OBJECTIVE To investigate the distribution of drug resistance of Pseudomonas aeruginosa(PAE) among neonates and analyze the characteristic of the PAE infection.METHODS API system was used for the identification of 131 PAE clinical isolates and the resistance to 17 kinds antibiotics was determined by K-B method.RESULTS Most of 131 strains were isolated from sputum(42.0%) and gastric juice(32.8%).All strains were mainly isolated from neonate intensive care unit(NICU).The sensitivity to amikacin,levofloxacin,ofloxacin,ciprofloxacin,piperacillin/tazobactam,cefoperazone/sulbactam,imipenem and meropenem was respectively over 70.0%.PAE was inferior sensitivity to piperacillin,mezlocillin,cefoperazone,ceftriaxone,ceftazidime and aztreonam.CONCLUSIONS PAE is one of the most common pathogens causing nosocomial infection especially for neonates.Its susceptibility to antibiotics showed multidrug resistance.In order to reduce or prevent the occurrence of resistant isolate,we should rationally choose and use antibiotics combining with trait of neonate.

Objective To clone human vacuolar protein sorting 4A gene(hVPS4A)and to construct its eukaryotic expressive plasmid.Methods Primers were designed to amplify the full length hVPS4A by PCR using cDNA of Huh7 cell as a template,then the target DNA was inserted into the eukaryotic vector pRK5.The recombinant plasmid was confirmed by PCR,restriction enzyme digestion and DNA sequencing.Results A 1 300 bp fragment was successfully amplified by PCR from the cDNA of Huh7 cells.Af-ter recycled,purified and ligated with the vector pRK5,the recombinant plasmid was transformed into E.coli DH5α.The positive re-combinant plasmid identified by PCR was selectred and digested by EcoRⅠto get a 5 900 bp fragment;and two fragments including 4 600 bp and 1 350 bp were obtained using EcoRⅠand HindⅢ digestion;the size of these two fragments were consistent with the pRK5 target fragment and the inserted hVPS4A as expected.Moreover,DNA sequencing results confirmed that the inserted frag-ment was in accordance with the hVPS4A reference sequence.Conclusion The eukaryotic expression vector containing hVPS4A gene is constructed successfully,which provides the condition for further study on the hVPS4A biological functions.

This study examined the anti-hepatitis B virus (HBV) effect of wild-type (WT) vacuolar protein sorting 4B (VPS4B) and its dominant negative (DN) mutant VPS4B-K180Q in vivo in order to further explore the relationship between HBV and the host cellular factor VPS4. VPS4B gene was amplified from Huh7 cells by RT-PCR and cloned into the eukaryotic expression vector pXF3H. Then, the VPS4B plasmid and the VPS4B-K180Q mutation plasmid were constructed by using the overlap extension PCR site-directed mutagenesis technique. VPS4B and HBV vectors were co-delivered into mice by the hydrodynamic tail-vein injection to establish HBV vector-based models. Quantities of HBsAg and HBeAg in the mouse sera were determined by ElectroChemiLuminescence (ECL). HBV DNA in sera was measured by real-time quantitative PCR. Southern blot analysis was used to assay the intracellular HBV nuclear capsid-related DNA, real-time quantitative PCR to detect the HBV-related mRNA and immunohistochemical staining to observe the HBcAg expression in the mouse liver tissues. Our results showed that VPS4B and its mutant VPS4B-K180Q could decrease the levels of serum HBsAg, HBeAg and HBV-DNA. In addition, the HBV DNA replication and the mRNA level of HBV in the liver tissues of treated mice could be suppressed by VPS4B and VPS4B-K180Q. It was also found that VPS4B and VPS4B-K180Q had an ability to inhibit core antigen expression in the infected mouse liver. Furthermore, the anti-HBV effect of mutant VPS4B-K180Q was more potent than that of wild-type VPS4B. Taken together, it was concluded that VPS4B and its DN mutant VPS4B-K180Q have anti-HBV effect in vivo, which helps develop molecular therapeutic strategies for HBV infection.

Objective:To detect the inhibitory effects of CAL-101, a selective inhibitor of phosphoinostitide-3'-kinase delta (PI3Kδ), on Burkitt's lymphoma cell line Raji and diffused large B-cell lymphoma cell line SUDHL-10 and elucidate its relative mechanism. Methods:Raji and SUDHL-10 cells were treated with various concentrations of CAL-101. Methyl thiazolyl tetrazolium (MTT) assay was performed to determine the inhibitory effect of CAL-101 on lymphoma cells, and cell apoptosis was measured by Annexin V/PI and DAPI staining. Migration assays were performed with transwell to detect the migration of lymphoma cells derived from the stromal cell line HK. Western blot was used to detect the phosphorylation status of the ERK pathway. MTT and CalcuSyn software analyses were preformed to detect whether or not combining CAL-101 with bortezomib induces synergistic cytoxicity. Results:CAL-101 at con-centrations of 5, 10, 15, and 20μmol/L inhibited cell proliferation in a dose-dependent manner. The proliferation rates of the Raji cells treated with 5, 10, 15, and 20μmol/L for 48 h were 29.17%± 1.23%, 38.15%± 1.51%, 46.46%± 1.78%, and 55.8%± 2.01%, respec-tively, which were significantly higher (P<0.05) than that of the control group (1.15% ± 0.02%). Similar results were found in the SUDHL-10 cells after treatment with CAL-101 (P<0.05). CAL-101 also exerted an apoptotic effect on the lymphoma cells. The apop-totic rates of the Raji cells treated with CAL-101 for 21 h were 22.69%± 3.83%and 49.96%± 7.36%, respectively, which were signifi-cantly higher (P<0.05) than that of the control group (5.23%± 2.04%). Similar results were found in the SUDHL-10 cells (P<0.05). Treatment with 5 and 10 μmol/L CAL-101 dose-dependently inhibited the migration activity of lymphoma cells to stromal cells (P<0.05). Western blot analysis showed that the expression level of ERK phosphorylation protein was significantly downregulated in the cells treated with CAL-101. A synergistic effect between CAL-101 and bortezomib was verified. That is, these two drugs can signifi-cantly inhibit the proliferation of lymphoma cells with CI values less than 1. Conclusion:The PI3Kδ-specific inhibitor CAL-101 sup-pressed the proliferation of Raji and SUDHL-10 cells, induced apoptosis, and inhibited stromal cell-derived migration. This inhibitory effect may be induced by blocking the ERK pathway. Overall, our study indicated that CAL-101 is a novel and potential agent in the therapeutic strategy against aggressive B-cell lymphoma.

Objective To compare the intubating conditions between dexmedetomidine and remifentanil when combined with sevoflurane-nitrous oxide (N2O) for anesthesia induction in the pediatric patients.Methods A total of 122 pediatric patients,aged 4-10 yr,of American Society of Anesthesiologists physical status Ⅰ,undergoing elective plastic surgery,were randomly divided into dexmedetomidine group (group D,n =61) and remifentanil group (group R,n=61).Eight percent sevoflurane and 60％ N2O were inhaled for induction of anesthesia,and the fresh gas flow was set at 6 L/min.After disappearance of eyelash reflex,dexmedetomidine 1 μg/kg and remifentanil 1 μg/kg were intravenously injected over 50-60 s in D and R groups,respectively,and 1 min later tracheal intubation was performed.The intubating conditions were graded,and the satisfactory intubating conditions and successful intubation were recorded.The development of adverse cardiovascular reactions and complications such as hyoxemia and laryngospasm before and after intubation and postoperative pharyngodynia was recorded.Results Compared with group D,no significant change was found in the success rate of intubation,rate of satisfactory intubation,intubating condition grade or incidence of postoperative pharyngodynia (P＞ 0.05),and the incidence of hypertension and sinus tachycardia after intubation was significantly increased in group R (P＜0.05).No pediatric patients developed hyoxemia,laryngospasn or sinus tachycardia in two groups.Conclusion When 8％ sevoflurane and 60％ N2O are inhaled for anesthesia induction,combing with dexmedetomidine 1 μg/kg produces better clinical efficacy than combing with remifentanil 1 μg/kg in improving the intubating conditions for pediatric patients.

This study examined the anti-hepatitis B virus (HBV) effect of wild-type (WT) vacuolar protein sorting 4B (VPS4B) and its dominant negative (DN) mutant VPS4B-K180Q in vivo in order to further explore the relationship between HBV and the host cellular factor VPS4. VPS4B gene was amplified from Huh7 cells by RT-PCR and cloned into the eukaryotic expression vector pXF3H. Then, the VPS4B plasmid and the VPS4B-K180Q mutation plasmid were constructed by using the overlap extension PCR site-directed mutagenesis technique. VPS4B and HBV vectors were co-delivered into mice by the hydrodynamic tail-vein injection to establish HBV vector-based models. Quantities of HBsAg and HBeAg in the mouse sera were determined by ElectroChemiLuminescence (ECL). HBV DNA in sera was measured by real-time quantitative PCR. Southern blot analysis was used to assay the intracellular HBV nuclear capsid-related DNA, real-time quantitative PCR to detect the HBV-related mRNA and immunohistochemical staining to observe the HBcAg expression in the mouse liver tissues. Our results showed that VPS4B and its mutant VPS4B-K180Q could decrease the levels of serum HBsAg, HBeAg and HBV-DNA. In addition, the HBV DNA replication and the mRNA level of HBV in the liver tissues of treated mice could be suppressed by VPS4B and VPS4B-K180Q. It was also found that VPS4B and VPS4B-K180Q had an ability to inhibit core antigen expression in the infected mouse liver. Furthermore, the anti-HBV effect of mutant VPS4B-K180Q was more potent than that of wild-type VPS4B. Taken together, it was concluded that VPS4B and its DN mutant VPS4B-K180Q have anti-HBV effect in vivo, which helps develop molecular therapeutic strategies for HBV infection.
ATPases Associated with Diverse Cellular Activities ; Adenosine Triphosphatases ; physiology ; Animals ; Endosomal Sorting Complexes Required for Transport ; physiology ; Female ; Genes, Dominant ; genetics ; Hepatitis B ; metabolism ; virology ; Hepatitis B virus ; physiology ; Liver ; virology ; Mice ; Mice, Inbred BALB C ; Mutation ; genetics ; Virus Inactivation

Objective To evaluate and compare the efficacy and safety of sedative and analgesic anesthesia in surgical patients with supine and prone position .Methods Sixty female patients, American Society of Anesthesiologists physical status Ⅰ or Ⅱ, aged from 18 -53 years, scheduled for elective plastic operations under sedative and analgesic anesthesia combined with local anesthesia were divided into two groups according to their surgical positions: supine group ( n=30 ) and prone group ( n=30 ) .All patients received Ⅳ dexmedetomidine ( DEX) 1 μg/kg over 15 min followed by 0.4 -0.7 μg/kg/h infusion. Both groups were administered Ⅳ midazolam 0.04 mg/kg and a continuous infusion of remifentanil of 0.1 μg/kg/min at the beginning of anesthesia .Heart rate ( HR) , mean arterial pressure (MAP), pulse oximetry (SpO2), respiratory rate (RR), bispectral index (BIS) and Ramsay sedation scores ( RSS) were recorded at the following time points: before anesthesia ( T0 ) , 5 min after induction with midazolam ( T1 ) , 10 min after induction of midazolam ( T2 ) , immediately after induction with DEX( T3 ) , the beginning of local anesthesia ( T4 ) , the beginning of surgery ( T5 ) , 30 min after anesthesia induction ( T6 ) , 60 min after anesthesia induction ( T7 ) , immediately after turning off DEX infusion (T8), the end of surgery (T9).Incidences of respiratory depression, incidences of apnea, oxygen supplementation by facial mask and jaw-thrust, frequencies of body movements and additional rescue medication were also recorded .After surgery , recall of events during surgery , the visual analogue scales (VAS) for pain in PACU, the satisfaction levels of patients and surgeons were also assessed .Results No significant differences were found in MAP , SpO2 , RR, BIS, RSS scores at any time point between two groups (all P >0.05).There were no significant differences in incidences of respiratory depression , frequencies of body movements and additional rescue medication during surgery between groups ( all P>0. 05).Neither were recall of events during surgery , the visual analogue scales (VAS) for pain and the satisfaction levels of patients and surgeons after surgery (all P>0.05).The HR at time points of T0, T1, T2 in prone group were significantly higher than those in supine group (all P<0.05).Compared with the supine group , the incidences of apnea , oxygen supplementation by facial mask and jaw-thrust in prone group were significantly lower .Conclusions Sedative and analgesic anesthesia is effective and safe for patients with prone surgical position and has a lower incidence of upper airway obstruction during surgery than patients in supine surgical position .

Objective To evaluate and compare the efficacy and safety of sedative and analgesic anesthesia in surgical patients with supine and prone position .Methods Sixty female patients, American Society of Anesthesiologists physical status Ⅰ or Ⅱ, aged from 18 -53 years, scheduled for elective plastic operations under sedative and analgesic anesthesia combined with local anesthesia were divided into two groups according to their surgical positions: supine group ( n=30 ) and prone group ( n=30 ) .All patients received Ⅳ dexmedetomidine ( DEX) 1 μg/kg over 15 min followed by 0.4 -0.7 μg/kg/h infusion. Both groups were administered Ⅳ midazolam 0.04 mg/kg and a continuous infusion of remifentanil of 0.1 μg/kg/min at the beginning of anesthesia .Heart rate ( HR) , mean arterial pressure (MAP), pulse oximetry (SpO2), respiratory rate (RR), bispectral index (BIS) and Ramsay sedation scores ( RSS) were recorded at the following time points: before anesthesia ( T0 ) , 5 min after induction with midazolam ( T1 ) , 10 min after induction of midazolam ( T2 ) , immediately after induction with DEX( T3 ) , the beginning of local anesthesia ( T4 ) , the beginning of surgery ( T5 ) , 30 min after anesthesia induction ( T6 ) , 60 min after anesthesia induction ( T7 ) , immediately after turning off DEX infusion (T8), the end of surgery (T9).Incidences of respiratory depression, incidences of apnea, oxygen supplementation by facial mask and jaw-thrust, frequencies of body movements and additional rescue medication were also recorded .After surgery , recall of events during surgery , the visual analogue scales (VAS) for pain in PACU, the satisfaction levels of patients and surgeons were also assessed .Results No significant differences were found in MAP , SpO2 , RR, BIS, RSS scores at any time point between two groups (all P >0.05).There were no significant differences in incidences of respiratory depression , frequencies of body movements and additional rescue medication during surgery between groups ( all P>0. 05).Neither were recall of events during surgery , the visual analogue scales (VAS) for pain and the satisfaction levels of patients and surgeons after surgery (all P>0.05).The HR at time points of T0, T1, T2 in prone group were significantly higher than those in supine group (all P<0.05).Compared with the supine group , the incidences of apnea , oxygen supplementation by facial mask and jaw-thrust in prone group were significantly lower .Conclusions Sedative and analgesic anesthesia is effective and safe for patients with prone surgical position and has a lower incidence of upper airway obstruction during surgery than patients in supine surgical position .

OBJECTIVETo investigate the proliferation inhibitory role and mechanism of PI3Kδ inhibitor CAL-101 on multiple myeloma (MM) cells, and to provide new therapeutic options for MM treatment.

METHODSMM cell lines U266 and RPMI8226 cells were treated with various concentrations of CAL-101. MTT assay and CalcuSyn software were performed to determine the inhibitory effect of CAL-101 and the synergistic effect with PCI- 32765, SAHA (suberoylanilide hydroxamic acid), BTZ (Bortezomib) on MM cells. The protein expression level of p-AKT, p-ERK, AKT, ERK and PI3Kδ processed by CAL-101 were analyzed by Western blot.

RESULTSCAL-101 at concentration of 15, 20, 25, 30 and 40 μmol/L could induce significant dose-dependent proliferation inhibition on U266 cells after treatment for 48 hours. The cell proliferation inhibition rates were (33.54 ± 1.23)%, (41.72 ± 1.78)%, (53.67 ± 2.01)%, (68.97 ± 2.11)% and (79.25 ± 1.92)%, respectively. Similar results were found in RPMI8226 cell line. Western blots showed high expression level of p-AKT, p-ERK, AKT, ERK and PI3Kδ in cell lines and MM primary cells. p-AKT and p-ERK protein expression levels were down-regulated significantly by CAL-101 treatment. Synergistic effect has been verified between CAL-101 and PCI-32765, SAHA and Bortezomib in U266 cell line, and PCI-32765, Bortezomib in RPMI8226 cell line with CI values less than 1.

CONCLUSIONCAL-101 could inhibit proliferation of MM cell lines. High levels of p-AKT, p-ERK, AKT, ERK and PI3Kδ protein expression were observed in both cell lines and primary cells. Down-regulation of p-AKT and p-ERK probably related with the mechanism of CAL-101 in MM cell proliferation inhibition. CAL-101 has significant synergistic effect with PCI-32765, SAHA and BTZ.

Boronic Acids ; Bortezomib ; Cell Line, Tumor ; Cell Proliferation ; Down-Regulation ; Humans ; Multiple Myeloma ; pathology ; Phosphatidylinositol 3-Kinases ; antagonists & inhibitors ; Protein Kinase Inhibitors ; pharmacology ; Purines ; pharmacology ; Pyrazines ; Pyrazoles ; Pyrimidines ; Quinazolinones ; pharmacology