核心蛋白聚糖对兔肌腱细胞增殖及细胞周期的影响

目的 观察核心蛋白聚糖(decorin,DCN)对兔肌腱细胞体外增殖和细胞周期的影响,以探讨decorin在肌腱愈合中对肌腱细胞的作用.方法 原代培养兔肌腱细胞,分别加入不同浓度(0.25、1.25、2.5、5μg/ml)的decorin;培养12、24、48 h用二甲基噻唑二苯基四唑溴盐(MTT)法测定细胞增殖速度;镜下观察低浓度DCN作用24 h后对兔肌腱细胞增殖的影响;低浓度DCN培养24 h后用流式细胞仪检测细胞周期.结果 0.25、1.25、2.5μg/ml浓度decorin作用12 h后对兔肌腱细胞增殖有着减少的趋势,但是24 h后却明显地促进其增殖(P＜0.05),但是48 h后差异不显著.而5μg/ml浓度组作品12 h对兔肌腱细胞增殖有着增加的趋势,24 h后促进增殖作用显著(P＜0.05),48 h后却抑制兔肌腱细胞的增殖.0.25μg/ml低浓度decorin作用兔肌腱细胞24h后镜下观察和流式细胞分析,细胞增殖明显增强,S期增加,PI明显大于对照组(P＜0.05).结论 低、中浓度decorin对兔肌腱细胞的增殖起着先延迟后促进的作用,提示在肌腱愈合中如果在肌腱和腱鞘之间运用DCN,可能起到早期隔离TGF-β对腱鞘成纤维细胞的作用,而后期增强TGF-β对腱内膜细胞的作用,从而促进内源性愈合,减少外源性愈合,达到减少粘连的效果.     

结缔组织生长因子及抗体对体外培养鸡滑膜细胞增殖活性的影响

目的：研究不同浓度结缔组织生长因子（CTGF）及其抗体对鸡肌腱滑膜细胞增殖活性的影响.方法：以来亨鸡屈趾肌腱腱周滑膜组织为材料,经体外分离、培养和鉴定后,加入不同浓度的CTGF（浓度为0,5,10,20,50,100,200,500 ng/ml）及CTGF抗体（浓度为0,1,2,4,8,16,32,64 μg/ml）分组培养滑膜细胞,并以CCK-8染色法检测滑膜细胞的增殖情况.结果：滑膜细胞增殖在CTGF浓度为5 ng/ml至200 ng/ml时,呈逐步递增趋势,各组之间差异有统计学意义（P＜0.05）;而在200 ng/ml及500 ng/ml浓度组间差异无统计学意义（P＞0.05）,加入浓度为4,8,16,32 μg/mlCTGF抗体时,滑膜细胞增殖呈逐步递减趋势,各组之间差异有统计学意义（P＜0.05）;而CTGF抗体浓度为0,1,2 μg/ml和浓度为32,64μtg/ml时,各组间差异无统计学意义（P＞0.05）.结论：CTGF可促进体外培养的鸡肌腱滑膜细胞增殖,最佳浓度为200 ng/ml;CTGF抗体对滑膜细胞增殖活性可产生一定程度的抑制作用.     

体外诱导大鼠脂肪源间充质干细胞成肌腱潜能的研究

目的 体外诱导脂肪源间充质干细胞(adipose tissue-derived mesenchymal stem cells,AT-MSCs)成肌腱细胞,为临床肌腱损伤的修复提供理论基础.方法 从SD大鼠腹腔脂肪组织中分离出脂肪源间充质于细胞,经生长分化因子-5(growth differentiation factor 5,GDF-5)诱导,使其向肌腱细胞分化.应用形态观察和RT-PCR等方法,从形态和分子水平上对诱导分化的细胞进行鉴定.结果 从SD大鼠腹腔脂肪组织中分离出的AT-MSCs呈散在分布,排列不规则,以星形和多角形细胞为主,能稳定增殖传代.经5、10ng/mL GDF-5诱导8d后,大部分细胞呈梭形.RT-PCR结果显示经GDF-5诱导的细胞,I型和Ⅲ型胶原蛋白的表达明显高于未经诱导的对照组;GDF-5诱导的细胞Bcl-2/Bax均高于对照组.结论 从脂肪组织中可以获得具有多向分化能力的AT-MSCs,并能在体外稳定传代.经GDF-5诱导后,AT-MSCs有向肌腱细胞分化的倾向.GDF-5在一定时间内对细胞凋亡有抑制作用,有利于AT-MSCs向肌腱细胞的分化.AT-MSCs有可能替代骨髓间充质干细胞作为组织工程学的种子细胞,用于肌腱损伤的修复.     

Synoviolin基因修饰滑膜细胞对肌腱术后的生物力学研究

目的探讨在Synoviolin基因修饰滑膜细胞条件下预防肌腱粘连的效果。方法 36只Leghom鸡按随机数字表法分为实验组和对照组,每组18只。将2组鸡的左侧第3、4趾深屈肌腱(Ⅱ区)间腱鞘作部分切除,肌腱切断,并行原位缝合后,实验组将Synoviolin基因重组的腺病毒载体直接注入腱鞘残端表面;对照组注入生理盐水。术后1、3、8周取材,行大体观察、生物力学测试、组织学观察及生化检测。结果实验组在手术区域形成新生腱鞘,对照组手术区域只有瘢痕生成;实验组肌腱的滑动距离、模拟主动屈曲度及肌腱最大延伸率均高于对照组(均P<0.05);实验组术后1、8周肌腱最大抗拉力及腱段羟脯氨酸含量均优于对照组(P<0.05),而术后3周比较差异无统计学意义(P>0.05)。结论 Synoviolin基因修饰滑膜细胞能使无滑膜肌腱滑膜化,提高肌腱愈合质量,有效地预防肌腱粘连。     

肿瘤坏死因子-α对滋养细胞增殖及功能的影响

[目的]探讨肿瘤坏死因子-α(TNF-α)对滋养细胞增殖、凋亡及绒毛膜促性腺激素(HCG)分泌调节的影响.[方法] 建立人早期妊娠滋养细胞体外培养体系,检测不同浓度TNF-α作用下滋养细胞增值状态及HCG的浓度.[结果] TNF-α在低浓度时促进滋养细胞的增殖活性,在高浓度时则抑制其增殖活性; TNF-α浓度为10 ng/mL、100 ng/mL 可使滋养细胞凋亡,其凋亡率随TNF-α浓度升高而增加(P<0.05);在0～100 ng/mL浓度TNF-α范围内,滋养细胞分泌HCG水平随TNF-a浓度的增加而升高.[结论] TNF-α可影响滋养细胞增殖与凋亡;且具有促进滋养细胞分泌HCG的功能.     

生长因子对肌腱细胞表型及分化的影响

目前对于肌腱细胞体外培养的难点较为突出,其中一个难点为,肌腱细胞在体外长时间培养难以维持其表型的表达,继而影响其分化;另外一个难点是,在培养基内添加了大量的胎牛血清,而大量胎牛血清的使用会造成动物-人之间的疾病传播。已有研究显示在培养基中加入不同的生长因子以减少胎牛血清的使用并促进肌腱细胞的分化。本综述较为全面地总结了几种肌腱细胞较为重要的表型和分化标志物以及几种维持肌腱细胞表型的生长因子。     

去甲基化作用对胃癌细胞生物学行为及xaf1基因表达的影响

目的:探讨去甲基化药物5-脱氧杂氮胞苷(5-Aza.CdR)对体外培养的胃癌BGC823细胞增殖活性、细胞周期和凋亡以及对此细胞株xafl基因表达的影响.方法:用MTT法检测不同浓度5-Aza-CdR对细胞增殖活性的影响;PI染色和流式细胞仪检测不同浓度5-Aza-CdR处理72 h后细胞周期分布和细胞凋亡率;RT-PCR法检测用药前后xafl基因表达的变化.结果:用1×103,5×103.10×103nmol/L的5-Aza-CdR处理BGC823细胞6 d后,试验组细胞增殖抑制率较对照组明显升高(P<0.05),并呈剂量依赖关系;流式细胞仪分析表明,各药物浓度处理72 h后凋亡率明显增加:试验组凋亡率分别为(4.53±0.21)%,(8.11±1.01)%和(11.56±0.86)%,与对照组(0.51±0.01)%相比较差异显著(P<0.05).在5-Aza-CdR处理前,未检测到BGC823细胞株xafl基因mRNA表达,经过5-Aza-CdR处理后,xafl mRNA重新表达.结论:5-Aza-CdR可抑制BGC823细胞增殖;促进细胞凋亡;使xafl基因甲基化状态得到逆转,而重新表达.     

姜黄素对人结肠癌细胞SW480增殖周期及凋亡的影响

目的探讨姜黄素对人结肠癌细胞SW480细胞生长的抑制作用及其对细胞增殖周期和凋亡的影响。方法采用甲基噻唑（MTT）比色法和生长曲线测定不同浓度姜黄素在不同作用时间内对在体外培养SW480细胞增殖的影响，同时应用流式细胞术检测细胞增殖周期及凋亡率的变化。结果姜黄素可在G0／G1期阻滞人结肠癌细胞SW480的增殖，使S期细胞比率降低；在一定范围内，姜黄素的浓度越高、作用时间越长，对肿瘤细胞生长的抑制作用越强，凋亡率也越高。结论姜黄素能抑制人结肠癌细胞SW480细胞生长，并诱导细胞凋亡而阻止细胞周期。     

阿霉素对体外培养的猪毛囊真皮鞘细胞增殖和凋亡的影响

目的 探讨阿霉素对体外培养的猪毛囊真皮鞘细胞增殖和凋亡的影响。方法 用MTT法通过比色分析测定吸光度值，检测不同浓液(0、5、10、20mg／L)阿霉素作用24h和10mg/L阿霉素作用不同时间(4、8、12、24、48h)对猪毛囊真皮鞘细胞增殖的影响；用流式细胞仪检测不同剂量的阿霉素作用24h后猪毛囊真皮鞘细胞的凋亡率。结果 阿霉素抑制了体外培养的猪毛囊真皮鞘细胞的增殖，其抑制效应随阿霉素浓度的增大和作用时间的延长而增强；细胞凋亡率随阿霉素浓度增大而升高，阿霉素浓度为10mg/L时细胞凋亡率最高。结论 阿霉素抑制体外培养的猪毛囊真皮鞘细胞增殖，诱导细胞凋亡，其效应与阿霉素的浓度和作用时间相关。     

姜黄素诱导类风湿关节炎滑膜细胞凋亡及其机制研究

目的 观察姜黄素(CUR)对体外培养的类风湿关节炎成纤维样滑膜细胞(RA-FLSs)凋亡的诱导作用及其机制.方法 用不同浓度CUR处理在体外培养的RA-FLSs细胞后,CCK-8法检测CUR对细胞增殖能力的影响;流式细胞术检测CUR对RA-FLSs细胞凋亡的诱导效应;蛋白质免疫印迹方法(WB)检测细胞凋亡的外源途径中相...     

A histological observation on the flexor tendon healing within intact sheath

Summary In this study we allowed the sutured chicken flexor tendons to glide back into the uninjured sheaths in order to keep the healing process of flexor tendon from being affected by the healing of surrounding wounded tissues. By observing 12 chickens, 72 digits, with light microscope and transmission electron microscope, it was found that the visceral and parietal synovium of the sheath were the regions with earliest and most active cell proliferation and the major source of repairing cells during the healing process of the flexor tendon. Tendon cells had the ability of intrinsic healing, but delayed as compared to synovium cells. Adhesion between intact parietal synovium and healing tendon and its surrounding tissue could not be avoided.     

Effect of growth and differentiation factor 6 on the tenogenic differentiation of bone marrow-derived mesenchymal stem cells

Background Recent studies showed that bone marrow-derived mesenchymal stem cells (BMSCs) had risk of ectopic bone formation.In this study,we aimed to investigate the effect of growth and differentiation factor 6 (GDF-6) on the tenogenic differentiation of BMSCs in vitro,and then combined with small intestine submucous (SIS) to promote tendon regeneration in vivo.Methods The BMSCs were isolated from the green fluorescent protein (GFP) rats,and were characterized by multi-differentiation assays following our previous study protocol.BMSCs cultured with different concentrations of GDF-6,without growth factors served as control.After 2 weeks,mRNA expression and protein expression of tendon specific markers were examined by qRT-PCR and Western blotting to define an optimal concentration of GDF-6.Mann-Whitney U-test was used to compare the difference in relative mRNA expression among all groups; P ≤0.05 was regarded as statistically significant.The GDF-6 treated BMSCs combined with SIS were implanted in nude mice and SD rat acute patellar tendon injury model,the BMSCs combined with SIS served as control.After 12 and 4 weeks in nude mice and tendon injury model,the samples were collected for histology.Results After the BMSCs were treated with different concentration of GDF-6 for 2 weeks,the fold changes of the specific markers (Tenomodulin and Scleraxis) mRNA expression were significantly higher in GDF-6 (20 ng/ml) group (P ≤0.05),which was also confirmed by Western blotting result.The BMSCs became parallel in orientation after GDF-6 (20 ng/ml) treatment,but the BMSCs in control group were randomly oriented.The GDF-6 (20 ng/ml) treated BMSCs were combined with SIS,and were implanted in nude mice for 12 weeks,the histology showed neo-tendon formation.In the SD rat patellar tendon window injury model,the histology also indicated the GDF-6 (20 ng/ml) treated BMSCs combined with SIS could promote tendon regeneration.Conclusions GDF-6 has tenogenic effect on the tenogenic differentiation of BMSCs,and GDF-6 (20 ng/ml) has better tenogenic effect compared to other concentrations.The GDF-6 (20 ng/ml) treated BMSCs combined with SIS can form neo-tendons and promote tendon regeneration.     

Experimental studies of flexor tendon repair in "no man's land"

The FDP of the middle toes of 51 chickens were divided and repaired. All of the chickens were divided into three groups, each had a procedure of protected passive mobilization beginning at 1, 5, 21 postoperative days respectively. Through studies by micrography, light-microscopy, transmission and scanning electronmicroscopy, we found that passive motion beginning at 1 day and 5 days after operation has similar effects on the healing tendons by preventing adhesions, stimulating proliferation of epitenon and remoulding tendon callus. On the basis of the above results, another 32 chickens were used for the following experiments: The effects of early stage passive motion on the results of double tendon repair and free tendon graft in the "No Man's Land". It was disclosed that passive motion can significantly prevent the adhesions between FDS and FDP, but this was less effective on adhesions between FDS nd sheath. For tendon graft, the results of the mobilized side were extraordinarily good, and the free tendon itself had taken part in the healing process.     

腱鞘移植在严重腱周组织损伤中肌腱修复的应用

为探索屈指肌腱鞘背侧重建技术的屈指肌腱修复效果。对24例屈指肌腱Ⅱ区损伤手指,运用游离腱鞘移植修复其屈指腱背侧腱鞘。游离腱鞘取自同侧手腕背第一间隔。屈指腱鞘背侧重建技术用于早期或延迟早期屈指肌腱损伤、二期肌腱移植修复和肌腱粘连松解术时有骨面裸露或挫伤者。根据Strickland评价标准,结果优7指,良11指,中5指,差1指,优良率为75％。用游离自体移植腱鞘重建背侧屈指腱鞘,是提高严重腱周组织损伤的肌腱修复的一种有效方法。     

黄芪注射液对乳腺癌MCF-7细胞增殖周期以及细胞凋亡、迁移影响的实验研究

目的 探讨黄芪注射液对乳腺癌MCF-7细胞增殖、周期、凋亡及迁移的影响。方法 通过MTT法观察药物处理后对乳腺癌MCF-7细胞的生长增殖情况的影响;通过流式细胞术检测黄芪注射液对乳腺癌MCF-7细胞增殖周期、凋亡的影响;利用划痕实验检测黄芪注射液对乳腺癌MCF-7细胞的迁移的影响。结果 MTT结果发现,不同浓度的黄芪注射液（100、200、400、600、800mg/ml）对乳腺癌MCF-7细胞的增殖具有不同程度的抑制作用;流式细胞术结果显示,与空白对照组相比,200和400mg/ml黄芪注射液均能导致G₁期增加,且两个浓度的凋亡率都大于空白对照组;划痕实验结果显示200和400mg/ml黄芪注射液组较空白对照组划痕愈合率低。结论 不同浓度的黄芪注射液在体外对乳腺癌MCF-7细胞增殖具有一定的抑制作用;同时可将细胞增殖周期阻滞在G₁期,促进MCF-7细胞凋亡;一定浓度的黄芪注射液对MCF-7细胞的迁移具有抑制作用,为乳腺癌的临床应用提供实验基础。     

过度运动对大鼠跟腱组织及腱细胞凋亡的影响

目的探讨过度运动对大鼠跟腱组织和腱细胞的影响。方法制作大鼠跟腱过度运动动物模型。采用40只清洁级S-D大鼠随机均分为8组,其中运动组7组,对照组1组,每组5只。运动组大鼠按跑台训练(17m/min,1h/d,5d/w),分为1、2、3、4、5、6w组(每组5只),分别在复合麻醉下处死后取出运动组和对照组大鼠的跟腱,另一组训练6w后,再于笼内休息2w后麻醉下取材,为8w运动组。对照组大鼠于笼内自由活动,视为0w组。行常规HE染色,观察跟腱细胞的形态学改变,行免疫组化TUNEL染色,测定跟腱在过度运动不同时间段腱细胞凋亡指数及与跟腱细胞凋亡有关的蛋白Caspase-3活性的表达。结果过度运动1、2、3w组HE观察未见明显细胞排列紊乱及腱细胞水肿断裂,TUNEL法及Caspase-3活性检测跟腱细胞凋亡不明显。4、5、6w组HE观察见跟腱止点处有类软骨细胞增生,TUNEL法及Caspase-3检测跟腱细胞凋亡,明显高于对照组(P<0.05)。8w组跟腱细胞凋亡指数高于对照组(P<0.05),而与5、6w组相比差异均无统计学意义(P>0.05)。结论过度运动可引起跟腱细胞损伤,使跟腱细胞内出现类软骨样细胞及腱细胞的凋亡和Caspase-3活化表达的增高。     

腓肠肌肌腱皮瓣修复术治疗陈旧性跟腱断裂

目的：探讨腓肠肌肌腱皮瓣修复术治疗陈旧性跟腱断裂的临床疗效。方法：2001年1月-2007年1月我院采用腓肠肌肌腱皮瓣修复术治疗陈旧性跟腱断裂15例,术后6周去长腿石膏后托进行功能康复训练,术后随访1年,采用Arner-Lindholm疗效评定标准评估。结果：13例患者伤口均为一期愈合,无感染、生硬和跟腱再次断裂等发生;2例肌皮瓣皮肤发生部分坏死,行植皮术后延期愈合。Arner-Lindholm疗效评定,优9例,良4例,差2例,优良率为86.67%。结论：采用腓肠肌肌腱皮瓣修复术治疗陈旧性跟腱断裂能较好地恢复跟腱的功能,近、远期疗效满意。     

Effects of substance P on growth of fibroblast-like cells derived from the bile duct: an in vitro cell culture studyReferences

Background Possible role of substance P (SP) during wound healing has been the primary research focus in recent years, but its effect on the healing process after bile duct injury is little understood. The present study aimed to investigate the effects of SP on growth of fibroblast-like cells derived from rabbit bile duct. Methods Fibroblast-like cells derived from rabbit bile duct were identified and divided randomly into the control and the experimental groups. SP-treated cells at different concentrations at 10-9-10-5 mol/L, and control group were incubated respectively for 48 h. After incubating, effects of SP on cell proliferation were assessed by cell counts and MTT test. Apoptosis rate of cells was measured by flow cytometry. Results Cultured rabbit bile duct cells were fibroblast-like in morphology, and these cells stained positively for vimentin and negatively for desmin. After SP was added to nonconfluent cells for 48 h, cells numbers were significantly increased in experimental groups than that of controls (P < 0.05). The maximum stimulation of cell proliferation was achieved at SP of 10-5 mol/L. Bile duct fibroblast-like cells in the SP group showed a higher proliferating activity and lower apoptosis rate than those in the control group or in the SP Spantide group (P < 0.05). Spantide partly inhibited the effects of SP on fibroblast-like cells. Examination under transmission electron microscope revealed rough endoplasmic reticulum and prominent Golgi complexes after SP treatment. Conclusions SP has a growth regulatory property on cultivated bile duct fibroblast-like cells in vitro, suggesting that SP may involve in wound healing after bile duct injury by promoting wound fibroblast proliferation and inhibiting apoptosis and participate in pathological scar formation.