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1.
目的 研究外源性神经生长因子(NGF)对QBC939细胞67-kDa层黏连蛋白受体(67LR)表达的影响,探讨胆管癌神经浸润的机制.方法 (1)细胞免疫荧光染色法检测QBC939细胞表面NGF高亲和力酪氨酸激酶受体A(TrkA)表达情况.(2)用不同浓度的重组人神经生长因子(β-NGF)对QBC939细胞进行处理,采用Real-Time PCR和Western blot检测QBC393细胞67LR mRNA和蛋白表达情况.实验分为对照组、β-NGF(1、10、100、200μg/L)组.(3)根据上述实验结果选定最佳β-NGF浓度,再加入不同浓度的TrkA阻断剂K252a,再次检测QBC939细胞67LR mRNA和蛋白表达情况.实验分为对照组、100μg/Lβ-NGF组、K252a(100、200、300 nmol/L)阻断组.数据采用单因素方差分析,两两比较采用LSD法.结果 (1)QBC939细胞膜上TrkA表达明显.(2)对照组和β-NGF(1、10、100、200 μg/L)组QBC939细胞67LRmRNA和蛋白表达水平分别为0.35±0.06、0.38±0.14、0.62±0.14、0.90±0.08、0.70±0.10和0.32±0.05、0.50±0.09、0.69±0.13、0.93±0.07、0.76±0.07,两者比较,差异有统计学意义(F=22.4,14.6,P<0.05),其中100ng/ml β-NGF作用最明显(t=19.0,21.0,P<0.05).(3)对照组、100 μg/L β-NGF组、K252a(100、200、300 nmol/L)阻断组QBC939细胞67LR mRNA和蛋白表达水平分别为0.35±0.10、0.88±0.14、0.80±0.08、0.67±0.12、0.43±0.07和0.41±0.10、0.84±0.10、0.76±0.04、0.61±0.09、0.50±0.12,两者比较,差异有统计学意义(F=14.1,8.9,P<0.05),其中经300 nmol/L K252a处理后,QBC939细胞67LR mRNA和蛋白表达水平与对照组比较,差异无统计学意义(t=1.02,0.85,P>0.05).结论 外源性NGF通过与其高亲和力受体TrkA结合,可增强QBC939细胞67LR表达,这可能是促进胆管癌细胞延外周神经纤维间隙浸润的机制之一.
Abstract:
Objective To investigate the effects of nerve growth factor (NGF) on the expression of 67-kDa laminin receptor (67LR) in human bile duct carcinoma QBC939 cells, and study the possible mechanism of perineural invasion and metastasis of bile duct carcinoma. Methods ( 1 ) The expression of a high-affinity receptor for NGF, TrkA, was detected by immunofluorescence staining. ( 2 ) QBC393 cells were pretreated by β-NGF at different concentrations ( 1, 10, 100,200 μg/L), and then the mRNA and protein expressions of 67LR were examined by Real-Time PCR and Western blot assay. QBC939 cells were divided into control group and β-NGF (1, 10, 100,200 μg/L) groups. (3) The ideal concentration of β-NGF was selected according to the results of previous tests, and then the mRNA and protein expressions of 67LR were re-examined by adding specific TrkA inhibitor K252a at different concentrations ( 100,200,300 nmol/L). QBC939 cells were divided into control group, β-NGF 100 μg/L group and K252a ( 100,200,300 nmol/L) groups. All data were analyzed by one-way analysis of variance or LSD-test. Results (1) A strong expression of TrkA was detected in the membrane of QBC939 cells. (2) The mRNA and protein expressions of 67LR in QBC939 cells were 0.35 ± 0.06 and 0. 32 ± 0.05 in the control group, 0.38 ±0.14 and 0.50 ±0.09 in the β-NGF 1 μg/L group, 0.62 ±0.14 and 0. 69 ±0. 13 in β-NGF 10 μg/L group, 0.90 ± 0.08 and 0.93 ± 0.07 in the β-NGF 100 μg/L group, and 0. 70 ± 0. 10 and 0. 76 ±0.07 in the β-NGF 200 μg/L group, there were significant differences among the five groups (F = 22. 4, 14. 6,P <0.05). The mRNA and protein expressions of 67LR in the β-NGF 100 μg/L group were significantly higher than those in the control group ( t = 19. 0, 21.0, P < 0. 05 ). (3) The mRNA and protein expressions of 67LR in the QBC939 cells were 0.35 ±0.10 and 0.41 ±0.10 in the control group, 0. 88 ±0. 14 and 0.84 ±0.10 in the β-NGF 100 μg/L group, 0.80±0.08 and 0.76 ±0.04 in the K252a 100 nmol/L group, 0.67 ±0.12 and 0.61 ± 0.09 in the K252a 200 nmol/L group, and 0. 43 ± 0.07 and 0. 50 ± 0. 12 in the K252a 300 nmol/L group, there were significant differences among the five groups ( F = 14. 1, 8. 9, P < 0.05 ). There were no significant differences in the mRNA and protein expressions of 67LR between the K252a 300 nmol/L group and the control group (t =1.02, 0. 85, P>0.05). Conclusion In bile duct carcinoma cells, NGF enhances the expression of 67LR by combining with TrkA, which might be the mechanism of NGF mediating perineural invasion of bile duct carcinoma.  相似文献   

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Objective To investigate the effect of resveratrol on the proliferation and invasion of human pancreatic cancer PANC-1 cells. Methods Five groups including blank control group, 0. 1% dimethylsulfoxide (DMSO) group and resveratrol groups (50, 100, 200 μmol/L) were established. The proliferation of PANC-1 cells was detected by MTT assay. The apoptosis and cell cycle change were analyzed by flow cytometry. The invasive ability of PANC-1 cells was observed with a Transwell cell culture chamber. The expressions of Bax, Bcl-2,matrix metalloproteinases 2 (MMP-2) and 9 (MMP-9) of the PANC-1 cells were assayed by real-time quantitative PCR and Western blot. All data were analyzed using the analysis of variance. Results ( 1 ) The inhibition rate of resveratrol on the proliferation of PANC-1 cells was 0 in the blank control group, 3.25% ±0.42% in the 0. 1% DMSO group, 13.23% ± 1.68% in the 50 μmol/L of resveratrol group, 42.25% ± 3.20% in the 100 μmol/L of resveratrol group, and 56.94% ±5.31% in the 200 μmol/L of resveratrol group. There was a significant difference in the inhibition rate among the five groups (F=460. 10, P<0.05). (2) The apoptosis rate was 0.05% ±0.03% in the blank control group, 3.39% ± 1.77% in the 0. 1% DMSO group, 6.92% ± 1.85% in the 50 μmol/L of resveratrol group, 19.05% ± 2.01% in the 100 μmol/L of resveratrol group, and 27. 17% ±6.43% in the 200 μmol/L of resveratrol group. There was a significant difference in the apoptosis rate among the five groups (F = 38.84, P < 0.05). (3) There was no significant effect of 0. 1% DMSO on the cell cycle of PANC-1 cells. The number of PANC-1 cells in the G0/G1 and S phase was increased. (4) The average number of invading PANC-1 cells was 61 ± 13 in the blank control group, 54 ± 13 in the 0. 1% DMSO group, 48 ± 15 in the 50 μmol/L of resveratrol group, 23 ±6 in the 100 μ mol/L of resveratrol group and 18 ±7 in the 200 μmol/L of resveratrol group. There was a significant difference in the number of invading PANC-1 cells among the five groups (F = 69.08, P < 0.05 ). (5) There were up-regulated mRNA and protein expressions of Bax and down-regulated mRNA and protein expressions of Bcl-2, and the expressions of MMP-2 and MMP-9 of the PANC-1 cells were inhibited in the resveratrol groups. The changes of the protein expressions of Bax, Bcl-2, MMP-2, MMP-9 were consistent with the changes of the mRNA expressions of the four indexes. Conclusion Resveratrol can significantly inhibit the proliferation and invasion, as well as induce apoptosis of PANC-1 cells in vitro.  相似文献   

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Objective To construct β-NGF controlled delivery system and evaluate the biological effects of β-NGF on the growth of chick embryo dorsal root ganglion (DRG) axons in vitro. Methods Delivery systems releasing β-NGF at 50/μL, 100/μg/L and 250 μg/L concentration were constructed. To determine the optimal dose response effects of NGF in the controlled delivery system, DRG were co-cultured with of β-NGF at above concentrations while using DRG basic culture as control. Axonal growth was observed. DRG were also cocultured with the components in the controlled delivery system to detect the effects on growth of DRG axons. The experiment was divided into 5 experimental groups and 1 control group: control group, DRG+ fibrin; Group A,DRG+ fibrin+ peptide + heparin + 100 μg/L β-NGF; Group B, DRG + fibrin + heparin + 100/μg/L β-NGF;Group C, DRG + fibrin + peptide + 100 μg/L β-NGF; Group D, DRG + fibrin + 100 μg/L β-NGF; Group E,DRG + fibrin + peptide + heparin. Results The growth of DRG axons in 50 μg/L, 100μg/L and 250/μg/Lconcentration of β-NGF controlled delivery system was 1.31 ( P > 0. 05), 3.78 ( P < 0. 01 ) and 3.05 ( P <0.01) folds of the control respectively. The growth of DRG axons in 100 μg/L group was significantly better comparing to that in 250 μg/L group. The growth of DRG axons in Groups A, B, C, D and E was 3.75, 1.15,1.12, 1.10 and 1.09 folds of the control group, respectively. The difference was only statistically significant between Group A and the control group ( P < 0. 01 ). Conclusion β-NGF released from the β-NGF controlled delivery system was bioactive. It could promote the growth of DRG axons.  相似文献   

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Objective To construct β-NGF controlled delivery system and evaluate the biological effects of β-NGF on the growth of chick embryo dorsal root ganglion (DRG) axons in vitro. Methods Delivery systems releasing β-NGF at 50/μL, 100/μg/L and 250 μg/L concentration were constructed. To determine the optimal dose response effects of NGF in the controlled delivery system, DRG were co-cultured with of β-NGF at above concentrations while using DRG basic culture as control. Axonal growth was observed. DRG were also cocultured with the components in the controlled delivery system to detect the effects on growth of DRG axons. The experiment was divided into 5 experimental groups and 1 control group: control group, DRG+ fibrin; Group A,DRG+ fibrin+ peptide + heparin + 100 μg/L β-NGF; Group B, DRG + fibrin + heparin + 100/μg/L β-NGF;Group C, DRG + fibrin + peptide + 100 μg/L β-NGF; Group D, DRG + fibrin + 100 μg/L β-NGF; Group E,DRG + fibrin + peptide + heparin. Results The growth of DRG axons in 50 μg/L, 100μg/L and 250/μg/Lconcentration of β-NGF controlled delivery system was 1.31 ( P > 0. 05), 3.78 ( P < 0. 01 ) and 3.05 ( P <0.01) folds of the control respectively. The growth of DRG axons in 100 μg/L group was significantly better comparing to that in 250 μg/L group. The growth of DRG axons in Groups A, B, C, D and E was 3.75, 1.15,1.12, 1.10 and 1.09 folds of the control group, respectively. The difference was only statistically significant between Group A and the control group ( P < 0. 01 ). Conclusion β-NGF released from the β-NGF controlled delivery system was bioactive. It could promote the growth of DRG axons.  相似文献   

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Objective To construct β-NGF controlled delivery system and evaluate the biological effects of β-NGF on the growth of chick embryo dorsal root ganglion (DRG) axons in vitro. Methods Delivery systems releasing β-NGF at 50/μL, 100/μg/L and 250 μg/L concentration were constructed. To determine the optimal dose response effects of NGF in the controlled delivery system, DRG were co-cultured with of β-NGF at above concentrations while using DRG basic culture as control. Axonal growth was observed. DRG were also cocultured with the components in the controlled delivery system to detect the effects on growth of DRG axons. The experiment was divided into 5 experimental groups and 1 control group: control group, DRG+ fibrin; Group A,DRG+ fibrin+ peptide + heparin + 100 μg/L β-NGF; Group B, DRG + fibrin + heparin + 100/μg/L β-NGF;Group C, DRG + fibrin + peptide + 100 μg/L β-NGF; Group D, DRG + fibrin + 100 μg/L β-NGF; Group E,DRG + fibrin + peptide + heparin. Results The growth of DRG axons in 50 μg/L, 100μg/L and 250/μg/Lconcentration of β-NGF controlled delivery system was 1.31 ( P > 0. 05), 3.78 ( P < 0. 01 ) and 3.05 ( P <0.01) folds of the control respectively. The growth of DRG axons in 100 μg/L group was significantly better comparing to that in 250 μg/L group. The growth of DRG axons in Groups A, B, C, D and E was 3.75, 1.15,1.12, 1.10 and 1.09 folds of the control group, respectively. The difference was only statistically significant between Group A and the control group ( P < 0. 01 ). Conclusion β-NGF released from the β-NGF controlled delivery system was bioactive. It could promote the growth of DRG axons.  相似文献   

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Objective To construct β-NGF controlled delivery system and evaluate the biological effects of β-NGF on the growth of chick embryo dorsal root ganglion (DRG) axons in vitro. Methods Delivery systems releasing β-NGF at 50/μL, 100/μg/L and 250 μg/L concentration were constructed. To determine the optimal dose response effects of NGF in the controlled delivery system, DRG were co-cultured with of β-NGF at above concentrations while using DRG basic culture as control. Axonal growth was observed. DRG were also cocultured with the components in the controlled delivery system to detect the effects on growth of DRG axons. The experiment was divided into 5 experimental groups and 1 control group: control group, DRG+ fibrin; Group A,DRG+ fibrin+ peptide + heparin + 100 μg/L β-NGF; Group B, DRG + fibrin + heparin + 100/μg/L β-NGF;Group C, DRG + fibrin + peptide + 100 μg/L β-NGF; Group D, DRG + fibrin + 100 μg/L β-NGF; Group E,DRG + fibrin + peptide + heparin. Results The growth of DRG axons in 50 μg/L, 100μg/L and 250/μg/Lconcentration of β-NGF controlled delivery system was 1.31 ( P > 0. 05), 3.78 ( P < 0. 01 ) and 3.05 ( P <0.01) folds of the control respectively. The growth of DRG axons in 100 μg/L group was significantly better comparing to that in 250 μg/L group. The growth of DRG axons in Groups A, B, C, D and E was 3.75, 1.15,1.12, 1.10 and 1.09 folds of the control group, respectively. The difference was only statistically significant between Group A and the control group ( P < 0. 01 ). Conclusion β-NGF released from the β-NGF controlled delivery system was bioactive. It could promote the growth of DRG axons.  相似文献   

7.
Objective To construct β-NGF controlled delivery system and evaluate the biological effects of β-NGF on the growth of chick embryo dorsal root ganglion (DRG) axons in vitro. Methods Delivery systems releasing β-NGF at 50/μL, 100/μg/L and 250 μg/L concentration were constructed. To determine the optimal dose response effects of NGF in the controlled delivery system, DRG were co-cultured with of β-NGF at above concentrations while using DRG basic culture as control. Axonal growth was observed. DRG were also cocultured with the components in the controlled delivery system to detect the effects on growth of DRG axons. The experiment was divided into 5 experimental groups and 1 control group: control group, DRG+ fibrin; Group A,DRG+ fibrin+ peptide + heparin + 100 μg/L β-NGF; Group B, DRG + fibrin + heparin + 100/μg/L β-NGF;Group C, DRG + fibrin + peptide + 100 μg/L β-NGF; Group D, DRG + fibrin + 100 μg/L β-NGF; Group E,DRG + fibrin + peptide + heparin. Results The growth of DRG axons in 50 μg/L, 100μg/L and 250/μg/Lconcentration of β-NGF controlled delivery system was 1.31 ( P > 0. 05), 3.78 ( P < 0. 01 ) and 3.05 ( P <0.01) folds of the control respectively. The growth of DRG axons in 100 μg/L group was significantly better comparing to that in 250 μg/L group. The growth of DRG axons in Groups A, B, C, D and E was 3.75, 1.15,1.12, 1.10 and 1.09 folds of the control group, respectively. The difference was only statistically significant between Group A and the control group ( P < 0. 01 ). Conclusion β-NGF released from the β-NGF controlled delivery system was bioactive. It could promote the growth of DRG axons.  相似文献   

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Objective To construct β-NGF controlled delivery system and evaluate the biological effects of β-NGF on the growth of chick embryo dorsal root ganglion (DRG) axons in vitro. Methods Delivery systems releasing β-NGF at 50/μL, 100/μg/L and 250 μg/L concentration were constructed. To determine the optimal dose response effects of NGF in the controlled delivery system, DRG were co-cultured with of β-NGF at above concentrations while using DRG basic culture as control. Axonal growth was observed. DRG were also cocultured with the components in the controlled delivery system to detect the effects on growth of DRG axons. The experiment was divided into 5 experimental groups and 1 control group: control group, DRG+ fibrin; Group A,DRG+ fibrin+ peptide + heparin + 100 μg/L β-NGF; Group B, DRG + fibrin + heparin + 100/μg/L β-NGF;Group C, DRG + fibrin + peptide + 100 μg/L β-NGF; Group D, DRG + fibrin + 100 μg/L β-NGF; Group E,DRG + fibrin + peptide + heparin. Results The growth of DRG axons in 50 μg/L, 100μg/L and 250/μg/Lconcentration of β-NGF controlled delivery system was 1.31 ( P > 0. 05), 3.78 ( P < 0. 01 ) and 3.05 ( P <0.01) folds of the control respectively. The growth of DRG axons in 100 μg/L group was significantly better comparing to that in 250 μg/L group. The growth of DRG axons in Groups A, B, C, D and E was 3.75, 1.15,1.12, 1.10 and 1.09 folds of the control group, respectively. The difference was only statistically significant between Group A and the control group ( P < 0. 01 ). Conclusion β-NGF released from the β-NGF controlled delivery system was bioactive. It could promote the growth of DRG axons.  相似文献   

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Objective To construct β-NGF controlled delivery system and evaluate the biological effects of β-NGF on the growth of chick embryo dorsal root ganglion (DRG) axons in vitro. Methods Delivery systems releasing β-NGF at 50/μL, 100/μg/L and 250 μg/L concentration were constructed. To determine the optimal dose response effects of NGF in the controlled delivery system, DRG were co-cultured with of β-NGF at above concentrations while using DRG basic culture as control. Axonal growth was observed. DRG were also cocultured with the components in the controlled delivery system to detect the effects on growth of DRG axons. The experiment was divided into 5 experimental groups and 1 control group: control group, DRG+ fibrin; Group A,DRG+ fibrin+ peptide + heparin + 100 μg/L β-NGF; Group B, DRG + fibrin + heparin + 100/μg/L β-NGF;Group C, DRG + fibrin + peptide + 100 μg/L β-NGF; Group D, DRG + fibrin + 100 μg/L β-NGF; Group E,DRG + fibrin + peptide + heparin. Results The growth of DRG axons in 50 μg/L, 100μg/L and 250/μg/Lconcentration of β-NGF controlled delivery system was 1.31 ( P > 0. 05), 3.78 ( P < 0. 01 ) and 3.05 ( P <0.01) folds of the control respectively. The growth of DRG axons in 100 μg/L group was significantly better comparing to that in 250 μg/L group. The growth of DRG axons in Groups A, B, C, D and E was 3.75, 1.15,1.12, 1.10 and 1.09 folds of the control group, respectively. The difference was only statistically significant between Group A and the control group ( P < 0. 01 ). Conclusion β-NGF released from the β-NGF controlled delivery system was bioactive. It could promote the growth of DRG axons.  相似文献   

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Objective To construct β-NGF controlled delivery system and evaluate the biological effects of β-NGF on the growth of chick embryo dorsal root ganglion (DRG) axons in vitro. Methods Delivery systems releasing β-NGF at 50/μL, 100/μg/L and 250 μg/L concentration were constructed. To determine the optimal dose response effects of NGF in the controlled delivery system, DRG were co-cultured with of β-NGF at above concentrations while using DRG basic culture as control. Axonal growth was observed. DRG were also cocultured with the components in the controlled delivery system to detect the effects on growth of DRG axons. The experiment was divided into 5 experimental groups and 1 control group: control group, DRG+ fibrin; Group A,DRG+ fibrin+ peptide + heparin + 100 μg/L β-NGF; Group B, DRG + fibrin + heparin + 100/μg/L β-NGF;Group C, DRG + fibrin + peptide + 100 μg/L β-NGF; Group D, DRG + fibrin + 100 μg/L β-NGF; Group E,DRG + fibrin + peptide + heparin. Results The growth of DRG axons in 50 μg/L, 100μg/L and 250/μg/Lconcentration of β-NGF controlled delivery system was 1.31 ( P > 0. 05), 3.78 ( P < 0. 01 ) and 3.05 ( P <0.01) folds of the control respectively. The growth of DRG axons in 100 μg/L group was significantly better comparing to that in 250 μg/L group. The growth of DRG axons in Groups A, B, C, D and E was 3.75, 1.15,1.12, 1.10 and 1.09 folds of the control group, respectively. The difference was only statistically significant between Group A and the control group ( P < 0. 01 ). Conclusion β-NGF released from the β-NGF controlled delivery system was bioactive. It could promote the growth of DRG axons.  相似文献   

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膝关节后交叉韧带断裂治疗临床分析   总被引:7,自引:3,他引:4  
郭臻伟  杨茂清  朱惠芳 《中国骨伤》2001,14(10):582-584
目的 对35例膝关节后交叉韧带断裂治疗进行临床分析,重点探讨了有关交叉韧带断裂的治疗问题。方法 经明确诊断后,分析采用胫骨附着处撕脱骨折复位固定手术治疗26例、早期髌韧带中1/3移植重建3例、单纯长腿石膏固定6例。结果 本组病例全部进行随访,随访时间13个月-5年,胫骨附着处撕脱骨折复位固定及髋韧带中1/3移植重建29例为优良、单纯长腿石膏固定6例为差。结论 后交叉韧带断裂后应该及时给予手术修复;膝后外侧手术入路,操作简单,暴露充分;少于3个月的陈旧性病例仍适应手术治疗。  相似文献   

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目的:探讨不同方法重建指尖离断静脉回流的疗效。方法:2008年3月-2013年2月收治指尖离断患者80例,38例吻合指侧方静脉重建回流,术中吻合动静脉比例1:1或1:2或2:2,平均1:2;22例吻合指腹静脉重建回流,术中吻合动静脉比例1:1;20例未吻合静脉,术中仅吻合1条动脉,行侧切口或甲床放血。观察各组治疗效果。结果:吻合指侧方静脉组手指全部成活,无一例发生回流障碍;吻合指腹静脉组19例发生静脉危象,其中4例手指坏死;未吻合静脉组20例均发生回流障碍,其中6例手指坏死。58例获随访,随访时间6~28个月。吻合指侧方静脉组32例,指尖外形佳、指腹饱满;吻合指腹静脉组14例,指体轻度萎缩,指甲生长不平整;未吻合静脉组12例,指体萎缩明显。吻合指侧方静脉组指甲生长近平整,长度长于其他两组[(14.4±3.2)mm比(12.5±2.3)mm和(12.2±2.2)mm],远侧指间关节活动度大于其他两组[(63±5)°比(48±3)°和(45±7)°],两点分辨觉小于其他两组[(4.6±0.4)mm比(7.1±1.2)mm和(7.3±0.6)mm],感觉级别高于其他两组[S(3.45±0.39)级比S(2.57±0.42)级和S(2.55±0.49)级],差异均具有显著性(P〈0.05)。吻合指腹静脉组和未吻合静脉组在指甲长度、运动和感觉方面差异无统计学意义(P〉0.05)。结论:吻合指侧方静脉能有效解决指尖再植静脉回流问题,可避免回流障碍,成活率高,促进指甲生长,可恢复 DIPJ 活动度及感觉。  相似文献   

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目的:明确不同固定器械在胫骨干不同骨折类型固定中的特点,以指导临床应用。方法:68例胫骨干骨折,行加压钢板螺钉、交锁髓内钉、单侧外固定架固定后,作临床疗效分析。结果:加压钢板固定组42例,感染5例,骨不连1例,平均愈合时间3.8个月;交锁髓内钉固定组13例,无感染及骨不连,平均愈合时间5.4个月;单侧外固定架组13例,骨不连1例,踝关节背伸受限3例,平均愈合时间4.5个月。结论:胫骨骨折交锁髓内钉固定并发症少,功能恢复好,适用范围广,但要注意及时进行动力加压。加压钢板及外固定架固定应选择各自的最佳适应证,以达到理想的疗效。  相似文献   

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