基质金属蛋白酶-3和血管内皮生长因子在不同月龄大鼠椎间盘组织中的表达及意义

目的检测不同月龄大鼠的椎间盘组织中基质金属蛋白酶-3(MMP3)和血管内皮生长因子(VEGF)的表达,探讨MMP3和VEGF与椎间盘自发退变的关系。方法采用苏木精-伊红(HE)染色和链酶亲和素-过氧化物酶复合物(SABC)免疫组化方法及显微图像分析技术,测定50只Wistar大鼠(1、3、6、12、18月龄各10只)椎间盘组织中MMP3和VEGF的表达情况并观察椎间盘组织学变化。结果随着大鼠月龄的增长,其椎间盘中MMP3和VEGF的表达亦发生变化,由低月龄组(1-3月龄)至成龄组(12月龄)MMP3表达逐渐增多,而VEGF表达逐渐下降,且这种变化有显著统计学意义(P<0.01)。椎间盘的组织结构也发生变化,随月龄增加,椎间盘中软骨细胞数量减少,胶原纤维增生、粗大、走行紊乱。结论大鼠椎间盘结构随月龄增长发生性改变,椎间盘中MMP3和VEGF表达发生变化,这可能引起椎间盘的自发退变,而且这种改变与人的椎间盘的自然退变具有相似性和可比拟性。     

缺氧诱导因子-1α和葡萄糖转运蛋白-1在不同月龄大鼠椎间盘中的表达

目的检测缺氧诱导因子-1α(hypoxia-inducible factor-1α,HIF-1α)和葡萄糖转运蛋白-1(glucose transporter-1,GLUT-1)在不同月龄大鼠椎间盘纤维环组织中的表达及其相关性,探讨HIF-1α及GLUT-1在椎间盘退变过程中的作用。方法取Wistar大鼠50只,分别以1,3,6,12,18个月龄分为5组。采用免疫组化法及Western blot法检测各组椎间盘中HIF-1α和GLUT-1表达情况。结果随着大鼠月龄的增长,其椎间盘纤维环组织中HIF-1α和GLUT-1的表达也发生变化,由低月龄组(1-3月龄)至成年组(6-12月龄)HIF-1α和GLUT-1表达逐渐减少,而老年组(18月龄)二者表达显著增加。且这种变化有显著统计学意义(P0.01)。纤维环中HIF-1α和GLUT-1的蛋白表达呈正相关。结论HIF-1α、GLUT-1表达水平的变化与椎间盘退变的发生关系密切相关,HIF-1α可以通过上调GLUT-1等相关因子并延缓椎间盘退变,可能作为椎间盘退变治疗研究的切入点。     

Bcl2、Fas在人颈椎间盘软骨终板、纤维环及髓核中的表达及意义

目的 检测正常和突出的人颈椎间盘软骨终板、纤维环及髓核中Bcl2、Fas的表达情况,探讨其在各组中不同表达的意义。方法 先将各正常及突出颈椎间盘在解剖显微镜下分出软骨终板、纤维环、髓核并进行苏木精-伊红（HE）染色行形态学观察,再用免疫组化（SABC法）检测Bcl2、Fas在各组中的表达情况,并进行计算机显像系统图像分析。结果 Bcl2的表达在正常颈椎间盘的软骨终板、纤维环、髓核中均高于突出颈椎间盘的各相应组;Fas的表达在正常颈椎间盘的软骨终板、纤维环、髓核中均低于突出颈椎间盘的各相应组;在异常颈椎间盘的软骨终板中Bcl2的表达低于其在纤维环和髓核中的表达（P〈0.01）;Fas的表达高于其在纤维环和髓核中的表达（P〈0.01）;Bcl2及Fas的表达在纤维环和髓核中的差别无统计学差异（P〉0.05）。结论 细胞凋亡是椎间盘退变的重要原因,且凋亡可能始于软骨终板。     

PeroxiredoxinⅡ在退变椎间盘髓核中的表达及其临床意义

目的：检测PeroxiredoxinⅡ在腰椎间盘髓核组织中的表达,分析其在椎间盘退变中的临床意义。方法：用蛋白免疫印迹（Western blot）的方法检测PeroxiredoxinⅡ在正常、突出及脱出腰椎间盘髓核中的表达情况。结果：PeroxiredoxinⅡ在退变椎间盘髓核中表达丰富,而在正常椎间盘髓核中表达微弱,两者比较差异显著（P〈0.05）,在突出和脱出椎间盘髓核中表达无显著性差异（P〉0.05）。结论：PeroxiredoxinⅡ在正常及退变腰椎间盘髓核组织中差异表达。     

缺氧诱导因子-1α和血管内皮生长因子在突出腰椎间盘中的表达及意义

目的探讨缺氧诱导因子-1α（hypoxia—induciblefactor-1α，HIF-1α）和血管内皮生长因子（vascular endothelial growth factor，VEGF）在突出腰椎间盘组织中的表达及意义。方法采用链霉亲和素-过氧化物酶复合物（SABC）免疫组化方法，测定40例腰椎间盘突出症患者椎间盘组织中HIF-1α和VEGF的表达情况。结果退变椎间盘组织中HIF-1α和VEGF呈高表达，HIF-1α和VEGF在髓核的表达显著高于纤维环；纤维环破裂型显著高于纤维环完整型；各组中HIF-1α和VEGF的表达均高度相关。结论HIF-1α和VEGF共同参与了椎间盘退变；HIF-1α可能通过上调VEGF的表达来促进椎间盘组织中新生血管的形成，进而延缓椎间盘退变的发生。     

TGF-β1/Smad 2/3信号通路在椎间盘退变中的作用

临床实践表明,富含血小板的血浆(PRP)注射是延缓椎间盘退变的有效方法,但其作用机制尚不清楚。已有研究表明,活化的血小板可释放转化生长因子-β1 (TGF-β1),它可以正向调节髓核细胞的细胞外基质。本研究旨在揭示TGF-β1/Smad 2/3信号通路在PRP缓解椎间盘退变过程中的作用机制。本研究通过制备新西兰白兔PRP,并使用PRP和TGF-β1抑制剂(SB431542)处理白兔髓核细胞,检测PRP对髓核细胞增殖、软骨形成标志物(CollagenⅡ和Aggrecan) m RNA的表达、Smad 2/3及相关髓核细胞功能蛋白表达的影响。结果显示,用2%PRP处理的髓核细胞的增殖显著增强。PRP处理显著增加髓核细胞中的CollagenⅡ和Aggrecan的mRNA水平。Western blotting结果显示,TGF-β1信号通路的特异性抑制剂可显著抑制Smad 2/3和基质蛋白的表达。在兔椎间盘退变模型中,PRP+抑制剂注射组的Smad 2/3和CollagenⅡ的表达水平显著低于PRP处理组。这些结果表明,PRP中由于TGF-β1含量高,可激活TGF-β1/Smad 2/3途径,并促进CollagenⅡ和其他基质成分的合成和分泌,从而延缓了椎间盘退变。     

持久脊柱负荷与椎间盘组织中MMP-2表达关系的研究

目的：建立一压力可控型椎间盘退变模型,并探讨持久的脊柱负荷对椎间盘MMP-2表达的影响。方法：选用54只成年Wistar大鼠随机分为三组,分别模拟人类在站立（A组1.12N）、坐位直立（B组1.68N）、坐位前屈（C组3.08N）三种状态下椎间盘内的负荷情况,给予大鼠尾椎Co9/10椎间盘恒定压力加压,以相邻Co8/9椎间盘不加压作为对照（D组）。三组分别在3、7、14天后取受压及对照椎间盘标本,进行HE染色组织学观察及免疫组织化学分析,观察椎间盘退变情况及MMP-2在椎间盘组织中的含量变化。结果：随时间与压力的增加,椎间盘组织学评分与MMP-2表达增高（P〈0.05）,MMP-2表达与椎间盘退变程度成正相关（r=0.870,P〈0.05）。结论：持久的脊柱负荷可引起椎间盘退变及MMP-2表达增加,MMP-2可能在椎间盘退变的过程中发挥重要作用。     

Sox9治疗兔椎间盘退变的效果及机制研究

目的:探究Sox9用于治疗椎间盘退变的效果及调控机制。方法:将Ad-sox9和Ad-GFP各20μL分别转染至椎间盘退变兔的髓核组织中,转染后3、7、30、60天取材,采用免疫组化、免疫荧光和MRI等研究方法检测椎间盘髓核组织中II型胶原、蛋白多糖的表达情况,并分析对椎间盘退变的改善情况。结果:免疫组化染色显示sox9组中椎间盘髓核组织中II型胶原、蛋白多糖的表达明显升高,MRI显示sox9组椎间盘T2像信号有明显改善(P<0.05)。结论:体内转染腺病毒介导的sox9基因能够增加椎间盘内II型胶原和蛋白多糖的表达,并抑制椎间盘的退变进程。     

正常与退变椎间盘髓核和纤维环中生化成份的研究

本工作测定了椎间盘的髓核和纤维环中的生化成份,结果提示在退变的椎间盘中,糖醛酸,总己糖胺、半乳糖胺均明显下降,而羟脯氨酸的含量明显增加。在退变的纤维环中尚有组成硫酸角质素的主要成份葡萄糖胺量的明显下降。在退变的髓核和纤维环中,总蛋白质含量呈增加趋势,水的含量呈下降趋势。提示退变椎间盘中蛋白多糖含量降低,组成改变及胶原蛋白的增加可能是诱发椎间盘突出症的物质基础。     

Peroxiredoxin Ⅱ在退变椎间盘髓核中的表达及其临床意义

目的:检测PeroxiredoxinⅡ在腰椎间盘髓核组织中的表达,分析其在椎间盘退变中的临床意义。方法:用蛋白免疫印迹(Western blot)的方法检测PeroxiredoxinⅡ在正常、突出及脱出腰椎间盘髓核中的表达情况。结果:PeroxiredoxinⅡ在退变椎间盘髓核中表达丰富,而在正常椎间盘髓核中表达微弱,两者比较差异显著(P<0.05),在突出和脱出椎间盘髓核中表达无显著性差异(P>0.05)。结论:PeroxiredoxinⅡ在正常及退变腰椎间盘髓核组织中差异表达。     

Effects of mechanical loading on the expression of pleiotrophin and its receptor protein tyrosine phosphatase beta/zeta in a rat spinal deformity model

Mechanical loading of the spine is a major causative factor of degenerative changes and causes molecular and structural changes in the intervertebral disc (IVD) and the vertebrae end plate (EP). Pleiotrophin (PTN) is a growth factor with a putative role in bone remodeling through its receptor protein tyrosine phosphatase beta/zeta (RPTPβ/ζ). The present study investigates the effects of strain on PTN and RPTPβ/ζ protein expression in vivo. Tails of eight weeks old Sprague-Dawley rats were subjected to mechanical loading using a mini Ilizarov external apparatus. Rat tails untreated (control) or after 0 degrees of compression and 10°, 30° and 50° of angulation (groups 0, I, II and III respectively) were studied. PTN and RPTPβ/ζ expression were evaluated using immunohistochemistry and Western blot analysis. In the control group, PTN was mostly expressed by the EP hypertrophic chondrocytes. In groups 0 to II, PTN expression was increased in the chondrocytes of hypertrophic and proliferating zones, as well as in osteocytes and osteoblast-like cells of the ossification zone. In group III, only limited PTN expression was observed in osteocytes. RPTPβ/ζ expression was increased mainly in group 0, but also in group I, in all types of cells. Low intensity RPTPβ/ζ immunostaining was observed in groups II and III. Collectively, PTN and RPTPβ/ζ are expressed in spinal deformities caused by mechanical loading, and their expression depends on the type and severity of the applied strain.     

IL-13抑制剂在损伤大鼠尾椎间盘退变中的作用研究

为了探讨IL-13细胞因子在损伤后大鼠椎间盘退变中的影响,建立了大鼠尾椎间盘退变模型,给予IL-13抑制剂sIL-13Rα2-Fc进行干预,将实验分为空白、对照、低剂量、中剂量、高剂量干预组。分别于4周及6周后通过HE染色和Masson染色观察椎间盘形态变化并评分;DMMB法定量分析椎间盘中的糖胺多糖(glycosaminoglycan,GAG)、硫酸软骨素(chondroitin sulfate,CS)、硫酸角质素(keratan sulfate,KS)、透明质酸(hyaluronic acid,HA)含量变化;RT-PCR分析Ⅰ型和Ⅱ型胶原蛋白的mRNA表达水平;蛋白质印迹分析Ⅰ型和Ⅱ型胶原蛋白含量。HE和Masson染色显示与对照组相比,干预组椎间盘病理改变减小,纤维环排列更规则,破裂部位减小,NP细胞数量增加,胶原纤维减少。sIL-13Rα2-Fc干预增加了糖胺多糖、透明质酸含量,增加了硫酸软骨素/硫酸角质素比,减少了Ⅰ型胶原蛋白的表达,并增加了Ⅱ型胶原蛋白。结果表明IL-13抑制剂sIL-13Rα2-Fc可有效减轻椎间盘退变,并且与作用时间和浓度成正相关。     

Fibronectin content of the annulus fibrosus in diabetic and non-diabetic sand rats

Annulus fibrosus of intervertebral discs from diabetic and non-diabetic sand rats were examined by microspectrophotometry for fibronectin content. This was higher in the diabetic animals both in the dorsal and ventral parts and in the outer and inner lamellae of the annulus. It is suggested that diabetes-related changes in fibronectin are similar to changes in annular collagen observed in species other than sand rats.     

正常山羊腰椎间盘营养途径的DCE-MRI观察

目的 研究正常山羊腰椎间盘软骨终板营养途径.方法 选取健康24月龄山羊8只,每只山羊观察4个腰椎间盘,共32个腰椎间盘.麻醉后,行磁共振动态增强扫描,观察感兴趣区的信号变化特点.分别测量增强前及增强后0 min、5 min、10 min、30 rain、1 h、1.5 h、2 h,2.5 h、3 h、3.5 h感兴趣区信号强度值,分析时间-信号强度曲线及峰值出现时间.结果 椎体磁共振信号强度在0 min时达到高峰后迅速下降;软骨终板区在30 min时缓慢达到第一高峰后轻度下降,于2 h上升达到第二高峰;髓核在5 min内为负值,之后缓慢上升于2 h达到高峰,随后逐渐下降.结论 正常山羊腰椎椎间盘主要通过软骨终板途径进行营养代谢.     

Dexamethasone treatment affects nuclear glucocorticoid receptor and glucocorticoid response element binding activity in liver of rats (Rattus norvegicus) during aging

Aging is associated with marked changes in the biochemical processes of many organs. Basal and glucocorticoid induced of liver nuclear glucocorticoid receptor (GR) on the level of protein expression and DNA-binding activity were investigated at different ages (3, 6, 12, 18 and 24 months old) in two groups of rats in: untreated and dexamethasone treated. The results showed a significant decline of GR protein immunopurified from untreated rats of advanced age. In dexamethasone-treated rats, the quantity of GR protein was lower than in controls at all ages. The interactions of liver nuclear proteins with radioactively labelled synthetic oligonucleotide analogue containing consensus GRE sequence were analysed during aging. The results showed that GRE binding activity demonstrated a decrease both in untreated and in dexamethasone treated rats. However, relative to untreated rats, dexamethasone treatment resulted in a significant increase in GRE binding at all ages, except that of three months old animals. In conclusion, the observed alterations in GR protein expression and its DNA binding activity may play a role in the changes of the cell response to glucocorticoid during aging.     

Effects of pulsed electromagnetic field on intervertebral disc cell apoptosis in rats

Despite numerous studies on pulsed electromagnetic field (PEMF) application, its effects of PEMF on intervertebral disc (IVD) have not yet been investigated in vivo. Accordingly, the effects of PEMF upon IVD in rats were evaluated through molecular surveys. Rats were divided into six groups: Group I and II were exposed to low and high frequency of PEMF (LF and HF, respectively). Group III and IV underwent induced disc degeneration and were exposed to low and high frequency of PEMF (LF/IDD and HF/IDD, respectively). Group V underwent induced disc degeneration (IDD), and group VI was control. The values of caspase 3, Bax, Bcl-2 and β-actin band density, as cell apoptotic markers, were obtained from band densitometry. Our results showed that the value of cleaved caspase-3 of cells and Bax/Bcl-2 ratio in IDD group increased significantly compared to the control group (p?p?相似文献   

Immunohistochemistry of chondromodulin-I in the human intervertebral discs with special reference to the degenerative changes

The expression of the matrix protein chondromodulin-I has been studied in human intervertebral discs of 101 people using immunohistochemical analyses. The purpose of this report is to present data on the metabolic changes that were found to occur in the chondrocytes of intervertebral discs during development and aging. Chondromodulin-I was highly expressed during the gestational period and gradually decreased after maturation. It was detected in both the extracellular matrix and chondrocytes in the zone of hypertrophic cartilage, the zone of proliferative cartilage and the zone of resting cartilage in fetal discs. It was also present in the annulus fibrosus, nucleus pulposus and end-plate cartilage in mature discs. In degenerative discs, chondromodulin-I immunoreactivity tended to be elevated in the remaining chondrocytes. Our findings suggest that the expression of the protein is developmentally regulated and upregulated through a defense mechanism against the degenerative processes of the aged intervertebral disc.     

椎间盘脱出大鼠模型的脊髓血流量变化及电针效应

目的 观察大鼠椎间盘脱出模型的脊髓血流量及其电针效应,为兽医临床犬椎间盘病病理和针灸作用机理研究积累资料.方法 通过外科手术将硅胶片填充于大鼠第13胸椎( T13)脊髓腹侧位制作胸腰段椎间盘脱出模型,通过激光散斑血流系统实时监测造模后第一腰椎(L1)脊髓背侧血流量及电针刺激双侧足三里穴和趾间穴对其的影响.结果 以硅胶片填充于大鼠T13的脊髓腹侧位可成功制作出类似犬胸腰段椎间盘脱出的后肢瘫痪模型,且重复性好;模型组大鼠在压迫后,L1段的脊髓背侧血流量呈极显著下降(P＜0.01),电针双侧足三里穴和趾间穴可显著升高血流量(P＜0.05),电针结束后约10 min其血流量逐渐恢复至电针前水平;电针组大鼠电针治疗14 d后,脊髓背侧血流量极显著高于对照组,且运动功能评分显示电针组大鼠的后肢运动功能显著改善.结论 通过外科手术将硅胶片填充于大鼠T13脊髓腹侧位可成功制作胸腰段椎间盘脱出模型;压迫后L1段脊髓血流量显著下降;电针治疗后脊髓血流量极显著升高,且瘫痪大鼠运动机能明显好转,这可能与电针改善脊髓血流量继而减轻脊髓损伤并促进损伤恢复或促进代偿机制有关.