Objective : To investigate the effect of a laser-etched pure titanium surface on proliferation of the human osteosarcoma cell line MG63 and to provide a basis for study of implant surface modification. Methods: The pure titanium plate was cut into titanium pieces by a numerical control machine tool and divided into smooth surface and laser etching groups. The titanium surface of the laser etching group was etched with an Nd:YAG continuous wave laser using predetermined parameters, and the surfaces were observed by scanning electron microscopy (SEM). The surface micromorphology of each titanium sheet was evaluated. The relative element content of the titanium surface was measured by energy dispersive X-ray spectroscopy (EDS). The Ra value of each surface was determined using the Veeco roughness tester. MG63 cells were inoculated on 2 sets of titanium tablets. At 1, 3, and 6 h postinoculation, cell adhesion to the two groups of titanium sheets was observed under the microscope. At 24 h after inoculation, cellular F-actin was directly stained using immunofluorescence, and the morphology of the cytoskeleton was observed by laser confocal microscopy. Cell proliferation was examined at 1, 3, and 5 d using a MTS kit, and the data were analyzed with SAS 9.4. Results : The surface of the smooth surface group was smooth and flat, the element composition was pure titanium, and the roughness Ra was 179.23 nm. The surface of the laser-etched group formed a regular and uniform pore structure. The composition was mainly Ti, O, C, etc, and the surface roughness Ra was 14.11 μm. A large number of cells were uniformly distributed on the two titanium sheets in the observations at 1, 3, and 6 h. At 24 h postinoculation, MG63 cells were completely stretched on the two sets of titanium sheets and had extended a large number of pseudopods and microfilaments to cross-link with peripheral cells; moreover, the cell division phase was observed. The cell proliferation of the two groups at 1, 3, and 5 d showed a significant increase with time, indicating that no cytotoxicity occurred on the surfaces of the two groups. However, the cell proliferation in the laser-etched group was superior to that in the mechanical smooth surface group. Conclusion The surface morphology of titanium can be controlled by laser etching, which is conductive to increase the microstructure of implants without cytotoxicity and promoting osteoblast proliferation in the early stage.

Objective To observe the influence of IL-12,IL-15 on CIK cell in the normal culture;to observe the anti-tumor effect in the circumstance of different combination of cytokines,and to provide a new insight for preparing high effective and qualified CIK cell in vitro.Methods The optimal concentrations of IL-2,IL-12 and IL-15 were determined,respectively.After the peripheral blood from healthy blood donors was collected,monocytes were selected and co-cultured with different cytokines into different groups,as group A(IL-2 normal culture group),group B(IL-2 and IL-12 group),group C(IL-2 and IL-15 group),group D(IL-2,IL-12 and IL-15 group),and group E(cytokine control group).The monocytes in different groups were calculated by globulimeter,the activity of cells was detected by Trypan blue staining,positive ratio of CD3,CD8,CD56 on the celluar membrane was detected by flow cytometry,and the anti-tumor effect of CIK to SMMC-7721 was detected by MTSmethod,inthedayof0,5,10,15,20 after the culture.Results Statistical analysis indicated that,the proliferation multiplication of CIK cells was significantly higher in group B,group C and group D after 10,15 and 20 days of culture than those in group A(P＜0.05);and group D had higher proliferation multi-plication than that of group C(P＜0.05).The percentage of CD3 + CD8 +,CD3 + CD56+ in CIK cell membrane in group B,C,D was significantly higher than that in group A after 15 and 20 days of culture (P＜0.05).The percentage of CD3+ CD8+,CD3+ CD56+ in CIK cell membrane in group D was significantly higher than that in group B after 15 and 20 days of culture (P＜0.05).The killing rate of CIK cells for liver cancer in each group at 10,15,20 days of culture was significantly higher than that of group A when the target target ratio was 5 ∶ 1 (P＜0.05).The killing rate of CIK cells for liver cancer in group D,C at 10,15,20 days of culture was significantly higher than that of group B(P＜0.05).Conclusion IL-12and IL-15 could improve the proliferation of CIK cells,and IL-15 also has the effect of enhancing CIK cells the tumor-killing to SMMC-7721 activity.

OBJECTIVETo explore the optimal time of acupuncture intervention in the assisted reproduction.

METHODSOne hundred and twenty female mice and 60 male mice were collected. 20 female mice were selected in the natural period group and the rest 100 female mice were prepared as the model of controlled ovarian hyperstimulation (COH). The model mice were randomized into a COH group, a down-regulation group, a gonadotropins (Gn) start group, an injection of human chorionic gonadotropin (HCG) group and an embryo culture group, 20 mice in each one. The donor mice and receptor mice were subdivided in each group, 10 mice in each subgroup. One week before the experiment, vas deferens ligature was done in 30 male mice and the other 30 male mice did not receive ligature. In the down-regulation group, the Gn start group, the HCG injection group and the embryo culture group, electroacupuncture (EA) was applied to "Guanyuan" (CV 4), "Zhongji" (CV 3) and "Sanyinjiao" (SP 6) at the time points accordingly. EA stimulation was in the condition of continuous wave, 2 Hz and 1 mA. No inter-vention was given in the natural period group and the COH group. On the day of HCG injection, the donor mice and the non-ligatured male mice were put in the same cage of each group. The fertilized ovum was collected with the date of fertilization marked and was fostered in the incubator. At the ratio of 1:1, the receptor mice and ligatured mice were put in the same cage in each group. The vaginal plug was examined in the next morning. The pseudopregnancy was marked with the date of plug observed. In the 68th hour of embryo culture, the embryo of the donor was shifted to the receptor on the same day when the plug was observed. The clinical pregnancy rate and embryo imbed site number were observed. RT-PCR assay was adopted to determine the expression of insulin-like growth factor-1 (IGF-1) mRNA in endometrium.

RESULTSIn the COH group, the pregnancy rate, average imbed site number and endometrial IGF-1 mRNA expression were all significantly lower than those in the natural period group (all<0.01). After EA treatment, in the Gn group, the HCG injection group and the embryo culture group, the pregnancy rates were higher significantly than those in the COH group (<0.05,<0.01). In the HCG injection group, the average imbed site number and IGF-1 mRNA expression were increased apparently as compared with those in the COH group (both<0.01), better than those in the Gn group and the embryo culture group (all<0.01).

CONCLUSIONSIn the treatment with acupuncture combined with IVF-ET for infertility, the intervention of acupuncture on the day of HCG injection is the optimal time point. It increases the secretion of endometrial IGF-1 so as to improve the clinical pregnancy rate, the mean imbed site number and the embryo implantation.

Objective Take frizzled class receptor 4(FZD4) as an example for exploring whether the transmembrane receptor protein is a suitable marker for wound age estimation. Methods A total of 78 Sprague-Dawley rats were divided randomly into the control group and the contusion groups of 4h, 8h, 12h, 16h, 20h, 24h, 28h, 32h, 36h, 40h, 44h, and 48h after injury (n=6). Using a drop-ball technique, a contusion was produced in the right lamb of rats. The samples were dissected from injury zones. Then the expression of FZD4 was detected using immunofluorescence, real-time PCR and western blot, respectively. Results FZD4 was located on the membrane of skeletal muscle cells. Compared to the control group, FZD4 mRNA showed a signiifcant increase (over 2-fold) in 8h, 12h, 36h, and 40h after injury by Real-time PCR (P<0.05), and FZD4 protein showed a statistical up-regulation (less than 2-fold) in 8h, 36h, 40h, 44h, and 48h post contusion by western blotting (P<0.05). Conclusion The expression of FZD4 mRNA and protein are both time-dependent during contused skeletal muscle repair. To some degree, FZD4 mRNA was more suitable than corresponding protein for determining wound age.

Objective T o investigate the relationship betw een the expression of secreted frizzled-related protein 5 (SFR P5) m RNA and the tim e interval after skeletal m uscle injury in rats by real-tim e PC R . Methods A total of ninety SD rats w ere random ly divided into the contusion groups at different tim es including 4 h, 8 h, 12 h, 16 h, 20 h, 24 h, 28 h, 32 h, 36 h, 40 h, 44 h, 48 h after contusion, incision groups at different tim es including 4h and 8h after incision and the control group. T he sam ples w ere taken from the contused zone at different tim e points. T he total RNA w as isolated from the sam ples and re-versely transcribed to analyze the expression levels of SFRP5 m RNA . Results C om pared to the control group, the expression of SFRP5 m RNA in contusion groups w ere dow n-regulated w ithin 48 h after con-tusion and reached the low est level at 20 h, and the expression of SFRP5 m RNA gradually increased from 20 h to 48 h after contusion. T he expression of SFRP5 m RNA in the incised groups w ere signifi-cantly low er than that of the contusion groups at 4 h after injury. A t the tim e of 8 h, the expression levels betw een the contusion and incision groups show ed no statistically significant difference. Conclusion It is suggested that SFRP5 m RNA analysis m ay show regular expression and can be a m arker for estim ation of skeletal m uscle injury age.

Objective To determine the in vitro antibacterial activity of levornidazole against 375 anaerobic isolates.Methods Agar dilution method was used to determine the minimum inhibitory concentrations (MICs)of levornidazole,3 comparators (metronidazole,ornidazole and dextrornidazole)against 375 anaerobic isolates.Results For anaerobic gram-negative and gram-positive bacilli,and anaerobic gram-positive cocci,levornidazole displayed activity similar to or slightly higher than that of met-ronidazole,ornidazole and dextrornidazole.Levornidazole showed good activity against B.fragilis,Bacteroides thetaiotaomi-cron ,Clostridium difficile ,Clostridium perfringens ,and Peptostreptococcus magnus .The MIC90 value of levornidazole a-gainst the above-mentioned anaerobes was 0.5,1,0.25,2 and 1 mg/L,respectively.However,levornidazole and the compa-rators had poor antibacterial activity against Veillonella spp.among anaerobic gram-negative cocci.Conclusions The in vitro anti-anaerobic activity of levornidazole is similar to or slightly higher than that of metronidazole, ornidazole and dex-trornidazole.Levornidazole has good activity against both gram-negative and gram-positive anaerobic bacilli,and gram-positive anaerobic cocci,suggesting its promising clinical use.

ObjectiveTo investigate the clinical pharmacokinetics (PK) and pharmacodynamics (PD) of gemifloxacin tablet in healthy Chinese volunteers and to provide evidences for optimal clinical dosing.MethodsTwenty volunteers were enrolled in the randomized (1∶1) double-blind study,and divided into administration group and control group.Each group received multiple oral doses of 320 mg of gemifloxacin tablet or placebo.The plasma and urine samples for gemifloxacin were analyzed by igh-performance liquid chromatogram(HPLC)-fluorometricmethod. Theminimuminhibition concentrations (MIC)of gemifloxacin against190clinical isolateswere determinedby broth microdilution method.The fAUC0～24 h/MIC and fCmax/MIC,with target value of 25 and 5,were used as the indices to evaluate PK and PD characteristics of gemifloxacin. The cumulative fraction of response (CFR) of gemifloxacin against each bacterium and the probability of target attainment (PTA) under various MIC level were evaluated using Monte Carlo simulation following multiple administration at steady state.ResultsThe Cmax of gemifloxacin after once-daily oral doses for 7 days were (1.55 ±0.32) μg/mL and (1.57±0.31) μg/mL for the first and last dose,while the AUC0～24 h were (7.91±1.52) and (8.91±1.15) h · μg · mL-1,respectively.The accumulation factor was 1.13±0.05.The time-profile of gemifloxacin could be described using two-compartment model and the half-life of distribution and elimination phase were (0.64 ± 0.17) and (7.10 ± 2.10) h,respectively. The cumulative urinary excretion rates within 24 h of gemifloxacin were 34.83 ％ and 38.95 ％ for the first and the last dose,respectively.PD study showed that the MIC90 of gemifloxacin were 0.25 mg/L and 0.125 mg/L against Streptococcus pneumoniae and Moraxelle catarrhalis,respectively,while the MIC90 was 2 mg/L against Hemophilus influenza. However,most of Klebsiella pneumoniae and methicillin-resistant Staphylococcus aureus (MRSA) were resistant to gemifloxacin ( MIC90 ＞ 32mg/L).The PTA values of fAUC0～24 h/MIC and fCmax/MIC of gemifloxacin 320 mg daily for 7 days were close to 100％ when MIC was ≤0.06 mg/L.ConclusionsGemifloxacin is rapidly absorbed after oral administration of single doses in healthy Chinese volunteers,and the plasma concentration could reach steady state at the third day,while a minimal accumulation is shown after consecutive 7 days dosing.The PK/PD analysis suggests that the favorable clinical and bacteriological efficacy could be obtained when using thisregimen in treatment of sensitive patients with community-acquired pneumonia and acute exacerbation of chronic obstructive pulmonary disease.

Objective To characterize clinical feature, frequency of isolation and antimicrobial susceptibility of pathogens isolated from patients with nosocomial bloodstream infections in Huashan Hospital, Fudan University from 1995 to 2004. Methods The clinical data of all patients who were diagnosed with nosocomial bloodstream infections based on national diagnostic criteria of nosocomial bloodstream infections were retrospectively analyzed. The pathogens were routinely isolated and identified. Susceptibilities against antimicrobial agents were determined by Kirby-Bauer methods and analyzed by WHONET 5.0 software. Results During the 10-year study period, a total of 395 patients were diagnosed with nosocomid bloodstream infection with 435 strains isolated from blood specimen.Gram positive bacteria, Gram negative bacilli and fungi, accounted for 47.4%, 45.1 % and 7.6%,respectively. Coagulase-negative Staphylococci (21.4%), S. aures (17.9%), E.coli (13.6%), K. pneumoniae (10.8%), Candidaspp (7.4%), Enterococci (6.0%), Pseudomonasspp (6.0%) and Acinetobacter spp (3.7%) were frequently identified isolates. S. aures and coagulase-negative Staphylococci resistant to methicillin were 62.8% and 87.1%, respectively. The susceptibilities of cefotaxime and ceftazidime against E. coli and K. preumonine were 46%-78% and 27.7%-40.4%, respectively. Conclusions The Gram positive cocci are slightly more prevalent than Gram negative bacilli in nosocomial bloodstream infections and resistance to the first line antibiotics is common among all pathogens isolated. Candida spp is the fifth leading cause of nosocomial bloodstream infections.

OBJECTIVE To characterize clinical feature,frequency of isolation and antimicrobial susceptibility of bacterial pathogens isolated from patients with healthcare-associated bacterial meningitis(HABM).METHODS We review the charts of all patients in whom the diagnosis was based on(national) diagnostic criteria of healthcare-(associated) infections at Huashan Hospital from 1995 through 2004.The pathogens were routinely isolated,(identified) and susceptibilities against antimicrobial agents were determined by Kirby-Bauer methods.RESULTS During the 10-year study period,109 patients were treated for 120 episodes of HABM.Most of patients had a(history) of recent and remote neurosurgery.Fever was present in all patients,while nuchal rigidity and decrease consciousness were present in less than half of all patients.CSF opening(pressure,) white blood cell count and(protein) were elevated with predominance of neutrophils.A total of 120 strains were isolated from CSF specimen,Gram-positive bacteria and Gram-negative bacilli were accounted for 35.8% and(64.2%) of all isolates,respectively.Acinetobacter spp(24.2%),coagulase-negative staphylococci(22.5%),Klebsiella pneumoniae(12.5%),Pseudomonas aeruginosa(10%),Enterobacter cloacae(8.3%) and Staphylococcus aureas(7.5%) were the 6 most frequent isolates and resistance to the first line antibiotics was common among all pathogens isolated.(CONCLUSIONS) The most common risk factor for HABM is neurosurgery.Gram-negative bacilli and staphylococci are important causes of HABM,resistance to the first line antibiotics is common among all pathogens isolated.

20 antimicrobial agents were determined by Kirby-Bauer methods by the participating institutions.RESULTS A total of 428 strains were isolated,Gram-positive cocci and Gram-negative bacilli accounted for 49.8% and 50.2%,respectively.The Gram-positive bacteria increased from 44.6% to 51.2% from 1995-1996 through 2003-2004 and Gram negative bacteria decreased from 55.4% to 48.2% in the meantime.The most frequent Gram-positive isolates were coagulase negative staphylococci,Staphylococcus aureus,Enterococcus spp and Streptococcus pneumoniae;the most frequent Gram-negative isolates were Acinetobacter spp,Klebsiella spp,Escherichia coli,Pseudomonas aeruginosa and Enterobacter spp;Neisseria meningitidis,Haemophilus influenzae,Flavobacterium spp and Citrobacter spp were relative less common.No strains resistant to vancomycin were found in staphylococci and enterococci.Gram-negative bacilli were highly susceptible to carbapenem.CONCLUSIONS Gram-positive cocci play an increasing role in central nervous sysytem infections,especilly coagulase negative staphylococci,and Gram-positive cocci have been increasing from 1995-1996 through 2003-2004,resistance to the first line antibiotics is common among all cerebrospinal fluids isolates.