不同时段超低温保存对兔肌腱力学性能的影响

目的 :探讨不同时段超低温保存对兔肌腱组织生物力学性能的影响。方法 :以 15 %DMSO(二甲基亚砜 )为低温保护剂 ,采用两步冷冻保存步骤 ,对青紫兰兔肌腱进行维持温度 - 5 0℃的冷冻预处理 ,- 196℃液氮保存 18d、180d ,测试前用 4 0℃水浴复温 ,每组肌腱 8段 ,与新鲜肌腱对照进行拉伸实验。结果 :肌腱的破坏荷载在对照组、18d组、180d组分别为 (6 6 .0± 19.4 9)N、(6 5 .0± 7.0 7)N和 (5 4 .0± 11.4 )N ,各组肌腱的延伸率 (% )依次为 6 3.5± 17.3、38.6± 0 .98和 30 .6± 0 .4 7。结论 :经不同时段超低温冷冻保存的兔肌腱 ,拉伸实验的生物力学指标无明显差异 (P>0 .0 5 )。     

连续腱周缝合对肌腱修复的生物力学影响

目的探讨连续腱周缝合（Running）法在肌腱修复中的生物力学作用,为临床提供理论基础和参考依据。方法成年AA白羽鸡爪50只．按缝合方法随机分为5组。锐刀横断Ⅱ区趾深屈肌肌腱,分别用Running法、单线改良Kessler（MK）法、单线改良Kessler＋Running（K＋R）法、津下双套圈（Tsuge）法、Tsuge＋Running（T＋R）法修复肌腱。缝合后立刻取下肌腱,用冰冻卡具固定两端．在生物力学材料动态力学性能测试仪上进行拉伸——断裂测试。测定极限载荷、应变．记录间隙形成负荷,计算出各组肌腱的韧度、极限拉伸强度、弹性模量和断裂功耗并进行统计学分析。结果在间隙形成负荷、韧度、弹性模量、极限载荷、极限拉伸强度和断裂功耗方面,K＋R法均大于MK法（P〈0．001）,后者分别是前者的58．6％、49．9％、58.4％、56．1％、65．7％、66．3％。T＋R法均大于Tsuge法（P〈0．05）,后者分别是前者的57．8％、52．7％、84．0％、64．7％、91．0％、69．6％。结论Running法操作简单．能为核心缝合增加可观的抗拉强度和抗间隙形成能力,使吻合口对合良好、缝合外观光滑,从而减少肌腱滑动阻力．为术后早期功能锻炼提供有效保证。     

玻璃化冷冻保存组织工程肌腱植入大鼠体内的组织学变化

背景：应用玻璃化冷冻方法保存组织工程肌腱具有可行性和应用前景,有待进一步研究。 目的：探讨应用玻璃化冷冻保存组织工程肌腱体内植入对大鼠跟腱缺损修复的影响。 方法：选用成年SD大鼠64只,于大鼠跟腱中段制备5 mm肌腱缺损模型,随机摸球法均分为2组,分别植入玻璃化冷冻保存组织工程肌腱和新鲜非冷冻保存组织工程肌腱。植入后2,4,6,8周,观察植入肌腱材料及周围组织的大体形态和组织学变化。 结果与结论：两组肌腱材料体内植入后的大体形态和组织学反应无明显差异。植入后2,4周,植入的肌腱材料发生降解,材料中间及周围有大量炎性细胞浸润和纤维结缔组织增生。植入后6,8周,大量新生胶原纤维组织长入并替代降解的植入材料,形态和排列方式趋近于正常肌腱组织,大鼠跟腱缺损基本修复。结果表明玻璃化冷冻保存组织工程肌腱体内植入可修复大鼠跟腱缺损。     

组织工程化肌腱种子细胞深低温保存的实验研究

对肌腱种子细胞进行深低温保存，研究保存过程中多个环节的影响因索对细胞存活率的影响及深低温保存对种子细胞生物学特性、胶原分泌功能的影响。实验结果表明二甲基亚砜是肌腱种子细胞深低温保存中比较好的抗冻保护剂；冻存后使用与培养时不同的营养血清处理对细胞有损害，可降低细胞存活率；在冷冻保存过程中，细胞存活率与细胞浓度有一定关系，浓度太低可能降低细胞存活率；细胞在冷冻保存时，降温速度对细胞存活率有影响，慢速的分步降温组细胞存活率明显高于直接人液氮的快速降温组；采用10％二甲基亚砜加15%小牛血清加75% DMEM配方保存肌腱种子细胞，对其分泌胶原功能无明显影响，对其生长曲线、细胞周期及染色体众数无明显改变，适于肌腱种子细胞的保存。     

玻璃化冷冻法保存兔腰椎间盘的实验研究

目的观察玻璃化冷冻法保存兔腰椎间盘的效果。方法选取30只新西兰大耳白兔,随机分为玻璃化冷冻组、常规低温冷冻组和新鲜髓核细胞组,每组10只,将取下的腰椎间盘标本通过玻璃化冷冻法与常规低温冷冻法分别冻存30 d,复温洗脱后,利用MTT比色法测量两组腰椎间盘纤维环及髓核细胞的相对活性,利用台盼蓝拒染法检测其细胞存活率。结果 MTT比色法和台盼蓝拒染法所得结果基本一致,玻璃化冷冻组和常规低温冷冻组兔腰椎间盘髓核细胞存活率和相对活性均低于新鲜髓核细胞组,玻璃化冷冻组的兔腰椎间盘髓核细胞存活率和相对活性均优于常规低温冷冻组,其差异具有统计学意义(P0.05)。结论玻璃化冷冻法保存兔腰椎间盘髓核细胞存活率及细胞活性高,保存效果优于常规低温冷冻法。     

胎兔周围神经低温保存温度值的实验研究

目的 :探讨胚胎兔周围神经低温冷藏后的改变。方法 :以 10 %DMSO(二甲基亚砜 )为低温保护剂 ,采用两步冷冻保存步骤 ,选择 4种不同的冷冻维持温度 (- 2 0℃、- 4 0℃、- 6 0℃、- 80℃ ) ,对孕龄 2 6d～ 2 8d的胎兔坐骨神经进行冷冻 ,维持 30min后 ,放入液氮中保存一周。复温后行四唑盐还原试验、组织学和超微结构观察 ,探讨冷冻维持温度对胎兔周围神经的影响。结果 :- 4 0℃维持温度冷冻后液氮保存的胎兔周围神经代谢活性和超微结构得到良好维持。结论 :两步冷冻法可以较好地保存胎兔周围神经的生物活性 ,为胎周围神经的冷冻保存和移植奠定了基础     

玻璃化法保存同种异体肌腱移植材料的可行性

背景：处理同种异体肌腱最主要目的是减少免疫原性和保持肌腱结构与活性。现在临床上常用的低温冷冻法操作复杂、费时,所保存的肌腱活性较低。 目的：以玻璃化法保存鸡屈趾深肌腱,探索其作为肌腱移植材料的可行性。 方法：将成年来亨鸡跖趾关节远侧屈趾深肌腱切断,以数字表法随机分为2组,分别移植同种异体玻璃化肌腱与自体肌腱。术后2,4,8,12,16周观察移植肌腱在形态结构、羟脯氨酸含量和力学性能等方面的变化。 结果与结论：玻璃化肌腱移植组早期存在轻度的免疫排斥反应,但不影响肌腱愈合与重建。两组肌腱周围产生中度粘连,移植后肌腱中段的羟脯氨酸含量先减少,8周后逐渐增高,而吻合口部羟脯氨酸含量随时间逐渐增高,两组间羟脯氨酸含量差异无显著性意义。在8周内玻璃化肌腱移植组吻合口部的破裂强度与弹性模量低于自体肌腱移植组(P < 0.05),12周后差别无显著性意义。两组肌腱中段部生物力学性能差异无显著性意义。说明玻璃化保存的同种异体肌腱生物活性好、免疫原性低,是良好的肌腱移植材料。     

玻璃化冷冻保存组织工程肌腱大鼠体内植入对血液免疫及生化指标的影响

目的研究组织工程肌腱体内植入修复跟腱缺损的同时,根据细胞与支架材料的选择、工程化组织构建与保存方式所引起的体内毒性反应特征,从血液免疫和生化学指标方面评价玻璃化冷冻保存组织工程肌腱材料对机体的安全性影响。方法随机将72只SD大鼠分为3组,A、B两组各32只,C组8只。手术造成A、B两组大鼠一侧跟腱0.5cm缺失,分别植入1cm长的玻璃化冷冻保存与非玻璃化冷冻保存组织工程肌腱材料,C组不经任何手术。三组大鼠分批于术后第2、4、6、8周股动脉放血处死,收集全血分离血清,采用双抗体夹心ELISA、速率、双缩脲、溴甲酚绿、尿酶紫外速率、苦味酸、已糖激酶、氧化酶、DMSO、Diaso法测定血液相关免疫及生化指标。结果 A、B两组各项指标在相同植入期比较,差异无统计学意义（P〉0.05）;A、B两组与C组比较,除A组4周、B组4周和6周AST值及A组2周ALB值、B组2周CREA值明显低于C组相应指标外（P〈0.05）,其他指标差异均无统计学意义（P〉0.05）。结论玻璃化冷冻保存组织工程肌腱大鼠体内植入后血液免疫及生化各项指标与非手术大鼠基本一致。证实了玻璃化冷冻保存组织工程肌腱用于肌腱缺损修复是安全可靠的,未发现免疫排斥反应,对肝肾功能无明显影响。     

不同低温保护剂对皮肤组织β1整合素表达影响的比较

目的比较两种不同低温保护剂对皮肤组织β1 integrin低温保存后表达的影响，为寻求皮肤组织低温保护剂的最佳配方提供试验依据。方法获取新鲜成人皮肤组织分为3组．新鲜对照组、海藻糖／二甲基亚砜（T／D）组、二甲基亚砜／丙二醇（D／P）作为低温保护剂保存组，-196℃液氮冻存7d、14d复温，免疫组织化学染色对各组间皮肤进行比较。并在此基础上，进一步采用RT-PCR方法对不同低温保护剂保存后皮肤的β1 integrin基因水平进行了深入的研究。结果通过光镜图象观察和基因水平分析结果说明，0．5M海藻糖／二甲基亚砜能够很好保护皮肤组织，β1 integrin的基因表达量与新鲜皮肤组相似。结论海藻糖与二甲基亚砜联合应用对皮肤组织β1 integrin的保护作用优于传统组。     

玻璃化冷冻法保存关节软骨

背景：同种异体骨软骨移植技术是治疗关节软骨缺损有效的方法之一,但由于移植物保存方法不理想,明显制约着该技术的临床应用。 目的：探讨玻璃化冷冻法保存关节软骨组织的可行性和优越性。 方法：切取成年猪骨软骨,制成约5 mm×6 mm(直径×长度)大小的圆柱形骨软骨块。以新鲜软骨组为对照,分别采用0.5 mol/L甘油 、1 mol/L二甲基亚砜、1 mol/L玻璃化溶液3种方法预处理软骨块,再行冷冻法保存软骨块8周,采用组织化学染色、免疫荧光染色观察并比较软骨细胞活性的变化。 结果与结论：玻璃化溶液预处理组的关节软骨细胞存活率达到74.5%,明显高于甘油和二甲基亚砜预处理组,软骨基质成分仅少量丢失。3种方法相比较,玻璃化溶液预处理后慢速梯度降温冷冻保存法可以明显提高冻存关节软骨组织的活性。     

实时剪切波弹性成像技术对成年人肝脏杨氏模量正常值的研究

目的探讨应用实时剪切波弹性成像技术评价正常人肝脏杨氏模量值的临床应用价值。方法553例健康志愿者,其中男性213例,女性340例;年龄2l～80岁,平均年龄50．4岁。按照年龄分为3组：青年组（20．39岁）183例,中年组（40～59岁）250例．老年组（60～80岁）120例。应用法国声威公司AixPlorer超声诊断仪,所有研究对象均首先进行常规超声检查,然后再进行肝脏超声剪切波弹性成像检查,并行肝脏杨氏模量值定量分析,于深度4cm处测量感兴趣区域（ROI）（直径：lcm）内测量肝组织杨氏模量值。结果553例研究对象总的肝脏杨氏模量值为（6．10±1．31）kPa;青年组、中年组、老年组分别为（5．72±1．25）kPa、（6．21±1．63）kPa、（6．26±1．74）kPa,青年组低于中年组、老年组（P〈0．05）;中年组与老年组及不同性别间比较差异无统计学意义（P〉0．05）。结论应用实时剪切波弹性成像技术可定量检测正常人肝组织的杨氏模量值．杨氏模量值与年龄有关．但与性别无关。应用实时剪切波弹性成像技术评价肝脏杨氏模量时应考虑年龄因素。     

Preface

The aim of this study was to evaluate if spontaneous (nonforced active) exercise and age (maturation process) alter the biomechanical and biochemical properties of superficial digital flexor tendon. Chickens aged 1, 5, and 8 months were divided into two groups: caged and penned. The caged group was reared in an area of 0.5 m² (3 animals/cage), while the penned group was reared in an area of 60 m² (3 animals/area). For biochemical analysis, the tendon was divided into tensile and compressive regions for quantification of hydroxyproline and glycosaminoglycan content. Biomechanical properties were analyzed from tensile tests of intact tendons. The biomechanical measurements were taken at maximum load and maximum stress. In both the caged and penned groups, maximum load and energy absorption increased with maturation; however, the elastic modulus, maximum stress, and maximum strain did not increase with maturation. Exercise resulted in a higher load, stress, and elastic modulus in the fifth month. Collagen content increased with age in the penned group and with exercise in the fifth and eighth months. Exercise results in a higher expression of glycosaminoglycans in young tendons compared to mature tendons. Thus, low-intensity mechanical stimuli promote the synthesis and possible rearrangement of molecules in immature tendons, whereas inactivity leads to deleterious effects on the material properties (maximum stress and elastic modulus) during growth and maturation.     

A Poly(lactic-co-glycolic acid) Knitted Scaffold for Tendon Tissue Engineering: An In Vitro and In Vivo Study

We have designed a composite scaffold for potential use in tendon or ligament tissue engineering. The composite scaffold was made of a cellularized alginate gel that encapsulated a knitted structure. Our hypothesis was that the alginate would act as a cell carrier and deliver cells to the injury site while the knitted structure would provide mechanical strength to the composite construct. The mechanical behaviour and the degradation profile of the poly(lactic-co-glycolic acid) knitted scaffolds were evaluated. We found that our scaffolds had an elastic modulus of 750 MPa and that they lost their physical integrity within 7 weeks of in vitro incubation. Autologous rabbit mesenchymal stem cell seeded composite scaffolds were implanted in a 1-cm-long defect created in the rabbit tendon, and the biomechanical properties and the morphology of the regenerated tissues were evaluated after 13 weeks. The regenerated tendons presented higher normalized elastic modulus of (60%) when compared with naturally healed tendons (40%). The histological study showed a higher cell density and vascularization in the regenerated tendons.     

利用罗曼鸡肌腱滑液鞘体内构建趾深屈肌腱的生物力学及组织学研究

目的 探索肌腱滑液鞘对肌腱体内再生的作用。 方法 将36只罗曼鸡随机平均分为A、B两组。A组把左趾深屈肌腱滑液鞘的上、下、右侧分离,不切断趾深屈肌腱。同种异体脱细胞肌腱用部分分离的腱滑液鞘包裹,并固定在趾深屈肌腱左侧。B组直接把同种异体脱细胞肌腱固定在去除肌腱滑液鞘的趾深屈肌腱左侧。另取右趾深屈肌腱作为正常对照组。分别在第4、8、12周取材进行趾深屈肌腱最大载荷、弹性模量的生物力学检测及HE染色的组织学检测。结果 术后第4、8、12周时,除第4周A、B组在弹性模量方面差异无统计学意义,其余时间点A组最大载荷和弹性模量都大于B组（P<0.05）。随时间延长,A组胶原纤维增多,排列紧密,方向一致,炎性细胞浸润及纤维组织增生程度较轻。而B组胶原纤维数量逐渐减少、排列逐渐散乱,炎性细胞浸润、纤维组织增生程度明显高于A组。结论腱滑液鞘有助于肌腱再生,说明合适的体内环境对肌腱构建具有重要作用,本研究结果对临床上肌腱缺损病人找到合适肌腱替代物有积极作用。     

Knitted poly-lactide-co-glycolide scaffold loaded with bone marrow stromal cells in repair and regeneration of rabbit Achilles tendon

The objectives of this study were to evaluate the morphology and biomechanical function of Achilles tendons regenerated using knitted poly-lactide-co-glycolide (PLGA) loaded with bone marrow stromal cells (bMSCs). The animal model used was that of an adult female New Zealand White rabbit with a 10-mm gap defect of the Achilles tendon. In group I, 19 hind legs with the created defects were treated with allogeneic bMSCs seeded on knitted PLGA scaffold. In group II, the Achilles tendon defects in 19 hind legs were repaired using the knitted PLGA scaffold alone, and in group III, 6 hind legs were used as normal control. The tendon-implant constructs of groups I and II were evaluated postoperatively at 2, 4, 8, and 12 weeks using macroscopic, histological, and immunohistochemical techniques. In addition, specimens from group I (n = 7), group II (n = 7), and group III (n = 6) were harvested for biomechanical test 12 weeks after surgery. Postoperatively, at 2 and 4 weeks, the histology of group I specimens exhibited a higher rate of tissue formation and remodeling as compared with group II, whereas at 8 and 12 weeks postoperation, the histology of both group I and group II was similar to that of native tendon tissue. The wound sites of group I healed well and there was no apparent lymphocyte infiltration. Immunohistochemical analysis showed that the regenerated tendons were composed of collagen types I and type III fibers. The tensile stiffness and modulus of group I were 87 and 62.6% of normal tendon, respectively, whereas those of group II were about 56.4 and 52.9% of normal tendon, respectively. These results suggest that the knitted PLGA biodegradable scaffold loaded with allogeneic bone marrow stromal cells has the potential to regenerate and repair gap defect of Achilles tendon and to effectively restore structure and function.     

多代连续饮用四种饮水的雌性大鼠骨生物力学性能和骨代谢水平的比较

目的观察雌性SD大鼠多代连续单纯饮用自来水、纯净水、矿物质水和天然水,对其骨质密度、骨生物力学性能及骨代谢的影响。方法取前期实验暴露于四种饮水条件下的F2代断乳健康雌性SD大鼠,各组30只,饮用与F1代相同饮水,除饮水外其他条件完全相同。饲养10个月后处死,腹主动脉取血并分离血清检测大鼠骨碱性磷酸酶（BALP）、骨钙素（BGP）、血清I型前胶原羧基端前肽（PICP）、I型胶原交联羧基末端肽（ICTP）,同时取右侧股骨,进行骨密度和骨生物力学检测。结果血清中BGP含量天然水组高于自来水组（P〈0.05）;PICP含量纯净水组和天然水组低于自来水组（P〈0.01）;ICTP含量天然水组低于自来水组（P〈0.01）。拉伸试验显示：自来水组大鼠股骨最大桡度高于天然水组、矿物质水组和纯净水组（P〈0.01）;自来水组弹性桡度高于天然水组（P〈0.01）和矿物质水组（P〈0.05）。自来水组大鼠股骨最大应变高于天然水组、矿物质水组和纯净水组（P〈0.01）;断裂应变自来水组高于矿物质水组和天然水组（P〈0.01）。杨氏模量自来水组低于天然水组（P〈0.05）。结论长期饮用不同水质的水,可能对大鼠骨骼的生长发育会造成一定的影响。     

Effects of mechanical stimulation on the biomechanics and histology of stem cell-collagen sponge constructs for rabbit patellar tendon repair

The objective of this study was to determine how mechanical stimulation affects the biomechanics and histology of stem cell-collagen sponge constructs used to repair central rabbit patellar tendon defects. Autogenous tissue-engineered constructs were created for both in vitro and in vivo analyses by seeding mesenchymal stem cells from 10 adult rabbits at 0.14x10(6) cells/construct in type I collagen sponges. Half of these constructs were mechanically stimulated once every 5 min for 8 h/day to a peak strain of 4% for 2 weeks. The other half remained in an incubator without mechanical stimulation for 2 weeks. Samples allocated for in vitro testing revealed that mechanically stimulated constructs had 2.5 times the linear stiffness of nonstimulated constructs. The remaining paired constructs for in vivo studies were implanted in bilateral full-thickness, full-length defects in the central third of rabbit patellar tendons. Twelve weeks after surgery, repair tissues were assigned for biomechanical (7 pairs) and histologic (3 pairs) analyses. Maximum force, linear stiffness, maximum stress, and linear modulus for the stimulated (vs. nonstimulated) repairs averaged 70% (vs. 55%), 85% (vs. 55%), 70% (vs. 50%), and 50% (vs. 40%) of corresponding values for the normal central third of the patellar tendons. The average force-elongation curve for the mechanically stimulated repairs also matched the corresponding curve for the normal patellar tendons, up to 150% of the peak in vivo force values recorded in a previous study. Construct and repair linear stiffness and linear modulus were also positively correlated (r = 0.6 and 0.7, respectively). Histologically both repairs showed excellent cellular alignment and mild staining for decorin and collagen type V, and moderate staining for fibronectin and collagen type III. This study shows that mechanical stimulation of stem cell-collagen sponge constructs can significantly improve tendon repair biomechanics up to and well beyond the functional limits of in vivo loading.     

Canine tendon studies. II. Biomechanical evaluation of normal and regrown canine tendons.

Some of the mechanical properties of regrown canine tendons are compared to those of normal tendons of young and mature animals. Patellar and Achilles tendons from 12 beagle dogs were removed and studied with their bone origin and insertions. Mechanical tests were performed within 24 hr and test conditions simulated the physiological function of the tendon in vivo at room temperature. Specimens were soaked in Ringers solution and mounted in an Instron testing machine with load deflection curves plotted automatically. The parameters used for analysis were load extension, stress relaxation, elastic limit, and strain rate dependence. The regrown tendons in young animals appeared to quickly adjust in dimension and structure so that their properties were not significantly different from those of normal tendons on a load extension basis. The normal tendons were stiffer than regrown ones but the modulus of elasticity increased with age. The Achilles were stiffer than patellar tendons. Cyclic loading with 25 kg did not affect reconstructed tendon models, although some increase in stiffness was noted. The elastic modulus decreased with an increase in ambient temperature and increasing strain rate.     

几种屈肌腱修复方法的生物力学研究

目的：分析和比较４种肌腱合方法的生物力学特性，为临床提供实验基础。方法：将新鲜成年猪后足２０只，随机分在４组，解剖中间二趾深屈肌腱后足第１组行改良Ｋｅａｓｌｅｒ法缝合，第２组行真皮嵌入法缝 ，第３组行Ｂｅｃｋｅｒ法缝合，第４组行Ｔａｎｇ法缝合。测定每组缝合方法的间隙形成负荷、２ｍｍ间隙负荷、最大负荷、最大功耗及弹性模量，并行统计学分析（ＡＮＯＶＡ）。结果：Ｔａｎｇ法的２ｍｍ间隙形成负荷显著高于其他