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BACKGROUND: Now, Shock Wave Therapy is used to cure the ununion and delayed union of bone, the avascular necrosis of the femoral head and the chronic injury of locomotor system, what has got a good curative effect. But, the mechanism is not clear. OBJECTIVE: To investigate the expression of c-Fos, c-Jun protein in human bone marrow mesenchymal stem cells (BMSCs) Influenced by shock wave.DESIGN, TIME AND SETTING: Randomized group, the controlled study was performed at the State Key Laboratory of Pathology, College of Basic Medicine, Jilin University, from March 2007 to March 2008. MATERIALS: Bone marrow was obtained from healthy volunteers.METHODS: Human BMSCs were cultured in vitro. And the fourth generation cells were digested into cell suspension with 2.5 g/L trypsin and adjusted at a density of 1.0×10~9/L with DMEM-LG. Then, the cells were divided into 6 1.5-mL Eerrendorf tubes, one was control group and the other five were experimental groups. The experimental groups were treated with an optimal dose of shock wave (8.5 kV, 120 times) by liquid-electric shock wave lithotripsy. Then the protein was extracted at different time points (5, 15, 30 minutes, 1, 2 hours) after treatment.MAIN OUTCOME MEASURES: Inverted microscope was used to observe the morphologic feature of BMSCs. The growth curves were charted by MTT method. The change of activations of c-Fos, c-Jun were tested by western blot. RESULTS: ①BMSCs were seeded in culture plate. The cells began to divide and proliferate slowly 24 hours later, and became fusiform shape after adhering to the wall. 3 days later, the speed of proliferation quickened, and cells accumulated colony. At day 10-14, the number of hMSCs grew till they covered the bottom of the culture plate. The round passage hMSCs adhered to the wall completely in 24 hours, which were similar to primary cells. The cells connected together during one week, and showed vortex-like. The speed of proliferation became slower and the cells became older when hMSCs were passaged to the tenth generation. ②MTT method showed that the growth curve of original, P2, P3 hMSCs looked like S shape, and the third to fifth days were growing period. ③The phosphorylation level of c-Fos and c-Jun began to increase after induced by shock wave and reached a peak at 30 and 15 minutes, respectively. And their phosphorylation level were 2.56-fold and 1.68-fold (P < 0.01) compared with the average level in control groups, respectively. Then they began to decrease. There was no apparent change in the total dose (P > 0.05). CONCLUSION: Following the treatment of shock wave, the activations of c-Fos and c-Jun in BMSCs increased.
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Thirty-two patients with comminuted proximal humerus fractures(Neer Ⅲ or Ⅳ) were admitted to Department of Orthopedics,First Hospital of Jilin University from March 2001 to April 2006.The patients consisted of 19 males and 13 females,aged 19-70 years,including 15 of 19-49 years and 17 of 50-70 years.All patients underwent treatment of cloverleaf plate-screw fixation.During the follow-up for 8 months,union time and scores from Constant & Murley functional scale were similar between male and female groups.Compared with patients aged 50 years,and the scores from Constant & Murley functional scale were significantly lower(P 50 years old is relatively longer with unfavorable functional recovery in the shoulder joint.
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BACKGROUND:The previous researches indicate that, shock waves can enhance the proliferation of T-cells and the expression of interleukin (IL)-2 through a mechanism that involves p38 mitogen activated protein kinase (MAPK)activation.OBJECTIVE: To investigate if adenosine triphosphate (ATP) release is an underlying mechanism through which low-density shock waves (LDSWs) augment T-cell function.DESIGN: Controlled repetitive measurement by groups, taking cells as subject.SETTING: Department of Orthopedics, the First Hospital of Jilin University.MATERIALS: KDE-2001 Extracorporeal Shockwave Lithotripter (Beijing Zhongke Jian An Meditechs Co., Beijing, China).p38 MAPK inhibitor SB203580 1 mg (BioSource Inc., Camarillo, CA); p38 MAPK kit for detecting phosphorylation (Cell Signaling Technology, Inc. U.S.A.); P2 receptor inhibitor suramin 50 mg (BIOMOL Research Laboratories Inc., PA) was prepared into 0.02 mol/L solution by 1.749 2 mL IMDM. ATP enzyme: apyrase 200 U (Sigma, U.S.A.); P2X7 receptor antagonist KN-62 (BioSource Inc., Camarillo, CA); ATP Bioluminescence Assay Kit CLS Ⅱ (Roche Diagnostics GmbH,Mannheim, Germany).METHODS: The experiment was carried out in the Orthopedic Laboratory of the First Clinical Hospital in Jilin University from January 2005 to December 2006. ①An Extracorporeal Shockwave Lithotripter (at 7 kV generator voltage, 0.3 μF capacitance, 23 MPa positive pressure, 0.18 mJ/mm2 energy flux density) was applied for LDSWs treatment ranging from 50 to 400 impulses. ②ATP release into the culture supernatant from Jurkat T-cells or human peripheral blood mononuclear cells (PBMCs) was determined with a specific ATP Bioluminescence Assay Kit. ③Negative control group excluded antagonist or inhibitor. Human PBMCs were used to determine the effect of LDSWs on activated T-lymphocyte proliferation. Human Jurkat cells were used to study the effects of LDSWs on IL-2 expression. Expression and phosphorylation of p38 MAPK in Jurkat T-cell were measured by Western Immunoblotting with anti-p38 MAPK antibodies and anti-p38 MAPK phospho-specific antibodies that recognized the phosphorylation (on Thr180/Tyr182).MAIN OUTCOME MEASURES: extra-cellular ATP release, IL-2 expression in cell suspension, cellular proliferation and the phosphorylation of p38 MAPK.RESULTS: ①ATP release under the condition without LDSWs was obviously lower than that with LDSWs of 100, 150,200, 250, 300, 360 and 400 impulses (P ＜ 0.01), and ATP release increased with the LDSWs impulse.②Compared with negative control group, the additions of apyrase, KN-62 or suramin attenuated the 3H-TdR incorporation of the phytohemagglutinin-stimulated PBMCs or CD3/CD28-stimulated Jurkat T-cells, which were effected with LDSWs of 100,150, 200, 250, 300, 330 impulses at 0.18 mJ/mm2 (P＜ 0.01). IL-2 expression in the cellular supernatant was also significant increased (P ＜ 0.01). ATPase, KN-62 or suramin all decreased the effect of LDSWs on p38 MAPK of Jurkat T-cells.CONCLUSION: ①LDSWs deform cellular membranes but have no effect on organelle, which results in ATP release from Jurkat cells. Exogenous ATP release activates P2X7 receptor and p38 MAPK, and increases IL-2 expression. LDSWs enhance T-cell proliferation and IL-2 expression through a mechanism that involves ATP release, P2X7 receptor and phosphorylized p38 MAPK activation. ②The release of ATP plays a key role in the mechanism through which LDSWs regulate the function of T cells.
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Objective To investigate the effects of mitogen-activated protein kinase p38(p38 MAPK)on CD3/CD28-stimulated T-cell proliferation and IL-2 expression which were enhanced by shockwaves.Methods Jurkat T cells or peripheral blood mononuclear cells(PBMC)were pretreated with the p38 MAPK-inhibitor(SB203580)(The Jurkat T cells or PBMC of the control groups were not pretreated with SB203580),then treated with shockwaves,and stimulated respectively with a suboptimal dose of anti-CD3 and anti-CD28 antibodies or PHA.Finally the IL-2 productions of Jurkat T cells or the proliferation of PBMC were respectively measured.The protein tyrosine phosphorylation of p38 MAPKin Jurkat T cells treated either with shockwaves or not was measured by Western blotting with anti-phosphotyrine antibodies(Thr180/Tyr182).The expressoins of p38 MAPK in Jurkat T cells treated either with shockwaves or not were determined by Western blotting with anti-p38 MAPK antibodies.Results Compared with negative control group without shockwave treatment,the 3H-TdR incorporation of the phytohemagglutinin-stimulated PBMC,which were treated with low dose shockwaves(LDSWs)of 100,150,200,250,300,330 impulses at(0.180?0.015)mJ?mm2,increased significantly(P
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Objective To investigate the reveral effect of Tanshitone(Tan) Microemulsion on multidrug resistance(MDR) of sensitive K562 cell line(KS) and ADM resistant K562 cell line(KA) and the mechanism.Methods The reversal effect of Tan Microemulsion(Tan and Microemulsion as controls) on MDR of KA cells was observed.The inhibitory effect of cell growth was investigated by MTT.The apoptosis of KS and KA cell lines was observed by flow cytometry.The intracellular drug concentration was measured by detecting fluorescent density.Results The result showed that Tan Microemulsion,Microemulsion and Tan reduced the expression levels of P-gp and Bcl-2 in KA cells and Tan Microemulsion had the most significant effect.The MDR reversal folds of Tan Microemulsion of KA was highest among all 3 groups(P
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BACKGROUND: Multidrug resistance (MDR) makes the treatment of tumor more difficult. Many factors contribute to the happening of MDR. The combination of two or more kinds of reverse reagent at the same time may be more effective to MDR.OBJECTIVE: To investigate the reversal effect of Tanshinone Microemulsion on MDR of human leukemia cell line (K562) and ADM resistant cell line (K562/ADM) cell line and the mechanism of it.DESIGN: An observational controlled experiment.SETTING: First Affiliated Hospital's Central Laboratory of Jilin University.MATERIALS: K562 and K562/ADM (Tianjin Blood Research Institute of CAS). Tanshinone and Tanshinone Microemulsion (Pharmaceutical College of Jilin University). ADM(Pfizer pharmaceutical corp). P-gp-PE fluorescent antibody and Bcl-2-FITC fluorescent antibody (Immunotech Biotech corp).DMSO(Beijing Chemical Plant).METHODS: The experiment was carried out in the Central Laboratory of First Hospital ffiliated to Jilin University from March 2003 to January 2004. ① K562 was cultured in liquid medium(RMPI1640) which contains 100 g/L calf serum, 100 U/mL penicillin and streptomycin. The culture tank's environment was kept at 37 ℃, with saturated humidity and 5%CO2. K562/ADM was cultured in the same environment as above, but ADM was added to the RMPI1640. ② The reverse effect of Tanshinone Microemulsion (Tanshinone and Microemulsion as control) on K562/ADM cells' MDR is observed. ③Tanshinone Microemulsion,Tanshinone and Microemulsion's MDR reverse fold to K562 cell line was detected by the method of MTT. ④ ADM of different concentrations (10, 20, 40, 60 mg/L)were added to grouped cells of exponential proliferation period. The intracellular drug concentration was observed by detecting fluorescent density.⑤ Expression rate of P-gp, Bcl-2 and apoptosis ratio of K562 and K562/ADM added with Tanshinone Microemulsion, Tanshinone and Microemulsion cell line were observed by the method of Flow Cytometry.MAIN OUTCOME MEASURES: ①Tanshinone Microemulsion, Tanshinone and Microemulsion's MDR reverse fold to K562 cell line. ②Tanshinone Microemulsion, Tanshinone and Microemulsion's effect to K562 cell's intracellular contration.③Tanshinone Microemulsion, Tanshinone and Microemulsion's influence to K562 cell's expression rate of P-gp, Bcl-2 and apoptosis ratio.RESULTS: ①Tanshinone Microemulsion, Tanshinone and Microemulsion could decrease the MDR of the K562/ADM. The effect of Tanshinone Microemulsion were significantly higher than Tanshinone and Microemulsion(P ＜ 0.05 ).②The intracellular concentration of ADM can be improved by Tanshinone Microemulsion, Microemulsion and Tanshinone. The ADM's concentration of K562/ADM treated by Tanshinone Microemulsion is highest and the difference is significant(P＜ 0.05). ③Tanshinone Microemulsion, Tanshinone and Microemulsion could reduce the expression level of P-gp and Bcl-2 in K562/ADM, and increase the apoptosis percentage of tumor cell. Tanshinone Microemulsion has the most significant effect on it,andincreases the apoptosis percentage of tumor cell with ADM (P ＜ 0.05 ).CONCLUSION: Tanshinone Microemulsion (0.5 mg/L) can increase the intracellular concentration of ADM, which is consist with the conclusion of down regulation of the expressions of P-gp and Bcl-2. Reversal effect of Tanshinone Microemulsion is more effective than that of Tanshinone and Microemulsion.
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Objective To study the properties of stress relaxation of the middle and lower cervical vertebrae, and to evaluate the effects of laminotomy and anterior diskectomy and spinal fusion (ASF) on the properties . Methods The dynamic responses of stress relaxation of six intact and post operative fresh human cadaveric cervical vertebrae were measured in vitro. Results Under the condition of the constant strain, the functional equations and the curves of stress relaxation of the six intact and post operative fresh human cadaveric cervical vertebrae were obtained. The Gex(t)(the ratio of the ensuing instant stress to the original stress) of the post operative ones was significantly bigger than that of the intact ones; The Gex(t) of the laminotomic ones was significantly bigger than that of the ones having undergone the anterior discectomy and spinal fusion(ASF). Conclusion Either in flexion or in extension, cervical vertebrae have the similar behaviors of stress relaxation. The laminotomy and anterior discectomy and spinal fusion (ASF) all reduce the stress relaxation effect of the cervical vertebrae, but ASF is more significant.