Age-related changes in fibromodulin and lumican in human intervertebral discs.

STUDY DESIGN: An analysis of proteoglycans of the intervertebral disc using immunoblotting of tissue extracts. OBJECTIVES: To investigate the changes in structure and abundance of fibromodulin and lumican in human intervertebral discs during aging and degeneration. SUMMARY OF BACKGROUND DATA: Fibromodulin and lumican are keratan sulfate proteoglycan constituents of the disc's extracellular matrix, whose interaction with collagen fibrils may contribute to the mechanical properties of the tissue. Changes in their abundance and/or structure that occur with aging and degeneration therefore may have an impact on disc function. METHODS: Lumbar intervertebral discs were obtained from individuals of different ages, and extracts of anulus fibrosus and nucleus pulposus were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting using antibodies specific for fibromodulin and lumican. RESULTS: The major changes in abundance observed with age were a decrease in fibromodulin in the adult nucleus pulposus and an increase in lumican in anulus fibrosus during early juvenile development. In addition, fibromodulin in the anulus fibrosus exhibited a structural change with increasing age, characterized by a shift toward the predominance of its glycoprotein form lacking keratan sulfate. Fibromodulin was more abundant in the anulus fibrosus than in nucleus pulposus at all ages, whereas lumican was much more abundant in nucleus pulposus than in anulus fibrosus in the young juvenile; in the adult, however, lumican was present in comparable levels in both tissues. With increasing degrees of degeneration, fibromodulin exhibited an increase in abundance. CONCLUSIONS: Growth, aging, and degeneration of the intervertebral disc are associated with changes in the abundance and structure of fibromodulin and lumican, which presumably influence the functional properties of the tissue.     

Viscoelastic properties of intervertebral disc cells. Identification of two biomechanically distinct cell populations

STUDY DESIGN: A combined experimental and theoretical biomechanical study to quantify the mechanical properties of living cells of the porcine intervertebral disc. OBJECTIVES: To quantify zonal variations in the mechanical properties and morphology of cells isolated from the intervertebral disc. SUMMARY OF BACKGROUND DATA: Cellular response to mechanical stimuli is influenced by the mechanical properties of cells and of the extracellular matrix. Significant zonal variations in intervertebral disc matrix properties have been reported. No information is currently available on the corresponding regional variations in the mechanical properties of intervertebral disc cells, despite evidence of significant differences in cellular phenotype and biologic response to loading. METHODS: The micropipette aspiration test was used in combination with a three-parameter viscoelastic solid model to measure the mechanical properties of cells isolated from the anulus fibrosus, transition zone, and nucleus pulposus. RESULTS: Intervertebral disc cells exhibited viscoelastic solid behaviors. Highly significant differences were observed in the morphology, cytoskeletal arrangement, and biomechanical properties of the nucleus pulposus cells as compared with anulus fibrosus or transition zone cells. Cells of the nucleus pulposus were approximately three times stiffer and significantly more viscous than cells of the anulus fibrosus or transition zone. CONCLUSIONS: The findings of this study provide new evidence for the existence of two biomechanically distinct cell populations in the intervertebral disc. These differences in mechanical behavior may be related to observed differences in the cytoskeletal architecture between these cells, and may further play an important role in the development, maintenance, and degeneration of the intervertebral disc.     

兔腰椎间盘成分抗原性差异的研究

目的 初步了解兔椎间盘不同成分的免疫源性差别,进一步提示人体内不同椎间盘细胞的抗原性差别.方法 根据生物基因相似性原理及在椎间盘不同部位髓核成分含量存在差异的基础上,采用手术切取实验动物不同区域椎间盘组织并埋植于术野中邻近的椎旁肌内,从而建立髓核糖蛋白埋植组(n=20)和纤维环胶原蛋白埋植组(n=20),并且建立埋植自体肌肉的空白对照组(n=20).分别于第1、2、4、6、8周分批处死动物取标本进行HE染色,观察新生血管化和淋巴细胞浸润;进行免疫组织化学染色,观察CD4、CD8 T淋巴细胞阳性率.结果 HE染色显示糖蛋白埋植组标本在术后1周可见淋巴细胞浸润,4周可见典型新生血管化,并且持续至第8周;胶原蛋白埋植组标本在术后4周才可见少量淋巴细胞浸润,不典型新生血管化;肌肉埋植组在各个时间点均未见典型淋巴细胞浸润和新生血管化形成.各组标本总新生血管化率差异有统计学意义.各时间点所取标本进行免疫组织化学染色,计数CD4+和CD8+T淋巴细胞,发现糖蛋白埋植组CD4+和CD8+淋巴细胞计数均高于胶原蛋白埋植组和肌肉埋植组,胶原蛋白埋植组CD4+和CD8+淋巴细胞计数高于肌肉埋植组,差异有统计学意义.结论 兔腰椎间盘成分中髓核糖蛋白相比纤维环胶原蛋白更易诱导T淋巴细胞分化为CD4+和CD8+细胞,进而推论糖蛋白的自身免疫源性强于胶原蛋白,而胶原蛋白强于肌肉组织,具有弱抗原性.     

兔腰椎间盘成分抗原性差异的研究

目的 初步了解兔椎间盘不同成分的免疫源性差别,进一步提示人体内不同椎间盘细胞的抗原性差别.方法 根据生物基因相似性原理及在椎间盘不同部位髓核成分含量存在差异的基础上,采用手术切取实验动物不同区域椎间盘组织并埋植于术野中邻近的椎旁肌内,从而建立髓核糖蛋白埋植组(n=20)和纤维环胶原蛋白埋植组(n=20),并且建立埋植自体肌肉的空白对照组(n=20).分别于第1、2、4、6、8周分批处死动物取标本进行HE染色,观察新生血管化和淋巴细胞浸润;进行免疫组织化学染色,观察CD4、CD8 T淋巴细胞阳性率.结果 HE染色显示糖蛋白埋植组标本在术后1周可见淋巴细胞浸润,4周可见典型新生血管化,并且持续至第8周;胶原蛋白埋植组标本在术后4周才可见少量淋巴细胞浸润,不典型新生血管化;肌肉埋植组在各个时间点均未见典型淋巴细胞浸润和新生血管化形成.各组标本总新生血管化率差异有统计学意义.各时间点所取标本进行免疫组织化学染色,计数CD4+和CD8+T淋巴细胞,发现糖蛋白埋植组CD4+和CD8+淋巴细胞计数均高于胶原蛋白埋植组和肌肉埋植组,胶原蛋白埋植组CD4+和CD8+淋巴细胞计数高于肌肉埋植组,差异有统计学意义.结论 兔腰椎间盘成分中髓核糖蛋白相比纤维环胶原蛋白更易诱导T淋巴细胞分化为CD4+和CD8+细胞,进而推论糖蛋白的自身免疫源性强于胶原蛋白,而胶原蛋白强于肌肉组织,具有弱抗原性.     

Effects of axial traction stress on solute transport and proteoglycan synthesis in the porcine intervertebral disc in vitro

Summary The effects of axial traction stress on intradiscal hydration, solute transport and proteoglycan synthesis were examined in 658 porcine coccygeal intervertebral discs in vitro. Measurements were performed in three tissue fractions: nucleus pulposus, inner and outer annulus fibrosus. At 0.80 MPa traction stress, the equilibrium hydration did not change in the nucleus pulposus. However, in the inner and outer annulus, the equilibrium hydration was reduced, and the change led to an increase of the effective fixed charge density. Diffusion of solute to the nucleus pulposus was significantly suppressed at 0.80 MPa traction stress. The fluid flow of the intervertebral disc tended to be suppressed during the creep recovery process after compression. The proteoglycan synthesis rate in the outer annulus was markedly suppressed by traction stress of 0.80 MPa for 4 h, but not that in the nucleus pulposus. These results suggest that a prolonged excessive axial traction stress induces a decrease in tissue hydration in the annulus fibrosus, and this may lead to an increase in the fractional volume of solid in the matrix and tissue osmotic pressure, resulting in diffusion inhibition of solute and suppression of proteoglycan synthesis. Thus, prolonged and excessive spinal traction may accelerate disc degeneration.Presented at the annual meeting of the International Society for the Study of the Lumbar Spine, May 12–16, 1991, Heidelberg, Germany     

Cyclic mechanical stretch stress increases the growth rate and collagen synthesis of nucleus pulposus cells in vitro

STUDY DESIGN: A rabbit model designed to investigate the effects of applied cyclic tensile stress on the cell division rate and the collagen synthesis in the rabbit nucleus pulposus cells in vitro. OBJECTIVE: To evaluate the effects of mechanical stress on nucleus pulposus cells, thus adding to the understanding of the adaptation of the intervertebral disc to mechanical stress. SUMMARY OF BACKGROUND DATA: Intervertebral disc cells in vivo are exposed to a multitude of physical forces during physical motion. Although it is known that in intervertebral disc disease, a common pathway of disc degeneration is mechanical stress on the nucleus pulposus or the anulus fibrosus or both, the underlying mechanism has been less well defined. METHODS: Nucleus pulposus cells were isolated from 4-week-old Japanese white rabbits. These cells were subjected to the mechanical cyclic stretch stress using a computerized, pressure-operated instrument that physically deformed the cells. The DNA synthesis rate, collagen synthesis rate, and cell cycle progression were measured. RESULTS: Cyclic tensile stretch increased the DNA synthesis rate in nucleus pulposus cells and in the population of cells in the S phase of the cell cycle during 1 to 2 days of subjugation to stress. Cyclic tensile stretch also increased collagenous protein synthesis in nucleus pulposus cells during 1 to 4 days of stress. CONCLUSIONS: Mechanical stress on nucleus pulposus cells promotes the proliferation of cells and alters the properties of intervertebral disc cells. This study may reflect the adaptation of the intervertebral disc to increased motion and stress.     

The effect of various nucleotomy techniques on biomechanical properties of the intervertebral disk]

AIM OF THE STUDY: The purpose of the present investigation was to determine the significance of the technique of nucleotomy for the biomechanical properties of the intervertebral disc. METHODS: From 29 human cadavers the lumbar-vertebral segment L2/3 was taken. The laminae were removed and the segment was tested by an mechanical testing device. Measurements were taken under physiological loads in flexion/extension, lateral bending and rotation direction. There after nucleotomy was simulated by four different procedures: perforation of the anulus with a diameter of 3 mm, rectangular excision (1 x 1 cm) of the anulus, discotomy with 3 g and 6 g nucleus pulposus. In each step the measurement was repeated. RESULTS: The size of the defect in the anulus fibrosus did not have significant influence on the stiffness of the disc. There was a significant relation between quantity of removed disc material and flexibility of the motion segment. CONCLUSION: The quantity of removed nucleus pulposus plays a crucial role in the segmental mobility whereas the size of the approach in the anulus fibrosus, f.e. endoscopic with a hole diameter of 3 mm or conventionally with a rectangular defect of 1 x 1 cm, did not influence mobility.     

Phenotypic characteristics of rabbit intervertebral disc cells. Comparison with cartilage cells from the same animals.

STUDY DESIGN: Intervertebral disc cells were extracted from the surrounding matrix, and their metabolic activities and phenotypes were studied. OBJECTIVES: To compare the metabolic activities and phenotypes of cell populations extracted from the intervertebral discs of young rabbits with those of articular and growth plate chondrocytes from the same animals. SUMMARY OF BACKGROUND DATA: The phenotype of intervertebral disc cells has been poorly studied and still is debated. METHODS: The intervertebral discs as well as articular and vertebral growth plate cartilage of rabbits were digested enzymatically. The morphology of freshly isolated cells was examined. Their contents of collagen II and X mRNAs were determined by Northern blot analysis, and their sulfation activity by 35S-sulfate incorporation as chondrocytic markers. Cells were cultured at high density or low density and grown in primary culture. The stability of their phenotype was monitored by evaluating the collagen I and II mRNA ratio. The proteoglycans newly synthesized by the cells also were quantified, and their elution profile analyzed on Sepharose 2B columns. RESULTS: The anulus fibrosus cells were morphologically undistinguishable from articular chondrocytes. The nucleus pulposus contained mainly large vacuolated cells and a few smaller cells. All freshly extracted cells expressed different levels of collagen II mRNA. Anulus fibrosus and nucleus pulposus cells contained, respectively, 22% and 8% of collagen II mRNA compared with that found in articular or growth plate chondrocytes from the same animal. Only growth plate chondrocytes expressed collagen X. When anulus fibrosus cells were incubated for 48 hours at high density, they had collagen II mRNA contents similar to those of articular and growth plate chondrocytes, but synthesized five to six times fewer sulfated proteoglycans. When seeded at low density, anulus fibrosus cells divided more slowly than articular chondrocytes and incorporated four times fewer 35S-sulfate into proteoglycans. Their collagen II mRNA content was 2.75-fold lower than that of chondrocytes, and the procollagen alpha 1II/alpha 1I mRNA ratio was 3.1 for anulus fibrosus cells and 7 for chondrocytes. No collagen X mRNA was detected. When incubated for 48 hours at high density, the nucleus pulposus giant cells had four times less collagen II mRNA content than cartilage cells but synthesized the same amounts of sulfated proteoglycans. They did not divide during 21 days in culture and still contained collagen II mRNA but no collagen X mRNA. CONCLUSIONS: Findings showed that intervertebral disc cells all express cartilage-specific matrix proteins with quantitative differences, depending on their anatomic situation. It is suggested that anulus fibrosus cells are chondrocytic cells at a different stage of differentiation than articular and growth plate chondrocytes. The phenotype of nucleus pulposus cells still is unclear. They could be chondrocytic or notochordal. A definitive answer to this important question requires differentiating markers of notochordal cells.     

Cell culture of the intervertebral disc of rats: factors influencing culture, proteoglycan, collagen, and deoxyribonucleic acid synthesis.

To establish cell culture of the nucleus pulposus and anulus fibrosus of rat intervertebral disc, the effects of culture conditions on the growth of cells and the synthesis of DNA, proteoglycan, and collagen were studied. For cell culture of the nucleus pulposus, the use of 3-week-old rats and a medium adjusted to pH 7.0 was optimal. There was almost no difference in growth between cells in Ham's F12 medium and those in Dulbecco's Modified Eagle Medium. In cells isolated from the anulus fibrosus, a medium adjusted to pH 7.0-7.6 was preferable, but irrespective of rat age. Culture cells of the nucleus pulposus were composed of large cells with vacuoles and small polygonal cells. These cells had a slight growth activity and a fair capability of proteoglycan and collagen synthesis. Culture cells of the anulus fibrosus were composed of polygonal and spindle-shape cells, and the growth was more vigorous with the potentials for proteoglycan and collagen synthesis than the nucleus cells.     

Water diffusion pathway, swelling pressure, and biomechanical properties of the intervertebral disc during compression load

The behavior of water in the intervertebral disc of pig tail and its physiologic and biomechanical properties were investigated in relation to compression load. The water content, chemical composition, and swelling pressure in the intervertebral disc were measured, and the mechanism of the generation of the swelling pressure in relation to compression load stress was studied. The swelling pressure, through regulation of the water content of the disc and the resistance of the external load, differs with the region of the intervertebral disc. In the nucleus pulposus and the inner layer of the anulus fibrosus, the swelling pressure rises in proportion to the load, but few changes occur in the outer layer of the anulus fibrosus, and the constant pressure environment is thus maintained. The tritiated water (3H2O) uptake of the disc under various loads was measured. The molar partition coefficient of tritiated water is almost equal to 1 even under a compression load, which suggests that water is freely exchangeable. The diffusion of 3H2O in the intervertebral disc was traced using two pathway models: the perianular route and the end-plate route. The diffusion of water in the unloaded disc for both uptake and washout was about 2 to 3 times larger in the perianular route than in the end-plate route. Under load, the water diffusion was inhibited in both pathways. The relation between the load and displacement revealed viscoelastic properties indicating creep and stress relaxation. Young's modulus and the stiffness increased with a rise in load speed.(ABSTRACT TRUNCATED AT 250 WORDS)     

实验性脊柱内固定后相应区域椎间盘超微结构观察

目的　观察脊柱内固定后相应区域椎间盘的超微结构变化。　方法　日本大耳白兔2 4只,随机分成实验组和对照组,每组12只。实验组骨膜下游离T1 0 ～L3棘突和关节突,克氏针制成“L”形,将钢丝横行穿过T1 1、1 2 ,L1、2 的关节突关节,并与置于T1 1 ～L3棘突两旁的克氏针系紧,对相应区域的脊柱行内固定术。对照组未行手术,仅喂养至实验完成。术后6个月,对两组动物摄X线片观察1次,随后处死动物。取两组动物的L1 椎间盘组织(髓核、纤维环内侧及纤维环外侧)行透射电镜观察,对两组T1 2 、L2 椎间盘组织分别行水平面和矢状面透射电镜及扫描电镜观察。　结果　X线片显示,实验组与对照组椎体及椎间隙差别不明显;透射电镜与扫描电镜观察,实验组椎间盘的髓核、纤维环内层细胞的结构改变较纤维环外层早;对照组的髓核、纤维环内层细胞的结构改变与纤维环外层差别不明显。在退变的椎间盘基质中,蛋白多糖颗粒和特殊结构明显减少。髓核与纤维环基质内有蛋白多糖颗粒和一种特殊结构,而特殊结构在髓核与纤维环内层的形态不一致。　结论　脊柱内固定术后6个月,实验组在异常应力环境下发生椎间盘退变。髓核、纤维环内层基质内的特殊结构分布有特殊规律,与蛋白多糖颗粒在椎间盘退变中的生物学行为密切相关。     

Temporo-spatial distribution of blood vessels in human lumbar intervertebral discs

While there is consensus in the literature that blood vessels are confined to the outer anulus fibrosus of normal adult intervertebral disc, debate continues whether there is a vascular in-growths into inner parts of the intervertebral disc during degeneration. We therefore tested the hypothesis that vascular in-growth is not a distinct feature of disc degeneration. The specific endothelial cell marker CD 31 (PECAM) was used to immunohistochemically investigate 42 paraffin-embedded complete mid-sagittal human intervertebral disc sections of various ages (0–86 years) and varying extent of histomorphological degeneration. Additionally, 20 surgical disc samples from individuals (26–69 years) were included in this study. In discs of fetal to infantile age, blood vessels perforated the cartilaginous end plate and extended into the inner and outer anulus fibrosus, but not into the nucleus pulposus. In adolescents and adults, no blood vessels were seen except for the outer zone of the anulus fibrosus adjacent to the insertion to ligaments. The cartilaginous end plate remained free of vessels, except for areas with circumscribed destruction of the end plate. In advanced disc degeneration, no vessels were observed except for those few cases with complete, scar-like disc destruction. However, some rim lesions and occasionally major clefts were surrounded by a small network of capillary blood vessels extending into deeper zones of the anulus fibrosus. A subsequent morphometric analysis, revealed slightly “deeper” blood vessel extension in juvenile/adolescent discs when compared to young, mature and senile adult individuals with significantly “deeper” extension in the posterior than anterior anulus. The analysis of the surgical specimens showed that only sparse capillary blood vessels which did not extend into the nucleus pulposus even in major disc disruption. Our results show that vascular invasion deeper than the periphery was not observed during disc degeneration, which supports the hypothesis that vascular in-growth is not a distinct feature of disc degeneration. This study was supported by a grant from the AO/ASIF Foundation Switzerland (00-B72) and a grant from the AO Spine (SRN 02/103).     

Mechanobiology of the intervertebral disc and relevance to disc degeneration

Mechanical loading of the intervertebral disc may contribute to disc degeneration by initiating degeneration or by regulating cell-mediated remodeling events that occur in response to the mechanical stimuli of daily activity. This article is a review of the current knowledge of the role of mechanical stimuli in regulating intervertebral disc cellular responses to loading and the cellular changes that occur with degeneration. Intervertebral disc cells exhibit diverse biologic responses to mechanical stimuli, depending on the loading type, magnitude, duration, and anatomic zone of cell origin. The innermost cells respond to low-to-moderate magnitudes of static compression, osmotic pressure, or hydrostatic pressure with increases in anabolic cell responses. Higher magnitudes of loading may give rise to catabolic responses marked by elevated protease gene or protein expression or activity. The key regulators of these mechanobiologic responses for intervertebral disc cells will be the micromechanical stimuli experienced at the cellular level, which are predicted to differ from that measured for the extracellular matrix. Large hydrostatic pressures, but little volume change, are predicted to occur for cells of the nucleus pulposus during compression, while the highly oriented cells of the anulus fibrosus may experience deformations in tension or compression during matrix deformations. In general, the pattern of biologic response to applied loads suggests that the cells of the nucleus pulposus and inner portion of the anulus fibrosus experience comparable micromechanical stimuli in situ and may respond more similarly than cells of the outer portion of the anulus fibrosus. Changes in these features with degeneration are critically understudied, particularly degeneration-associated changes in cell-level mechanical stimuli and the associated mechanobiology. Little is known of the mechanisms that regulate cellular responses to intervertebral mechanobiology, nor is much known with regard to the precise mechanical stimuli experienced by cells during loading. Mechanical factors appear to regulate responses of the intervertebral disc cells through mechanisms involving intracellular Ca(2+) transients and cytoskeletal remodeling that may regulate downstream effects such as gene expression and posttranslational biosynthesis. Future studies should address the broader biologic responses to mechanical stimuli in intervertebral disc mechanobiology, the involved signaling mechanisms, and the apparently important interactions among mechanical factors, genetic factors, cytokines, and inflammatory mediators that may be critical in the regulation of intervertebral disc degeneration.     

Magnetic resonance imaging measurement of relaxation and water diffusion in the human lumbar intervertebral disc under compression in vitro.

STUDY DESIGN: Twelve lumbar intervertebral disc specimens were imaged with magnetic resonance imaging to estimate relaxation constants, T1 and T2, and tissue water diffusion, before and after applying compression. OBJECTIVES: The objectives of the study were to measure T1, T2, and water diffusion for differences with loading state, region of the disc (anulus fibrosus or nucleus pulposus), and grade of degeneration. SUMMARY OF BACKGROUND DATA: Magnetic resonance imaging can be used qualitatively to estimate water content and degeneration of the intervertebral disc. Beyond structural information of images, the relaxation times T1 and T2 may contain information on the changes occurring with degeneration. A modified spin-echo sequence can be used to estimate tissue water diffusion in cartilage and disc specimens with the ability to measure anisotropy. METHODS: Specimens were imaged in a 1.5-Tesla clinical scanner. T1, T2, and water diffusion were estimated from midsagittal images. Magnetic resonance imaging parameters were calculated before and after axial loading. The measured T1, T2, and D (diffusion coefficient) were compared before and after compression, and for the diffusion data, also by direction to consider anisotropy. RESULTS: For the T1 data, a significant difference was found by region, nucleus > anulus, and loading state, loaded > unloaded. For the T2 values, there was a significant difference by region, nucleus > anulus, and Thompson grade. For diffusion, significant differences were found by region, nucleus > anulus, Thompson grade, direction of diffusion, and state of compression, loaded > unloaded. CONCLUSIONS: This study demonstrated that magnetic resonance imaging can be used to measure significant changes in T1, T2, or diffusion in intervertebral disc specimens by region, loading condition, or Thompson grade.     

Automated percutaneous lumbar discectomy with and without chymopapain pretreatment versus non-automated,discoscopy-monitored percutaneous lumbar discectomy

Summary Percutaneous lumbar discectomy has gained growing interest during recent years as an alternative to open surgery for protrusions and non-sequestrated subligamentous intervertebral disc herniations. As a less invasive method it competes with chemonucleolysis. At least two modifications are known to date: automated percutaneous lumbar discectomy (APLD) with a 2-mm suction probe and non-automated, discoscopy-monitored percutaneous lumbar discectomy with a suction rongeur and a motor-driven shaver (NAPLD). In this study these two methods are compared for the amount of material extracted, dependence upon the degree of degeneration of the disc and intrinsic technical problems, using 20 human cadaver lumbar specimens for experimental testing. Total nucleotomy was not possible with either method. APLD yielded significantly less material and proved to be less effective in severely degenerated intervertebral discs than the NAPLD procedure, as preexisting gaps within the degenerated nucleus pulposus allowed the tissue to shift away from the tip of the 2-mm probe and facilitated displacement of the probe within the anulus fibrosus. By contrast, the rongeur, which first cuts the material to be removed and then carries it away by suction, was much more effective. Further data to support the advantages of non-automated percutaneous nucleotomy are discussed. Pretreatment of the disc with chymopapain did not result in a higher yield of nucleus material when combined with APLD.     

The effect of age on inflammatory responses and nerve root injuries after lumbar disc herniation: an experimental study in a canine model

STUDY DESIGN: An experimental investigation on the effect of age on pathologic events surrounding the herniated disc and at the adjacent nerve root. OBJECTIVES: To investigate the role of age on the inflammatory responses and nerve root damage surrounding a sequestered lumbar disc fragment using a dog model. SUMMARY OF BACKGROUND DATA: Lumbar disc herniation is manifested in patients by variable clinical findings, natural history, and resorption phenomena in which the variability is particularly noted among patients with different ages. There are no previous reports on the effect of age on pathologic events induced by the herniated disc. METHODS: Six beagle dogs, including two animals of each age group of 6, 12, and 24 months (human equivalent ages of 10, 15, and 24 years), were used in this study. The dogs underwent L4-L5, L5-L6, and L6-L7 laminotomy and discectomy under general anesthesia. An autologous intervertebral disc from the tail was divided into anulus fibrosus and nucleus pulposus fragments. The anulus fibrosus and nucleus pulposus fragments were placed in the anterolateral epidural space of L5-L6 and L6-L7, respectively. The L4-L5 discectomy site served as a control. Dogs were killed at 12 weeks after surgery. The lumbar spine was removed en bloc, and histologic sections were prepared consecutively and examined. RESULTS: In the nucleus pulposus group at L6-L7, neovascularity, and intensive infiltration of lymphocytes, macrophages, and fibroblasts were observed surrounding the nucleus pulposus fragment in the 24-month-old group only. Degenerative changes of the nerve root fibers were observed in the 24-month-old group only. In the control and anulus fibrosus groups at L4-L5 and L5-L6, there were no marked inflammatory reactions in all age groups. The nerve root fibers around the anulus fibrosus were normal in all age groups. CONCLUSIONS: There is an effect of age on the inflammatory response and nerve root injury caused by the herniated disc. The apparent neuroprotective mechanism in the young animal, and the apparent inflammatory and resorption changes of the nucleus pulposus fragment in the older animal are quite intriguing.     

Effects of anulus fibrosus and experimentally degenerated nucleus pulposus on nerve root conduction velocity: relevance of previous experimental investigations using normal nucleus pulposus

STUDY DESIGN: Nerve conduction velocity was measured in the pig cauda equina after local application of anulus fibrosus or in vitro/postmortem degenerated nucleus pulposus from the same pig. OBJECTIVES: To analyze the effects of anulus fibrosus and degenerated nucleus pulposus on nerve conduction velocity. SUMMARY OF BACKGROUND DATA: Previous studies on nucleus pulposus-induced effects on nerve roots have used normal, nondegenerated nucleus pulposus. Because both anulus fibrosus and degenerated nucleus pulposus are commonly seen in the clinical situation of disc herniation, the value of the previous work could be questioned. METHODS: Anulus fibrosus and nucleus pulposus were harvested using a retroperitoneal approach. The nucleus pulposus was degenerated artificially either by addition of sodium lactate with HCl added to form a pH of either 6.0 or 3.5 (in vitro degeneration), or by storing the nucleus pulposus at 4 C until a pH of 6.0 (postmortem degeneration) was reached. After epidural application, the nerve conduction velocity was determined at 7 days (anulus fibrosus) or 3 days (degenerated nucleus pulposus). RESULTS: Application of anulus fibrosus did not induce any reduction of nerve conduction velocity. In vitro and postmortem degenerated nucleus pulposus induced a reduction of nerve conduction velocity similar to that of normal nucleus pulposus. CONCLUSIONS: Although only nerve function and not pain was assessed, it seems likely that previous experiments using normal nucleus pulposus may be relevant for evaluating the pathophysiologic mechanisms behind the nucleus pulposus-induced nerve root injury, also in a clinical perspective.     

Histology of intervertebral disc protrusion: an experimental study using an aged rat model

STUDY DESIGN: In vitro experimental intervertebral disc ruptures of aged rats were examined histologically. OBJECTIVES: To clarify the mechanism of intervertebral disc herniations by microscopic investigation of ruptured discs. SUMMARY OF BACKGROUND DATA: Clinically, disc herniations have been classified into two types: extrusion and protrusion. However, the pathogenesis of protrusion type herniations has not yet been demonstrated by any studies. To clarify this issue, it is essential to establish an appropriate model producing disc herniations, and to examine the sequential changes in the structure of herniated discs. METHODS: Lumbar discs of 2-year-old rats were examined histologically and compared with human lumbar discs. To examine structural changes in discs subjected to repetitive motion stress, 400 repetitions of a sequence of flexion (30 degrees ) and axial rotation (6 degrees ) were applied in vitro to the lumbar discs of the animals. RESULTS: The microstructure of normal lumbar discs in aged rats was similar in many ways to the human lumbar discs in a 20- to 40-year-old adult. Of 10 discs subjected to repetitive stress, 4 were ruptured at the junction between the posterior anulus fibrosus and the sacral cartilage endplate. One had an extruded nucleus pulposus, and three had a protruded anulus fibrosus, which displayed disorganized structure containing widened and flaccid lamellae. CONCLUSIONS: The results from this study indicate that disc protrusion can be caused by disorganization of the ruptured annular lamellae, not by focal compression of the nucleus pulposus.