人椎间盘软骨细胞外基质小分子蛋白的双向电泳和质谱鉴定

目的初步研究双向电泳和质谱鉴定在人椎间盘的蛋白质组特性研究中的价值和作用．为进一步研究椎间盘疾病寻找有效途径。方法取人的正常椎间盘组织（包括纤维环和髓核），针对细胞外基厦分子蛋白进行蛋白萃取，通过氯化铯梯度离心去除蛋白多糖，通过固相pH梯度（IPG）等电聚焦和梯度聚丙烯酰胺双向电泳分别分离纤维环和髓核的蛋白质．分析电泳图像．使用基质辅助激光解吸附飞行时间质谱仪对蛋白质点进行分析鉴定。结果双向电泳显示。纤维环和髓核的细胞外基质小分子蛋白无明显差异．纤维环和髓核中的小分子蛋白在pH3～10、分子量16～250kd范围内能被很好地分离。对各蛋白质点行质谱仪鉴定．分别确定了萁中相同的19种蛋白质。结论双向电泳可有效地分离准间盘组织中细胞外基质小分子蛋白，通过质谱仪鉴定可以确定其组成成分，为进一步研究椎间盘软骨细胞外基质小分子蛋白的功能及在准间盘病变中的改变和所起的作用打下基础。     

Extracephalic jabs/idiopathic stabs. Vågå study of headache epidemiology

Ultrashort cephalic paroxysms are well known. In the parish of V?g?, Norway, 35.2% of the 18-65-year-old subjects (n = 1779) were recently found to have such jabs. In the present work, a search has been made for extracephalic 'jabs'. A questionnaire was in its entirety administered by the same investigator (O.S.) in a 'semistructured' way. Facial jabs were present in three women, and in one of them the pain spread to the head. Four subjects had jabs occurring at random throughout the body, also including the cephalic area. Pure nuchal jabs were present in 12 subjects, 10 of whom were males. This sex preponderance difference differs significantly from that in jabs in general (with 40.2% males). The characteristics of the extracephalic jabs, i.e. the duration and temporal pattern, do not seem to differentiate them essentially from jabs in general. The subjects were not asked specific questions regarding extracranial jabs. Most of the affected individuals gave information spontaneously about their jabs. For these reasons, this study is not a proper prevalence study. It does show, however, that extracranial jabs exist, and it gives some indications as to their frequency.     

Effects of Hydration and Fixed Charge Density on Fluid Transport in Charged Hydrated Soft Tissues

The effects of tissue hydration and fixed charge density on hydraulic permeability and creep behavior of cartilaginous tissues have been investigated using the triphasic theory and finite element methods. The empirical model for hydraulic permeability of uncharged gels and Mackie and Meares (1955) model for ion diffusivity were used in the numerical analysis. The hydraulic permeabilities of normal and trypsin-treated porcine annulus fibrosus tissues were measured indirectly. Analysis of the experimental data from this study and in literature indicates that the water content plays a more important role in regulating tissue permeability than fixed charge density for normal tissues. A change in glycosaminoglycan content will change both triphasic closed-circuit (or intrinsic) and biphasic open-circuit permeabilities of cartilaginous tissues. Analysis also shows that both fixed charge density and water content play an important role in tissue creep response. This study adds new knowledge to the permeability and creep behavior of cartilaginous tissues and is important for understanding the nutrition in intervertebral disk. © 2003 Biomedical Engineering Society. PAC2003: 8719Tt, 8710+e, 8719Uv     

Diffusivity of Ions in Agarose Gels and Intervertebral Disc: Effect of Porosity

The effect of tissue porosity on ion (sodium, potassium, and chloride) diffusivity in agarose gels and porcine intervertebral disc tissues was investigated using an electrical conductivity method. An empirical, constitutive model for diffusivity (D) of solutes in porous fibrous media was proposed: D/D_o=exp[–(r_s/^1/2)] where r_s is the Stokes radius of a solute, is the Darcy permeability of the porous medium, D_o is the diffusivity in free solution, and are two positive parameters whose values depend on material structure. It is found that =1.25±0.138, =0.681±0.059 (95% confidence interval, R²=0.92, n=72) for agarose gels and =1.29±0.171 and =0.372±0.088 (95% confidence interval, R²=0.88, n=86) for porcine annulus fibrosus. The functional relationship between solute diffusivity and tissue deformation was derived. Comparisons of our model prediction with experimental data on diffusion coefficients of macromolecules (proteins, dextrans, polymer beads) in agarose gels in the literature were made. Our results were also compared to the data on ion diffusivity in charged gels and in cartilaginous tissues reported in the literature. There was a good agreement between our model prediction and the data in the literature. The present study provides additional information on solute diffusivity in uncharged gels and charged tissues, and is important for understanding nutritional transport in avascular cartilaginous tissues under different mechanical loading conditions.     

Effects of swelling pressure and hydraulic permeability on dynamic compressive behavior of lumbar annulus fibrosus

The objective of this study was to investigate the effects of swelling pressure and hydraulic permeability on the dynamic behavior of intervertebral disk tissue in confined compression. Normal (served as a control) and trypsin-treated, axial annulus fibrosus (AF) specimens from the porcine lumbar disks were tested and their swelling strain, swelling pressure, equilibrium compressive modulus (H _A, dynamic modulus, and hydraulic permeability (k) were determined at 30% and 40% swelling strain levels. The proteoglycan depletion due to trypsin treatment resulted in significantly lower values of the free swelling strain, swelling pressure, equilibrium modulus, dynamic modulus, and higher value of hydraulic permeability for trypsin-treated group, comparing to those for the control group. At the 30% swelling strain level, the equilibrium moduli were 51.84±14.53 kPa (n=8) for the control group and 15.11±5.67 kPa (n=8) for the trypsin-treated group; and the hydraulic permeabilities were 4.50E-15±1.60E-15 m⁴/Ns and 8.43E-15±4.29E-15 m⁴/Ns for control and trypsin-treated groups, respectively. No statistically significant difference in wet tissue density or dry tissue density was found between control and trypsin-treated groups. There was a significant correlation between swelling pressure and compressive (aggregate) modulus (R²=0.824, m=22). The decrease in measured dynamic modulus for trypsin-treated group was attributed to the reduced swelling pressure (or modulus H _A and increased hydraulic permeability (k) due to PG depletion. © 2002 Biomedical Engineering Society. PAC2002: 8719Rr, 8719Tt, 8360Jk     

The implantation of non-cell-based materials to prevent the recurrent disc herniation: an in vivo porcine model using quantitative discomanometry examination

Recurrent disc herniation is frequently observed due to leakage of nucleus pulposus through injured anulus fibrosus. There is no effective treatment to prevent recurrent disc herniation yet. In this study, we proposed to implant non-cell-based materials into the porcine disc to stimulate the growth of fibrous tissue and thereby increase the disc functional integrity. The disc herniation was simulated by anular punctures using the spinal needles. Four clinically used implantation materials, i.e., gelfoam, platinum coil, bone cement and tissue glue, were delivered into the discs via percutaneous spinal needles. Two months after the surgery, the swine were killed. The degree of disc integrity of intact, naturally healed and implanted discs, was examined by quantitative discomanometry apparatus. We found the disc injury could not recover after 2 months of healing, and the disc implantation affected the degree of disc integrity. The disc integrity of gelfoam-implanted discs was better than that of coil-, bone cement-, and glue-implanted discs. The implantation of non-cell-based material was proved to be a potentially clinically applicable method to recover the integrity of injured discs and to prevent recurrent disc herniation.     

兔椎间盘纤维环细胞单层和立体培养条件下细胞表型的比较

目的 观察单层培养和藻酸盐立体培养对兔椎间盘纤维环细胞基因表达的影响.方法 10只新西兰大白兔腰椎椎间盘纤维环细胞经体外单层培养扩增后取第一代细胞分别同时行单层培养和藻酸盐立体培养1周,通过实时RT-PCR检测细胞Ⅰ型胶原、Ⅱ型胶原和蛋白聚糖基因不同的表达情况.结果 在相差显微镜下,单层培养的椎间盘纤维环细胞呈贴壁生长的梭形或多角形细胞,其Ⅰ型胶原基因表达最高,Ⅱ型胶原基因表达其次,蛋白聚糖基因表达最低,符合成纤维细胞的细胞表型.而在藻酸盐凝胶珠内,同样的纤维环细胞呈圆形,被一层藻酸盐凝胶膜囿于凝胶珠内,其Ⅰ型胶原,Ⅱ型胶原基因表达明显下降,而蛋白聚糖基因明显升高,成为该培养条件下纤维环细胞表达最高的基因.结论 藻酸盐立体培养对兔椎间盘纤维环细胞维持软骨样细胞基因表型具有一定作用.     

新型组织工程椎间盘纤维环支架材料的制备和形态学观察

目的 探讨脱矿脱细胞骨基质环结构特点是否适合用于构建组织工程椎间盘纤维环细胞支架.方法 取兔股骨近端骨制成环状,经改进的脱矿脱细胞方法处理并制成支架材料,进行大体、光镜、电镜观察并测量平均孔径.结果 材料为环状,质地柔韧,多孔结构;HE染色光镜观察显示支架材料为嗜酸性,为不均匀的多孔结构;扫描电镜观察发现支架材料为相互沟通的多孔结构,平均孔径为99μm.结论 脱矿脱细胞骨基质环结构特点适合作为组织工程椎间盘纤维环细胞支架材料.