Objective To construct the eukaryotic plasmid expression vector mediated short hairpin RNA(shRNA) interference targeting TLR4 gene,and transfect it into pancreatic adenocarcinoma cell line PANC1,then screen stably transfected clonal cell line.Methods Three shRNA interference expression plasmid vectors targeting the TLR4 gene were constructed,named TLR4-1,TLR4-2,TLR4-3.The shRNA plasmid with highest inhibitory efficiency was selected and transfected into PANC1 cells with liposome.The silencing efficiency and transfection efficiency of TLR4-shRNA was assayed with real-time quantitative PCR and flow cytometry analysis.Monoclonal cell with stable transfection of TLR4-shRNA were selected by geneticin 418 (C418) and limiting dilution analysis.Results Transient transfection efficiency of PANC1 was (46.72 ±5.06) ％.TLR4 mRNA expressions were 0.025 ± 0.004,0.027 ± 0.003,0.019 ± 0.006in cells transfected with TLR4-1,TLR4-2,TLR4-3,respectively,which were significantly lower than that in untransfected group (0.061 ±0.018) and negative control group (0.057 ±0.015,P ＜0.05).The transfection efficiency of TLR4-3 vector in stably transfected clones [(82.79 ±8.16)％] was significantly higher than that of transient transfection (P =0.001 ).The expression of TLR4 mRNA was decreased to 0.010 ± 0.002,which was significantly lower than that of transient transfection ( P =0.001 ).The expression of TLR4 protein was (0.54±0.32) ％,which was significantly lower than that of untransfected cells [( 87.42 ± 5.00 ) ％] and that of negative control [(82.9±5.00)％,P =0.000].Conclusions Stable transfection PANC1 cell lines with TLR4 gene silencing are successfully identified.

BACKGROUND: Presently, it is a certain preventive effect of local emplacement of various isolated materials during operation and postoperatively drug-controlled inflammatory reaction on adhesion between epidural and nerve root after laminectomy.OBJECTIVE: To study the preventive effect of chitosan and titanium net on epidural cicatricle adhesion after laminectomy.DESIGN, TIME AND SETTING: Randomized controlled animal study, which was performed in Guiyang Medical College from June to December 2005.MATERIALS: Eighty adult rabbits, weighing (2.0±0.2) kg, were used to establish vertebral plate resection models. Chitosan was provided by Shanghai Qisheng Biological Agent Industry Company, and titanium net by Guizhou Kelun Pharmacology Company Limited.METHODS: Eighty adult rabbits were randomly divided into four groups, including control group (0.5 mL saline), titanium net group (titanium net in the size of 1.0 cm × 0.8 cm was covered on defect region of vertebral plate and fixed on processus spinosus of upper and lower cone, bilateral muscles and soft tissues), chitosan group (2 mL chitosan), and chitosan + titanium net group (combination of chitosan and titanium net). Six rabbits were sacrificed in the 2nd and 4th weeks postoperatively, and eight rabbits were chosen as the samples in the 8th week.MAIN OUTCOME MEASURES: Morphological changes of cicatricle.RESULTS: Cicatricle was clearer in the control group. Titanium net could relieve peripheral cicatricle of dura mater and effectively isolate peripheral cicatricial tissue and spinal dura meter. Chitosan could remarkably prevent from cicatricle adhesion and inhibit inflammatory reaction. The combination of chitosan and titanium net could effectively inhibit proliferation of cicatricle surrounding dura meter.CONCLUSION: Association of chitosan and titanium net can effectively inhibit cicatricle adhesion surrounding dura meter after laminectomy.

ObjectiveTo evaluate and compare long-term clinical outcomes of arthroscopic anterior cruciate ligament (ACL) reconstruction with remnant preservation versus remnant resection.MethodsFrom October 1999 to May 2005,standard ACL reconstruction with autogenous quadrupled hamstring tendons under arthroscopy using remnant resection with transtibial techniques was conducted in 87 patients.From June 2005 to May 2010,ACL reconstruction with autogenous quadrupled hamstring tendons using outside-in bone tunnel establishment and remnant preservation was performed in 221 patients.Only the patients who underwent simple ACL reconstruction together with more than three years follow-up records were enrolled in the study,including 66 patients in remnant preservation group and 39in remnant resection group.ResultsPatients of both groups were all followed up for 36-60 months,which showed insignificant difference between groups ( P ＞ 0.05).Compared with the preoperative data,the latest follow up data demonstrated satisfactory outcomes in both groups.The two groups showed no significant differences regarding the preoperative data including age and gender distribution,average time from injury to surgery,joint instability extent and functional score of the affected knee ( P ＞ 0.05 ).There was no statistical difference in regards of the thigh muscle atrophy and Lachman test between groups ( P ＞0.05).However,the remnant preservation group was superior to the remnant resection group in aspects of the range of motion of the affected knee,Lysholm score,stability in anterior drawer test,stability in pivot shift test and international knee documentation committee (IKDC) grade ( all P ＜ 0.05 ). ConclusionCompared with standard ACL reconstruction using remnant resection with transtibial techniques,the ACL reconstruction using outside-in bone tunnel establishment and remnant preservation has better longterm clinical outcomes including stability and function of the affected knee.

The genetically modified mice , as a helpful model , have been widely used in life scientific research . However, several new issues appeared subsequently with the wide application of the genetically modified mice .Here, we mainly discussed and analyzed the problems in the management and usage of genetically modified mice , which underlies the foundation for establishing management practice of the genetically modified mice .

Objective To evaluate the clinical application of multi-slice spiral CT(MSCT)for foreign body of trachea or bronchus in children.Methods Twenty cases suspected foreign body were examined with MSCT.The post-processing reconstruction included multiple planar reconstruction(MPR),CT virtual bronchoscopy(CTVB),shaded surface display(SSD),minimum intensity projection(MinIP),and volume rendering(VR).Results The foreign bodies located at trachea in 1 case and at bronchia in 19 cases.The foreign bodies included 18 fragments of peanut and melon seeds,one shrimp and one plastic-tube.All cases were confirmed by fiberoptic bronchoscopy or course of diseases.The position and form of foreign bodies,as well as the relationship between foreign bodies and bronchial muous membrane could be showed by MPR,CTVB,SSD,MinIP and VR.Conclusion MSCT scanning is a significant noninvasive and sensitive diagnostic technique in detecting the foreign body in trachea or bronchus.

Objective To investigate the inhibitory effects of trichostatin A(TSA)on bladder cancer cell lines and its synergetic effect with anticancer drugs in treating bladder cancer in vitro and in vivo.Methods The inhibitory effects of TSA on human bladder cancer cell lines in vitro were detected by MTT assay.Hoechst staining was used to observe morphology for apoptotic cells after TSA treatment.Western blot was used to detect expression of acetyl-histone H3 and survivin.In vivo synergetic effects of TSA with anticancer drugs were detected in bladder cancer model rats.Results TSA significantly inhibited growth of bladder cancer cell lines in concentration and time dependent manner.Better results of tumor inhibition have been achieved when it was combined with DDP,MMC and ADM than used alone.After TSA treatment,the survivin expression in bladder cancer cells decreased and acetyl-histone H3 expression increased.Intravesical application of TSA combined with MMC can significantly inhibited tumor growth and progression.Conclusion TSA has direct anti-cancer effect and can enhance the action of several chemotherapy agents markedly.TSA may be an excellent candidate agent for intravesical application to treat bladder cancer.

Objective To investigate the expression of miR-218 (microRNA-218) in breast cancer tissues and its clinical significance. Methods Totally 45 tissue biopsies gained from breast cancer patients and the adjacent normal tissue were collected. Real-time qPCR was used to detect the miR-218 expressions. The correlations of miR-218 expression with clinicopathological characteristics and prognosis of breast cancer patients were analyzed. Results The average expression level of miR-218 in breast cancer tissues was significantly lower than that in control tissues (P = 0.001). The average expression level of miR-218 in PR negative breast cancer tissues was significantly lower than that of PR positive breast cancer tissue (P = 0.037). The average expression level of miR-218 in Ki-67 positive breast cancer tissues was significantly lower than that in Ki-67 negative breast cancer tissue (P=0.018). Conclusion The expression of miR-218 is closely related to carcinogenesis ,progres-sion and prognosis of breast cancer ,so it may be served as a diagnostic biomarker and a prognostic predictor in breast cancer patients,which provides a new way for the treatment of breast cancer.

Objective To investigate the feasibility, safety and technical superiority of mechanical thrombectomy using a direct aspiration first-pass thrombectomy (ADAPT) in treatment of patients with acute cerebral artery occlusion. Methods A retrospective study was conducted on all patients with acute ischemic stroke treated with mechanical thrombectomy in our institution from January 2013 to August 2016.Patients using ADAPT or stent retriever as a first-line endovascular procedure were compared for clinical characteristics, procedural variables and clinical outcomes. The technical superiority of ADAPT was analyzed in depth. Results During observation period, a total of 91 cases were performed endovascular treatment with mechanical thrombectomy. ADAPT was designed in 46 cases as a first-line endovascular procedure and was utilized in 38 cases (82.6%;ADAPT group), while primary stent retriever thrombectomy was performed in 21 patients(stent group). There was no significant difference in baseline clinical or radiographic factors between ADAPT and stent groups. Although rates of good neurological outcome (modified Rankin Scale(mRS) score≤2) at 90 days were similar between the ADAPT and stent groups (61%(23/38) vs 48%(10/21), P=0.247), National Institute of Health Stroke Scale (NIHSS) score at seven days (6.0(2.0, 9.3) vs 9.0(5.5, 18.5),Z=-2.031,P=0.021) and full recovery rate of neurological outcome (mRS score=0, 37%(14/38) vs 10%(2/21), P=0.022) were significantly better in the ADAPT group than in the stent group. There were no significant differences in rates of embolus to new territory (21%(8/38) vs 29%(6/21), P=0.365), Thrombolysis In Cerebral Infarction (TICI) 2b/3 grade revascularization (84%(32/38) vs 81%(17/21), P=0.507) and symptomatic intracerebral hemorrhage (0%(0/38) vs 10%(2/21), P=0.123) between the ADAPT and the stent groups, but the figures were better in the ADAPT group. Conclusions Mechanical thrombectomy using ADAPT is feasible and safe compared with stent retriever, with higher full recovery rate of neurological outcome and better NIHSS score.It is a method worthy of further exploration for endovascular mechanical recanalization.

BACKGROUND: Chitosan wound dressing has been extensively used in the treatment of wounds and burns, not only because of its bacteriastasis, hemostasis and promoting the wound healing, but also its good biocompatibility,biodegradability and biological functions.OBJECTIVE: To evaluate the efficacy and safety of chitosan wound dressing for deep second-degree burn, thereby providing clinical basis for its registration.METHODS: Sixty patients with deep second-degree burn were randomly allotted to two groups, and then subjected to the external application of chitosan wound dressing (experimental group) or chitosan biomedical dressing (control group),respectively. Then, the therapeutic efficacy was evaluated through healing time, recovering rate and pain scores, and the safety was evaluated through the incidence of adverse reactions and laboratory indexes before and after treatment.RESULTS AND CONCLUSION: There were no significant differences in the wound healing time [(21.23±6.84) days vs.(23.77±4.26) days], recovery rate and pain scores between the experimental and control groups at 14, 21 and 28 days after treatment (P > 0.05). The blood routine, liver function and kidney function indexes before and after treatment did not differ significantly between groups. Additionally, neither adverse nor severe adverse events occurred in the two groups.These results indicate that the therapeutic efficacy and safety of the chitosan wound dressing are equivalent to the control product in the treatment of deep second-degree burn.

BACKGROUND: Both hydroxyapatite (HA) and large diameter TiO2 nanotubes have excellent biocompatibility, but bone-forming ability of nano-HA (nHA) deposited large diameter TiO2 nanotubes is rarely reported.OBJECTIVE: To evaluate the bone-forming ability of nHA/large-diameter TiO2 nanotube composite coating.METHODS: Large-diameter TiO2 nanotubes were prepared by anodic oxidation method, and then nHA was electrochemically deposited on the surface of TiO2 nanotubes. Preosteoblasts MC3T3-E1 were co-cultured with the nHA/large diameter TiO2 nanotube composite, pure titanium and TiO2 nanotube coatings, respectively. At 0.5, 1, 2 hours after culture, the initial cell adhesion was observed. At 1, 3, 5 day after culture, cell proliferation was assessed. At 2 days after culture, cell morphology was observed. At 3 and 7 days after osteogenic induction, intracellular alkaline phosphatase activity was detected. At 14 days after osteogenic induction, mineralization of extracellular matrix was detected.RESULTS AND CONCLUSION: (1) After 2 hours of culture, the number of adherent cells on the composite coating was significantly lower than that on the TiO2 nanotube coating (P < 0.05), but slightly higher than that on the pure titanium coating with no statistical difference. (2) After 1, 3, 5 days of culture, the cell proliferation on the composite coating was significantly lower than that on the TiO2 nanotube coating (P < 0.05), but slightly higher than that on the pure titanium with no statistical difference. (3) The cells on the pure titanium showed a spindle-shape, while those on the TiO2 nanotube coating processed filopodia. The cells on the composite coating showed polygonal shape with a larger number of filopodia. (4) The intracellular alkaline phosphatase activity of the composite coating group was significantly higher than that of the pure titanium group and TiO2 nanotube group. The trend of mineralization of extracellular matrix was ranked from high to low: the composite coating group > TiO2 nanotube group > pure titanium group. To conclude, the nHA/large diameter TiO2 nanotube composite coating not only has good biocompatibility, but also has the ideal ability to promote bone formation.