measured by laser capture microdissection and qRT-PCR, respectively. Dual-luciferase reporter gene assay was used to determine the target gene of miR-224-5p. The protein expressions of p53 and unc-51 like kinase 2 (ULK2) in CRC cells were detected by western blot. Flow cytometry was used to detect cell cycle and apoptosis. Cell proliferation was measured by CCK8 and EdU assay.Results The miR-224-5p expression was upregulated in CRC tissues and increased progressively with the rise of CRC stage. CRC cells secreted extracellular miR-224-5p mainly in an exosome-dependent manner, and then miR-224-5p could be transferred to surrounding tumor cells to regulate cell proliferation in the form of autocrine or paracrine. Moreover, ULK2 was characterized as a direct target of miR-224-5p and was downregulated in CRC tissues. Interestingly, ULK2 inhibited CRC cell proliferation in a p53-dependent manner. Furthermore, exosome-derived miR-224-5p partially reversed the proliferation regulation of ULK2 on CRC cells.Conclusion Our findings demonstrate that exosome-transmitted miR-224-5p promotes p53-dependent cell proliferation by targeting ULK2 in CRC, which may offer promising targets for CRC prevention and therapy.