OBJECTIVE: To establish an SPE(solid phase extraction)-HPLC method for the determination of diclofenac sodium in human plasma.METHODS: The separation of diclofenac sodium was performed on ODS-2 column with a detection wavelength of 280nm.The mobile phase was composed of phosphate buffer(pH6.5)-methanol(40∶ 60) at a flow rate of 1.0mL? min-1.RESULTS: The linear range of diclofenac sodium was 16.2～ 2 024.0ng? mL-1(r=0.998 9).The methodological recovery ranged between 90% to 110% and the extraction recovery was above 90%.Both intraday RSD and inter-day RSD were less than 5%.CONCLUSION: The method is simple,accurate and reliable,and suitable for the pharmacokinetic study of diclofenac sodium.

Nerve growth factor (NGF) has been the focus in area of angiogenesis.In differential,survival,development of nerves,NGF plays a crucial role.In addition,NGF activates migration,differential of endothelial cells and vascular smooth muscle cells,and stimulate the activation of the endothelial progenitor cells in angiogenesis.Crosstalk between vascular endothelial growth factor and NGF is also critical in the development of vessels.The function of NGF in angiogenesis blazes a trail in therapy of ischemia diseases.

AIM:ToexploretheeffectofcelastrolonapoptosisofSaos-2cellsanditsmechanism.METH-ODS:Saos-2 cells were treated with various concentrations of celastrol , and the cell viability was measured by MTT assay . Apoptosis and reactive oxygen species ( ROS) production were determined by flow cytometry .The protein levels of cleaved caspase-9, cleaved caspase-3 and phosphorylated JNK were evaluated by Western blot .RESULTS:The viability of Saos-2 cells was significantly inhibited by celastrol .Celastrol significantly induced apoptosis of Saos-2 cells.Celastrol signifi-cantly induced ROS production in the Saos-2 cells.Western blot analysis demonstrated that celastrol significantly increased the protein levels of cleaved caspase-9, cleaved caspase-3 and phosphorylated JNK in the Saos-2 cells.CONCLUSION:Celastrol induces caspase-dependent apoptosis through ROS/JNK pathway in Saos-2 cells.

Objective] The two-step massage treatment of tennis elbow and simple massage therapy clinical curative effect around the elbow. [Method] A total of 80 cases of external humeral epicondylitis were randomly divided into the treatment group(42 cases) and control group(38 cases). The treatment group were treated with the combination of the neck and elbows. The control group was treated only with simple elbow massage. [Results] The treatment group cure rate was 71.4%, the total efficiency of 92.9%;control group, the cure rate was 34.2%, the total effective rate was 71%. The comparison between the 2 groups, the cure rate and total effective rate differences were significant( P＜0.01, P＜0.05). [Conclusion]Two-step massage has better cure effect on external humeral cpicondylitis.

Objective\ To evaluate the immediate effects and strength of allograft fusion cage(AFC) used for reconstructing stability of cervical spine. Methods\ Discs of C 5 and C 6 were resected on 8 fresh human cervical spine specimens, and autogenous iliac bone grafts(AIBG), and AFCs were implanted into the intervertebral spaces respectively. Compression test, pull out test and segmental motion measurement were studied.Results\ Comparing with intact and AIBG groups, the range of motions of C5-6 in AFC group were decreased in all directions except for extension; with (502?114) N compressive load,the vertebrae in AFC group were broken while the AFC were intact, but the AIBG were broken at (135?42) N load; with 300 N drawing load,no loosening was found between AFCs and vertebra, but it was found between AIBG and vertebra at 60 N load. Conclusion\ AFC could provide enough support,anti slide ability and could remain or increase the height of intervertebral spaces. It completely meets clinical and biomechanical requirements.

Objective To evaluate the effects of allograft fusion cage(AFC) on anterior cervical interbody fusion. Methods AFCs were implanted in 61 degenerative cervical intervertebral spaces of 39 cases, who needed anterior cervical interbody fusion from September 1995 to December 1999. 31 cases were diagnosed as cervical spondylolysis， 2 cases as acute protrusion of cervical intervertebral disc and 6 cases as fracture and dislocation of cervical spine. The clinical effects and complications were observed, and the postoperative presentations of X－ ray examination of cervical spine were also evaluated. Results Thirty- nine cases were followed up with a mean period of 28.6 months. No neurologic complications appeared, and no AFCs shifted or dislocated. The clinical effects were satisfactory. 61 intervertebral spaces were confirmed to be solid fused completely by constant X－ ray examination at 3.9 months in average after operation. There were no collapse or angular deformities in 59 spaces of them,the other 2 spaces lost a little height because of removal of external fixation too early. Conclusion The implantation of AFC was simple, stable, less injury with similar intervertebral osseous fusion rate compared to the conventional anterior cervical interbody fusion. Forthermore, the implantation of AFC does not need auto iliac crest graft or the use of metal fixations. Some complications caused by implanting auto iliac crest and metal fixations can be avoided.

Aim To develop a convenient and effective method to isolate and culture primary rabbit aortic vascular smooth muscle cells(VSMCs).Methods The thoracic aortas were removed by dissection under sterile conditions.Aortic smooth muscle cells were excised and cleaned of fat and connective tissue,and the isolated vessel media was cut into 1 mm3 pieces.The explants were digested with different concentrations of collagenase typeⅠ,and incubated at 37℃ for different time,then undispersed explants were placed onto a sterile 100-mm plastic tissue culture dish with growth medium.Results VSMCs could emigrate from the explants digested 6 h by collagenase typeⅠ(1.5 g?L-1)for 24 hours,the cells would passage for another 4~5 days.Confluency could be reached within 3~4 days after subculturing.VSMCs were identified by immunoreactivity with ?-actin and by the smooth muscle cell-specific,hills and valley-like morphology.Conclusion It was an effective method to culture primary VSMCs from the explants digested for 6h by collagenase type Ⅰ(1.5 g?L-1),which could shorten primary culture time.

Aim To develop a convenient and effective method to isolate and culture primary rat aortic endothelial cells (RAECs). Methods The thoracic aortas were removed by dissection under sterile conditions. Aortas were turned over to expose the luminal surface, and the surfaces were digested with different concentrations of collagenase typeⅠ, incubated at 37℃ for different times, then, cut into pieces and placed luminal side down onto collagen-coated flask with growth medium. Results RAECs could emigrate from explants digested 1h by collagenase typeⅠ(2.0 g?L-1) for 24 h and cells would passage for another 4~5 days. Reached confluency within 3~4 d after subculturing. RAECs were identified by immunoreactivity with Factor-Ⅷ and by the endothelial cell-specific, cobblestone-like morphology. Conclusion It is an effective method to culture primary RAECs from explants digested for 1h by collagenase type Ⅰ(2.0 g?L-1),that can shorten primary culture time.

Objective To compare the survival conditions of patients with insulinoma after enucleation of insulinoma or partial resection of pancreas.Methods The clinical data of 99 patients with insulinoma,treated with surgery from May.2003 to Aug.2015 were retrospectively analyzed.Of the 99 patients,38 received enucleation of insulinoma alone and 61 received partial resection of pancreas.The overall data were analyzed by SPSS 21.0 software.Results Average survival of patients after enucleation of insulinoma (103.3 months) was longer than that of patients after partial resection of pancreas (77.5 months),and the difference had statistical significance (P=0.006).The difference of the incidence of most chronic or temporary complications had no statistical significance between the two groups (P＞0.05),except for new-onset diabetes (P=0.004).Conclusion Enucleation of insulinoma should be firstly recommended for patients with insulinoma in suitable size,which can provide patients with better survival condition.

[Objective] To determine the effects and possible mechanism of the thrombin antagonist r-RGD-Hirudin (HIR) on ventricular arrhythmia(VA) after acute myocardial infarction (AMI). [Methods] Seventy adult male Sprague-Dawley rats were randomly subjected to the 10 groups according to duration of left coronary occlusion: HIR 0 min, HIR 5 rain, HIR 10 min, HIR 20 min, HIR 30 min, and normal saline(NS) 0 min, NS 5 min, NS 10 min, NS 20 min, NS 30 min; and the average of every group is 7 rats. Acute myocardial infarction was produced by the occlusion of the left anterior descending coronary artery, then the measurements of arrhythmia and infarction sizing by Evans blue were assessed as well as the expression of three isoforms of inositol 1,4,5-trisphosphate receptors (IP3Rs) mRNA in isehemic myocardium by reverse transeriptase polymerase chain reactions (RT-PCR). [Results] Compared with NS groups, the measurements of VA in HIR were reduced significantly in 5 to 20 minutes after AMI (P<0.05). The incidence of VA was all positive related to the expression of three isoforms of IP3Rs mRNA (P<0.01). Compared with NS groups, the expression of type2,inositol 1,4,5-trisphosphate receptor (IP3R2) mRNA at 10 min and type3, inositol 1,4,5-trisphosphate receptor mRNA (IP3R3) at 10 min and 20 min after AMI were significant decreased (P<0.05) in HIR groups. [Conclusion] The thrombin antagonist r-RGD-Hirudin exerts its myocardial protection against ventricular arrhythmia after acute myocardial infarction possible through IP3R2 and IP3R3 and not typel, inositol 1,4,5-trisphosphate receptor (IP3R1).