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1.
背景:寻找一种同种异体组织保存液,使保存的肌腱具有最大程度的生物活性。目的:使用无水甘油在常温下对兔子肌腱进行保存,观察不同时间点的细胞形态和肌腱活性。方法:使用特殊的方法处理兔肌腱,并常温避光密封保存在无水甘油中,在保存2,4,7,12个月使用苏木精-伊红染色石蜡切片、透射电镜观察肌腱组织结构及细胞形态,使用超氧化物歧化酶酶学实验观察肌腱的生物学活性。结果与结论:2,4,7,12个月苏木精-伊红染色石蜡切片示大部分细胞细胞膜完整,细胞核规则,肌腱组织结构存在。透射电镜结果示细胞呈静止状态,核形态正常。超氧化物歧化酶酶学实验示肌腱具有超氧化物歧化酶酶活性。结果说明使用特殊的方法处理兔肌腱并在无水甘油中常温避光保存12个月,肌腱的组织结构、大部分细胞的细胞形态和肌腱生物活性得以保存。  相似文献   

2.
背景:最大程度保留肌腱的生物活性是进行同种异体肌腱移植的条件。 目的:通过比较选择出最佳的肌腱常温保存方法。 方法:使用无菌操作的方法将兔肌腱随机分为4组:新鲜肌腱对照组、生理盐水组、无水甘油Ⅰ组(肌腱梯度脱水后在无水甘油中保存)、无水甘油Ⅱ组(肌腱直接在无水甘油中保存),分别于保存的第2,4,7个月进行检测。 结果与结论:苏木精-伊红染色显示:无水甘油Ⅰ组肌腱细胞完整率明显高于无水甘油Ⅱ组。电镜观察发现,无水甘油Ⅰ组大部分肌腱细胞形态正常,肌腱组织结构完整;无水甘油Ⅱ组保存4,7个月后细胞核凝固、固缩、崩解状。无水甘油Ⅰ组的超氧化物歧化酶活性也明显高于无水甘油Ⅱ组。说明肌腱经梯度脱水后在无水甘油中常温保存效果更佳。  相似文献   

3.
背景:如何更好地以组织工程学方法修复关节软骨缺损并达到良好的远期疗效目前尚无公识。 目的:创新性地在膝关节腔内培养兔骨髓间充质干细胞复合同种异体脱钙骨的组织工程软骨。  方法:采用全骨髓贴壁筛选法分离培养兔骨髓间充质干细胞,DMEM/F12完全培养基培养,成软骨诱导条件培养基诱导分化。取同种异体兔的髂骨和椎体骨制作成脱钙骨支架,诱导后的骨髓间充质干细胞种植于脱钙骨支架上,培养1 d后将细胞-支架复合物用筋膜包裹置于兔左膝关节腔内培养,单纯脱钙骨支架筋膜包裹置入右膝关节腔。于培养第4,8,12周分别取材,行大体观察并制成石蜡切片,采用苏木精-伊红染色、甲苯胺蓝染色,Ⅱ型胶原免疫组化染色方法进行组织学观察。 结果与结论:培养4,8周,细胞-支架组标本Ⅱ型胶原免疫组化的平均吸光度值(A)分别为0.263±0.031,0.340±0.052,单纯支架组标本分别为0.147±0.027,0.165±0.030,两组比较差异有显著性意义(P < 0.05);培养12周细胞-支架组标本Ⅱ型胶原免疫组化A值平均为0.362±0.037,标本类似正常软骨外观,Ⅱ型胶原免疫组化反应呈阳性;而单纯支架组脱钙骨支架降解。培养12周细胞-支架组苏木精-伊红染色结果显示细胞数量多,脱钙骨支架基本被吸收;而甲苯胺蓝染色结果显示有被染成紫红色的异染性基质形成。结果提示兔骨髓间充质干细胞复合同种异体脱钙骨可在兔膝关节腔内培养出组织工程软骨。  相似文献   

4.
背景:自体肌腱修复肌腱缺损因可用肌腱有限且形成供区功能障碍,同种异体肌腱同样来源有限,并且价格昂贵,在临床上很难满足其需要。目的:观察不同时期异种肌腱修复肌腱缺损的微观变化,为异种肌腱作为临床组织工程化肌腱生长支架提供理论依据。方法:取6月龄的Leghorn鸡屈趾肌腱经化学去细胞处理后作为异种肌腱移植供体,健康成熟日本大耳白兔36只,建立双后肢跟腱中间束2 cm缺损模型,随机分为异种肌腱移植组和自体肌腱移植组,每组18只。肌腱移植缝合用4-0无创伤肌腱缝合线行双"8"字缝合,移植后伸直位管型石膏固定2周,对供体肌腱行去细胞前后大体观察、生物力学测定、组织学光镜及电镜观察,术后2,4,9周每组取6只兔对标本行组织学光镜及电镜检测。结果与结论:1肌腱经过化学去细胞处理后色泽变白,质地较前柔软,去细胞前可见细胞与胶原纤维交替紧密排列,去细胞后胶原排列相对松散,且无细胞及细胞碎片,去细胞后肌腱的力学强度较术前减弱。2由电镜图片直观看到:随移植时间的延长,移植的粗大鸡肌腱胶原纤维逐渐被再生的兔肌腱胶原纤维最终替代,而新生成的纤细胶原纤维经改造塑形变为粗细相等的较粗大纤维,排列方向逐渐趋于平行,在结构和功能上达到正常肌腱水平。结果表明,去细胞后的最大抗拉力是去细胞前的83.44%,能够满足肌腱移植生物力学的要求。最终肌腱修复是再生胶原纤维形成的结果,异种肌腱经过理化方法处理后可作为临床肌腱修复的生长支架使用。  相似文献   

5.
背景:临床发现膝关节内游离体能长期存在于关节腔内并能保持一定的软骨组织学特性和生理学特性,因此大胆提出假设:关节腔环境可能是软骨细胞生长、发育的较佳环境并提出"腔内培养,腔内移植"的理念。目的:观察兔骨髓间充质干细胞复合同种异体脱钙骨基质体外培养或关节腔内培养组织工程软骨与同腔软骨的性状差异。方法:实验分3组进行,体外培养组将经成软骨诱导的乳兔骨髓间充质干细胞与成年兔脱钙骨基质体外复合培养;腔内培养组将经成软骨诱导的乳兔骨髓间充质干细胞与成年兔脱钙骨基质以筋膜包裹,复合培养于成年新西兰兔膝关节腔内,以同腔内正常软骨为对照。结果与结论:培养12周后:①体外培养组苏木精-伊红染色见软骨细胞少量增生,胞核蓝染;甲苯胺蓝染色见软骨细胞排列无序,少量周围基质包绕;Masson染色阳性区域小,细胞排列无序;Ⅱ型胶原免疫组织化学见软骨细胞胞浆及胞外基质少量黄色颗粒。②腔内培养组苏木精-伊红染色见软骨细胞增生,胞核蓝染;甲苯胺蓝染色见软骨细胞成串排列,软骨陷窝形成,周围基质包绕;Masson染色阳性,软骨细胞多,基质蓝染,按一定应力方向排列;Ⅱ型胶原免疫组织化学见细胞外基质中出现较多棕黄色颗粒,Ⅱ型胶原染色阳性。说明骨髓间充质干细胞与同种异体脱钙骨基质复合物可在体外及膝关节腔内培养出组织工程软骨,关节腔内培养的软骨比体外培养的软骨更接近正常软骨。  相似文献   

6.
梁艳清 《解剖学研究》2012,34(2):159-160
<正>让冰冻切片的苏木精-伊红(HE)染色效果等同于石蜡切片,固定液的选择是最关键。目前很多相关文献报道说法不一。我们选用组织学常用和易配制的6种固定液进行苏木精-伊红(HE)染色效果的比较观察。  相似文献   

7.
龚正 《中国组织工程研究》2015,19(21):3329-3333
 背景:对组织器官进行脱细胞,细胞外基质在经过去除细胞和可溶性蛋白处理之后,可维持正常的器官外形及组织结构基质成分。 目的:利用灌注法制备肺脱细胞基质。 方法:将40只Wistar大鼠随机均分为2组,常规麻醉处理后打开胸腔,获得完整的肺组织,实验组利用Langendorff灌注模型构建大鼠全肺脱细胞基质支架,对照组不予以特殊处理。动态观察和记录实验组肺脏颜色和形态变化,分别从2组大鼠肺脏不同部位获取小块组织,利用电子显微镜进行组织学观察,观察两组的Weigert弹力纤维联合Von Gieson结缔组织染色和苏木精-伊红染色情况。 结果与结论:经1%脱氧胆酸钠溶液灌注后,实验组大鼠肺脏颜色逐渐发生改变,表现为从内至外呈分段分叶状逐渐变为白色半透明状,并可观察到清晰肺叶结构,最终肺脏呈现出均一的白色半透明状。经Weigert弹力纤维联合Von Gieson结缔组织染色和苏木精-伊红染色发现,对照组新鲜肺组织切片中含有大量细胞,其中包括毛细血管和成纤维细胞及内皮细胞等,细胞呈整齐排列状,具有完整的肺泡结构,弹性纤维结构清晰,胶原纤维也呈整齐排列状,结构较为紧凑;实验组肺组织细胞基本已消失,仍具有完整的肺泡形态结构,但呈较为疏松的状态且裂隙增大,弹性纤维保存良好,胶原纤维呈较疏松排列状。表明利用灌注法课可有效构建大鼠全肺组织基质支架。中国组织工程研究杂志出版内容重点:生物材料;骨生物材料; 口腔生物材料; 纳米材料; 缓释材料; 材料相容性;组织工程  相似文献   

8.
左健  孙皓  潘乐 《中国组织工程研究》2011,15(53):10039-10042
背景:近年来同种异体肌腱移植逐渐用于治疗膝关节前交叉韧带损伤。 目的:综述膝关节前交叉韧带损伤的特点及其同种异体肌腱移植重建后的免疫排斥反应问题。 方法:应用计算机检索1990-01/2011-10 PubMed数据库及维普数据库相关文献。英文检索词“anterior cruciate ligament,allograft,anatomy,transplantation”,中文检索词“前交叉韧带,同种,肌腱,移植”。检索文献量总计164篇,最终纳入符合标准的文献34篇。 结果与结论:国内外学者对同种异体肌腱移植重建前交叉韧带进行了几十年的研究,已逐步了解了膝关节前交叉韧带的生物力学特性;通过冷冻处理法、细胞毒物质处理法解决了同种异体肌腱移植后的免疫排斥反应,使其既能降低异体肌腱的抗原性,又能保存肌腱细胞的活性,促使肌腱的内源性愈合,提高肌腱愈合速度与强度,减轻或避免肌腱粘连的发生,临床应用取得了很好的效果。  相似文献   

9.
组织学考试用组织芯片制作初探   总被引:2,自引:0,他引:2  
目的实现组织芯片在组织学教学中的应用。方法取正常Wistar大鼠心、肝、肾、脾等器官,采用组织芯片仪制作组织微阵列蜡块,常规切片,经苏木精—伊红染色制成组织学考试用组织芯片。结果芯片中组织微阵列排列整齐,各样本组织染色清晰,组织定位良好,可判断性强。结论与传统组织切片相比,组织学考试用组织芯片具有低消耗、可比性强、简便易行、局限性小的特点,应用前景广阔。  相似文献   

10.
背景:理想的脱细胞方法要求既能完全去除供体细胞,降低免疫原性,又能保留天然瓣膜的胶原纤维、弹力纤维等细胞外基质成分,以保持足够的机械强度。 目的:采用不同洗剂制备脱细胞猪主动脉瓣膜支架,对比其组织结构,探讨最为有效的脱细胞瓣膜支架制备方法。 方法:20个新鲜猪主动脉瓣膜随机分为新鲜对照组、去污剂组、酶消化组和去污剂-酶消化组,后3组分别使用Triton X-100、胰蛋白酶以及二者联合的方法制备脱细胞瓣膜支架,对比支架大体形态、苏木精-伊红染色、Mallory-Heidenhain染色和电镜下超微结构的不同。 结果与结论:经脱细胞处理后,去污剂组瓣叶柔软、光滑,苏木精-伊红染色少量核物质存留、纤维排列规整,Mallory-Heidenhain染色胶原纤维和弹性纤维相交错,电镜下呈波浪状排列、原纤维横纹清楚;酶消化组瓣叶局部塌陷,苏木精伊红染色细胞完全去除、纤维排列较紊乱,Mallory-Heidenhain染色胶原纤维和弹性纤维呈网状排列,电镜下纤维部分断裂、原纤维横纹存在;去污剂-酶消化组瓣叶柔软、光滑,苏木精伊红染色细胞完全去除、纤维完整,Mallory-Heidenhain染色胶原纤维和弹性纤维平行排列,电镜下纤维完好,但排列稀疏,原纤维横纹清晰。说明3种方法均可有效去除供体细胞,保持纤维结构相对完整,在完全清除供体细胞并保持纤维支架完整性方面,Triton X-100联合胰蛋白酶的方法更为有效。  相似文献   

11.
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。  相似文献   

12.
Collagenase was injected into the Achilles tendon of both hind legs of 10 clinically normal adult male New Zealand white rabbits. One month after induction of the injury, beta-aminoproprionitrile (BAPN) or hyaluronic acid (HA) was injected into the tendon core of the right hind leg of each rabbit, the left hind leg being left untreated. The treatment effects were evaluated by electron microscopy and analysis of the glycosaminoglycan (GAG) content of samples at 2 and 6 months post-treatment. At 2 months, collagen fibrils in tendons from both hind legs were relatively small in diameter, irregularly arranged, and interspersed with abundant active tenocytes as compared with those in normal tendon uninjured by collagenase. In the matrix, the amount of HA increased, but chondroitin-6-sulphate was eliminated. At 6 months, BAPN-treated tendons had small-diameter, regularly arranged collagen fibrils. HA-treated tendons, on the other hand, had large diameters, as well as regularly arranged collagen fibrils by comparison with non-treated tendon. The results suggest that HA, unlike BAPN, promoted healing.  相似文献   

13.
背景:异体肌腱移植是目前修复肌腱缺损的理想方法,但移植后的排斥反应是其使用受到限制的主要原因。 目的:观察脱细胞处理的版纳近交系微型猪肌腱移植修复兔跟腱缺损的疗效,以及其作为异种肌腱移植支架材料的可行性。 方法:将40只日本大白兔制作双后肢跟腱缺损实验动物模型后,随机均分为2组,脱细胞猪肌腱组用脱细胞版纳近交系微型猪肌腱修复,自体肌腱组用自体肌腱修复,术后用3-0肌腱线改良HEMI-KESSLER法进行端端原位吻合。 结果与结论:①移植后2周内,脱细胞猪肌腱组与自体肌腱组白细胞计数、C-反应蛋白测量结果差异无显著性意义(P > 0.05)。②两组移植后局部反应小,伤口一期愈合,屈踝功能恢复正常。③组织学检查均未见明显淋巴细胞浸润,肌腱缝合处胶原纤维相互衔接。结果说明脱细胞版纳近交系微型猪肌腱能成功修复兔跟腱缺损,且具有组织相容性好、移植排斥反应轻的优点。  相似文献   

14.
背景:异体移植物可作为前交叉韧带重建翻修以及膝关节复合损伤修复的选择。 目的:通过建立异体韧带移植重建前交叉韧带的动物模型,观察早期活动对异体植入物止点腱骨愈合的组织形态以及关节活动功能恢复的影响。 方法:健康成年新西兰兔9只,随机取3只兔双侧跟腱作为供体,取6只兔切断一侧膝关节前交叉韧带,固定重建前交叉韧带。动物随机数字表法均分运动组和制动组,6周后观察关节活动功能、腱骨愈合大体观察以及组织学观察。 结果与结论:术侧肢体活动情况基本正常,所有动物前交叉韧带上下止点愈合情况均良好。前交叉韧带周围滑膜均可见明显增生。活动组可见较多Sharpey纤维,腱骨间接连接形成,而制动组未见明显Sharpey纤维。说明术后6周异体重建物止点已有腱骨愈合,早期活动对重建物的腱骨愈合并无明显不良影响,可能还更有利。  相似文献   

15.
This study explored the hypothesis that local administration of a polysulphated glycosaminoglycan (PSGAG) in the early phase of healing of a standard collagenase-induced tendon injury in the superficial digital flexor tendon of the rabbit would reduce the degenerative effects of inflammatory mediators and proteases and preserve normal tendon morphology, composition, and biomechanical properties. Histological and ultrastructural changes together with the mechanical properties, dry weight, collagen content, and amount of DNA in healing tissue at the site of the lesion were assessed in treated and untreated animals. In treated lesions 28 days after injury, the normal orientation of tenoblasts and collagen fibrils was well preserved compared with the disorganized scar formation seen in untreated animals. The degree of cellularity was significantly higher in the untreated lesions. At the ultrastructural level the collagen in the healing tissue of the treated animals consisted of a mixture of small diameter, new regenerated fibrils intermingled with well-preserved large diameter, old fibrils, aligned to the long axis of the tendon; in untreated animals small, randomly arranged new fibrils predominated. The diameters of treated tendons had returned to normal, but in untreated animals the injured tendons remained significantly thicker than their controls. The percentage dry weight and collagen contents of treated injured tendons approximated those of control normal tendons, whereas those of untreated tendons were significantly less than those of the control values. The DNA content of injured treated tendons was not significantly different from that of normal contralateral controls, while in the untreated tendons it was significantly higher. There were no significant differences between the normal and the contralateral treated injured tendons in ultimate strength, fatigue strength, stiffness, and maximum absorbed energy. However in the untreated animals, although the tendon diameter was significantly greater, the ultimate strength, fatigue strength, stiffness, and maximum absorbed energy were significantly lower than the contralateral control. These data suggest that polysulphated glycosaminoglycans are effective in restoring the morphological, biochemical, and biomechanical properties of injured soft connective tissues and may be of clinical value in the treatment of acute tendon injury.  相似文献   

16.
This study explored the hypothesis that local administration of a polysulphated glycosaminoglycan (PSGAG) in the early phase of healing of a standard collagenase-induced tendon injury in the superficial digital flexor tendon of the rabbit would reduce the degenerative effects of inflammatory mediators and proteases and preserve normal tendon morphology, composition, and biomechanical properties. Histological and ultrastructural changes together with the mechanical properties, dry weight, collagen content, and amount of DNA in healing tissue at the site of the lesion were assessed in treated and untreated animals. In treated lesions 28 days after injury, the normal orientation of tenoblasts and collagen fibrils was well preserved compared with the disorganized scar formation seen in untreated animals. The degree of cellularity was significantly higher in the untreated lesions. At the ultrastructural level the collagen in the healing tissue of the treated animals consisted of a mixture of small diameter, new regenerated fibrils intermingled with well-preserved large diameter, old fibrils, aligned to the long axis of the tendon; in untreated animals small, randomly arranged new fibrils predominated. The diameters of treated tendons had returned to normal, but in untreated animals the injured tendons remained significantly thicker than their controls. The percentage dry weight and collagen contents of treated injured tendons approximated those of control normal tendons, whereas those of untreated tendons were significantly less than those of the control values. The DNA content of injured treated tendons was not significantly different from that of normal contralateral controls, while in the untreated tendons it was significantly higher. There were no significant differences between the normal and the contralateral treated injured tendons in ultimate strength, fatigue strength, stiffness, and maximum absorbed energy. However in the untreated animals, although the tendon diameter was significantly greater, the ultimate strength, fatigue strength, stiffness, and maximum absorbed energy were significantly lower than the contralateral control. These data suggest that polysulphated glycosaminoglycans are effective in restoring the morphological, biochemical, and biomechanical properties of injured soft connective tissues and may be of clinical value in the treatment of acute tendon injury.  相似文献   

17.
背景:有实验证实外源性6-磷酸果糖能降低肌腱细胞Ⅰ型胶原的产生量,减轻肌腱术后粘连的形成。 目的:观察6-磷酸果糖对肌腱术后粘连形成的影响。 方法:取72只兔行中趾屈指肌腱切断吻合,随机分成实验组与对照组(n=36),分别于腱鞘内注入6-磷酸果糖与生理盐水,术后4,8周后行肌腱粘连检测、生物力学测定、组织学观察和扫描电镜观察;术后1,2,4,8周采用原位杂交方法测定肌腱转化生长因子β1和Ⅰ型胶原mRNA的表达。 结果与结论:术后4,8周,与对照组比较,实验组肌腱缝合处光滑,屈趾肌腱滑动距离较长,肌腱滑动受限较轻(P < 0.05),但两组最大抗断裂载荷差异无显著性意义(P > 0.05);扫描电镜和组织学观察结果显示,对照组胶原纤维排列紊乱,实验组胶原纤维排列整齐。实验组转化生长因子β1和Ⅰ型胶原mRNA表达水平均低于对照组(P < 0.05)。证实6-磷酸果糖能有效抑制转化生长因子β1在肌腱损伤修复中的作用,减轻粘连形成。  相似文献   

18.
目的:探究随机肌腱细胞外基质(ECM)支架对骨髓间充质干细胞(BMSCs)活力和分化的影响。方法:从Sprague-Dawley大鼠股骨和胫骨中提取BMSCs,体外培养,观察细胞形态,并利用流式细胞术鉴定细胞干性。采用1%Triton X-100和DNase/RNase混合液对鼠尾肌腱进行脱细胞处理,利用HE染色和DNA含量测定考察肌腱组织中细胞核残余情况。制备胶原纤维随机排列的肌腱ECM支架,培养BMSCs,以孔板中生长的细胞为对照组,利用Live/Dead染色和CCK8法考察细胞的活力和形态;利用RT-qPCR检测肌腱标志物I型胶原蛋白(Col I)、肌腱特异转录因子scleraxis(SCX)及成骨标志物碱性磷酸酶(ALP)和Runt相关转录因子2(RUNX2)的表达水平。结果:HE染色结果显示,经过脱细胞处理后肌腱组织内无细胞残余,且DNA含量从(481.7±15.8)μg/g显著性降至(31.0±3.8)μg/g(P<0.05),脱细胞处理成功。7 d时,种植在支架上的BMSCs的活力较对照组显著增强(P<0.05);14 d时,种植在支架上的BMSCs肌腱标志物Col I和SCX的表达量较对照组显著下调,而成骨标志物ALP和RUNX2的表达量较对照组显著上调(P<0.05)。结论:脱细胞随机肌腱ECM支架能增强BMSCs活力,并诱导其向成骨细胞分化。  相似文献   

19.
Regenerated tendon tissue replacing partial-thickness defects produced in rabbit calcaneal tendons was seen to undergo almost complete maturation between three and sixteen weeks after surgery. At this stage, most collagen fibers had large diameters as in normal tendon, and the majority of elastic fibers were mature. This findings, along with those of a previous study on completely severed and unsutured tendons, indicate that mechanical stress plays a primary role in maturation of regenerating tendon. This emphasizes the importance of close approximation of tendon ends in the management of tendon ruptures and lacerations. This is possibly by surgical repair.  相似文献   

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