Akihito Minamide

Adjacent segment motion after a simulated lumbar fusion in different sagittal alignments: a biomechanical analysis.

Study Design. An in vitro biomechanical study of adjacent segment motion (at L3–L4 and L5–S1) after a simulated lumbar interbody fusion of L4–L5 in different sagittal alignments was carried out. Objectives. To test the hypothesis that an L4–L5 fixation in different sagittal alignments causes increased angular motion at the adjacent levels (L3–L4 and L5–S1) in comparison with the intact spine. Summary of Background Data. Clinical experience has suggested that lumbar fusion in a nonanatomic sagittal alignment can increase degeneration of the adjacent levels. It has been hypothesized that this is the result of increased motion at these levels; however, to the authors’ knowledge no mechanical studies have demonstrated this. Methods. Eight fresh human cadaveric lumbar spines (L3–S1) were biomechanically tested. Total angular motion at L3–L4 and L5–S1 under flexion-extension load conditions (7-Nm flexion and 7-Nm extension) was measured. Each specimen was tested intact, and then again after each of three different sagittal fixation angles (at L4–L5): (1) in situ (21°lordosis), (2) hyperlordotic (31°lordosis), and (3) hypolordotic (7°lordosis). The simulated anterior/posterior fusion was performed at L4–L5 with pedicle screws posteriorly, vertebral body screws anteriorly, and an interbody dowel. Results. The averaged values for flexion-extension motion at L3–L4 were as follows: intact specimen 2.0°, in situ fixation 4.0°, hyperlordotic fixation 1.7°, hypolordotic fixation 6.5°. The averaged values for flexion-extension motions at L5–S1 were as follows: intact specimen 2.3°, in situ fixation 2.6°, hyperlordotic fixation 3.6°, hypolordotic fixation 2.9°. Conclusion. Hypolordotic alignment at L4–L5 caused the greatest amount of flexion-extension motion at L3–L4, and the differences were statistically significant in comparison with intact specimen, in situ fixation, and hyperlordotic fixation. Hyperlordotic alignment at L4–L5 caused the greatest amount of flexion-extension motion at L5–S1, and the difference was statistically significant in comparison with intact specimen but not in situ fixation or hypolordotic fixation.

Delivery of Recombinant Human Bone Morphogenetic Protein-2 Using a Compression-Resistant Matrix in Posterolateral Spine Fusion in the Rabbit and in the Non-Human Primate

Study Design. A rabbit and rhesus monkey model of posterolateral intertransverse process spine arthrodesis was used. Objective. To test two new soft tissue compression resistant ceramic/collagen sponge carriers for recombinant human bone morphogenetic protein-2. Summary of Background Data. After determining that a plain collagen sponge was too compressible for large animals in a posterolateral fusion application, the authors demonstrated good bone induction using biphasic ceramic phosphate granules (60% hydroxyapatite/40% tricalcium phosphate) as the carrier matrix for recombinant human bone morphogenetic protein 2 in rhesus monkeys. A limitation of 60:40 biphasic ceramic phosphate was its slow resorption time caused by the high hydroxyapatite content, making radiographic detection of new bone formation very difficult. Methods. Adult New Zealand white rabbits (n = 14) underwent posterolateral spine arthrodesis at L5–L6 using 5:95 biphasic ceramic phosphate (5% hydroxyapatite/95% tricalcium phosphate) impregnated Type I collagen sponges (17 × 35 × 2.5 mm, two per side) loaded with 0.86 mg recombinant human bone morphogenetic protein 2. Additional rabbits (n = 14) received 60:40 hydroxyapatite-tricalcium phosphate granules as the carrier for bone morphogenetic protein 2. Adult rhesus monkeys (n = 6) underwent posterolateral arthrodesis at L4–L5 with ceramic/collagen sponge carrier loaded with 9 mg recombinant human bone morphogenetic protein 2 per side. Two monkeys received ceramic/collagen sponges containing 15:85 biphasic ceramic phosphate (15% hydroxyapatite/85% tricalcium phosphate) with two pieces per side; two received sponges containing 5:95 biphasic ceramic phosphate with two pieces per side, and two received sponges containing 5:95 biphasic ceramic phosphate with four pieces per side. The rabbits were killed after 5 weeks and the monkeys after 24 weeks; the spines were evaluated by manual palpation, radiographs, tensile mechanical testing (rabbits only), and histology. Results. The recombinant human bone morphogenetic protein 2 delivered in the 5:95 biphasic ceramic phosphate/collagen sponge achieved fusion in 100% of rabbits and had improved handling properties compared with the biphasic ceramic phosphate granules. Biomechanical results with 5:95 biphasic ceramic phosphate/collagen carrier were comparable to those obtained with the 60:40 biphasic ceramic phosphate granules and superior to those of autogenous bone graft (P < 0.05). The recombinant human bone morphogenetic protein 2 delivered in the 15:85 or the 5:95 biphasic ceramic phosphate/collagen sponge carrier (two pieces per side) induced fusion in nonhuman primates with normal bone histology, less residual ceramic, and more bone in the center of the carrier matrix in comparison with BCO granules alone. The 15:85 biphasic ceramic phosphate/collagen sponge resulted in fusion mass sizes closer to the original size of the matrix implanted than did the 5:95 biphasic ceramic phosphate/collagen sponge, which was considered a desirable feature. The monkeys with 9 mg recombinant human bone morphogenetic protein 2 spread over four sponges per side instead of two had half the effective recombinant human bone morphogenetic protein 2 concentration per sponge and inferior results. Conclusions. The new compression-resistant biphasic ceramic phosphate/collagen sponge matrices were biologically compatible with recombinant human bone morphogenetic protein 2 bone formation, resulted in biomechanically stiffer fusion masses than autograft, better space maintenance than plain collagen sponges, and improved handling and radiographic resorption properties over the ceramic carriers previously tested.

Lower Dose of rhBMP-2 Achieves Spine Fusion When Combined With an Osteoconductive Bulking Agent in Non-human Primates

Study Design. A non-human primate lumbar intertransverse process arthrodesis model was used to evaluate the effectiveness of different formulations of recombinant human bone morphogenetic protein-2 (rhBMP-2) to induce consistent bone formation. Objective. To determine if the combination of rhBMP-2/absorbable collagen sponge (ACS) wrapped around a bulking agent, consisting of a biphasic calcium phosphate/collagen composite, could achieve posterolateral spine fusion with a dose of rhBMP-2 (3.0 mg/side) that previously failed to induce posterolateral fusion in rhesus monkeys with other carriers. Summary of Background Data. Successful bone induction in both human and non-human primates has required higher concentrations of BMP than were required in lower order models. The Food and Drug Administration approved concentration of rhBMP-2 for interbody fusion (1.5 mg/mL) when delivered on the ACS alone without a bulking agent (doses 3–9 mg/side) has failed to induce clinically relevant amounts of bone formation in a posterolateral spine fusion model in rhesus monkeys. Previously, a higher concentration of 2.0 mg/mL of rhBMP-2 delivered on stacked sheets of a biphasic calcium phosphate ceramic/collagen compression resistant matrix (CRM) was required to achieve fusion in the rhesus monkey and was the basis for this study (doses of 6–12 mg/side). Methods. Nine skeletally mature, rhesus macaque monkeys underwent single level posterolateral arthrodesis at L4–L5. Two different rhBMP-2 doses were evaluated in 3 delivery configurations. The first 3 monkeys received 10 mg/side (2.5 mL at 4.0 mg/mL) of rhBMP-2 loadeddirectly onto a CRM carrier (15% hydroxyapatite/85%&bgr;-tricalcium phosphate ceramic/collagen matrix), resulting in a final concentration of 2.0 mg/mL. The second 3monkeys received 3 mg/side (2.0 mL at 1.5 mg/mL) of rhBMP-2 loaded directly on the CRM carrier, resulting in a 0.6 mg/mL final concentration. Three additional monkeys also received the 3 mg/side (2.0 mL at 1.5 mg/mL) of rhBMP-2 delivered on an ACS, which was then wrapped around the dry CRM matrix used as a bulking agent, yielding a 1.5 mg/mL final concentration of rhBMP-2 on the sponge wrapped around the bulking agent. The monkeys were euthanized at 24 weeks after surgery. Manual palpation, plain radiographs, computerized tomography, and nondecalcified histology were used to evaluate fusion in a blinded fashion. Results. The 3 monkeys with 10 mg rhBMP-2 placed directly on the CRM carrier (2.0 mg/mL final concentration) achieved solid fusion. The 3 monkeys that underwent fusion with 3 mg of rhBMP-2 placed directly on the CRM carrier (0.6 mg/mL final concentration) failed to achieve fusion. In contrast, the 3 monkeys that underwent fusion with the same 3 mg dose of rhBMP-2 dispensed only on an ACS that was wrapped around the CRM achieved solid bilateral fusion. Conclusions. This study shows the importance of carrier optimization and final implant protein concentration for the successful delivery of rhBMP-2. By combining the properties of the ACS with the CRM, the required dosage of rhBMP-2 was diminished by more than 3-fold in the non-human primate model. This finding suggests that the currently available concentration of rhBMP-2 (1.5 mg/mL) could be successful for achieving posterolateral spine fusion when combined with an osteoconductive bulking agent that can support the induced new bone formation.

The use of cultured bone marrow cells in type I collagen gel and porous hydroxyapatite for posterolateral lumbar spine fusion.

Study Design. Posterolateral lumbar transverse process fusion was completed using the cultured bone marrow cells in type I collagen gel and porous hydroxyapatite. Objective. To compare the efficacy of cultured bone marrow cells with that of bone morphogenetic protein (BMP) as a graft alternative to autologous bone for posterolateral spine fusion. Summary of Background Data. The clinical application of BMP for spinal fusion may be limited by high dose and cost. Recently, mesenchymal stem cells have been studied in various fields because of their capability to differentiate into various cells, including those in the osteogenic lineage. Methods. Thirty adult rabbits were used. Each underwent single-level, bilateral, posterolateral intertransverse process fusions at L4–L5. The animals were divided into 4 groups, each according to the material implanted: (1) autologous bone (autograft, n = 9); (2) porous hydroxyapatite (HA) particles and type I collagen sheet with 100 &mgr;g rhBMP-2 (BMP-HA, n = 7); (3) bone marrow cells (1 × 106cells/mL, low-marrow-HA, n = 7); and (4) bone marrow cells (1 × 108cells/mL, high-marrow-HA, n = 7). Before implantation for groups 3 and 4, fresh bone marrow cells from the iliac crest of each animal were cultured in a standard medium for 2 weeks. For one additional week, the marrow cells were cultured in 10−8M dexamethasone, type I collagen gel, and HA. Animals were euthanized 6 weeks after surgery. Spinal fusions were evaluated by radiograph, manual palpation, and histology. Results. The fusion rates were 4 of 7 in the autograft group, 7 of 7 in the BMP-HA group, 0 of 7 in the low-marrow-HA group, and 5 of 7 in the high-marrow-HA group. The histology in the BMP-HA and high-marrow-HA groups showed that grafted HA fragments were connected with mature new bone. The pores of HA fragments were filled up with bone matrix. In the low-marrow-HA group, fibrous tissue was predominant in the grafted fragments. Conclusions. This study shows that the cultured bone marrow cells can act as a substitute for autograft or BMP in spine fusion. The current formulation may yield improved fusion success and better quality of fusion bone as compared to autograft.

Prevalence and distribution of intervertebral disc degeneration over the entire spine in a population-based cohort: the Wakayama Spine Study

OBJECTIVES The purposes of this study were to investigate the prevalence and distribution of intervertebral disc degeneration (DD) over the entire spine using magnetic resonance imaging (MRI), and to examine the factors and symptoms potentially associated with DD. DESIGN This study included 975 participants (324 men, mean age of 67.2 years; 651 women, mean age of 66.0 years) with an age range of 21-97 years in the Wakayama Spine Study. DD on MRI was classified into Pfirrmanns system (grades 4 and 5 indicating DD). We assessed the prevalence of DD at each level in the cervical, thoracic, and lumbar regions and the entire spine, and examined DD-associated factors and symptoms. RESULTS The prevalence of DD over the entire spine was 71% in men and 77% in women aged <50 years, and >90% in both men and women aged >50 years. The prevalence of an intervertebral space with DD was highest at C5/6 (men: 51.5%, women: 46%), T6/7 (men: 32.4%, women: 37.7%), and L4/5 (men: 69.1%, women: 75.8%). Age and obesity were associated with the presence of DD in all regions. Low back pain was associated with the presence of DD in the lumbar region. CONCLUSION The current study established the baseline data of DD over the entire spine in a large population of elderly individuals. These data provide the foundation for elucidating the causes and mechanisms of DD.

Effects of steroid and lipopolysaccharide on spontaneous resorption of herniated intervertebral discs: An experimental study in the rabbit

Study Design. Histologic examination was performed on autografted intervertebral disc materials of rabbit models, which were partially incised through a retroperitoneal approach at L1‐L2 and grafted within the posterior epidural space at L4. Objective. To evaluate whether the resorption process of the herniated intervertebral disc is influenced and controlled by treatments with medications. Summary of Background Data. Regarding resorption of herniated intervertebral discs, recent studies of magnetic resonance images and histologic investigations of surgically resected specimens in lumber disc herniation patients have been reported. It has been shown that inflammatory factors may play an important role in the mechanism of resorption of the herniated intervertebral disc. However, little is known about the origin of newly formed vessels and inflammatory cells detected in herniated disc specimens from patients. In this study, The resorption process of disc material grafted into the epidural space was observed in a rabbit model. Methods. Thirty‐six adult rabbits were used. The L1‐L2 intervertebral disc was partially incised through a retroperitoneal approach. The harvested disc material, which contains the nucleus pulposus and the anulus fibrosus were placed into the posterior epidural space at L4 of the same rabbit. The animals were divided into control, and steroid groups. The control group received no treatment after surgery. In the lipopolysaccharide group, rabbits were injected 1 mg/kg into the peritoneum immediately and at 7 days after surgery. In the steroid group, rabbits were injected with 1 mg/kg beta‐methasone into the epidural space daily from 1 to 7 days after surgery. Rabbits of each group were killed for histologic examination at 1, 2, 4, and 8 weeks after surgery. Results. At 1 and 2 weeks after surgery, inflammatory cells and newly formed vessels were more frequently observed in the lipopolysaccharide group than in the control and steroid groups. At 4 weeks after surgery, derangement and loosening of collagen fibers were also observed in the lipopolysaccharide group. At 8 weeks after surgery, fragmentation and partial disappearance of matrix were observed in the control and lipopolysaccharide groups. Most of the intervertebral discs were replaced by fibrous tissues in the lipopolysaccharide group. However, the matrix of the intervertebral disc almost remained. Conclusions. Autologous intervertebral disc material grafted into the epidural space was penetrated by newly formed vessels produced from the epidural fat tissue and resolved as the result of inflammatory reaction. Lipopolysaccharide accelerated the replacement of grafted intervertebral disc by fibrous tissue, which suggests the resorption of the disc in the epidural space of the rabbit, whereas high‐dose steroid suppressed the replacement.

Mechanism of bone formation with gene transfer of the cDNA encoding for the intracellular protein LMP-1.

Background: LIM mineralization protein-1 (LMP-1), an intracellular protein, is thought to induce secretion of soluble factors that convey its osteoinductive activity. Although evidence suggests that LMP-1 may be a critical regulator of osteoblast differentiation in vitro and in vivo, little is known about its mechanism of action. The purpose of the present study was to identify candidates for the induced secreted factors and to describe the time sequence of histological changes during bone formation induced by LMP-1.Methods: Human lung carcinoma (A549) cells were used to determine if LMP-1 overexpression would induce expression of bone morphogenetic proteins (BMPs) in vitro. Cultured A549 cells were infected with recombinant replication-deficient human type-5 adenovirus containing the LMP-1 or LacZ cDNA. Cells were subjected to immunohistochemical analysis after forty-eight hours. Finally, sixteen athymic rats received subcutaneous implants consisting of collagen disks loaded with human buffy-coat cells that were infected with one of the above two viruses. Rats were killed at intervals, and explants were studied with histological and immunohistochemical analyses.Results: In vitro experiments with A549 cells showed that AdLMP-1-infected cells express elevated levels of BMP-2, BMP-4, BMP-6, BMP-7, and TGF-&bgr;1 (transforming growth factor-beta 1) protein. Human buffy-coat cells infected with AdLMP-1 also demonstrated increased levels of BMP-4 and BMP-7 protein seventy-two hours after ectopic implantation in athymic rats, confirming the in vitro hypothesis.Conclusions: The osteoinductive properties of LMP-1 involve synthesis of several BMPs and the recruitment of host cells that differentiate and participate in direct membranous bone formation.Clinical Relevance: Ex vivo gene therapy with the LMP-1 cDNA-induced secretion of multiple BMPs may provide an alternative to implantation of large doses of a single BMP to induce new bone formation.

Prevalence of symptomatic lumbar spinal stenosis and its association with physical performance in a population-based cohort in Japan: the Wakayama Spine Study

OBJECTIVE The purpose of this study was to investigate the prevalence of symptomatic lumbar spinal stenosis (LSS) and to clarify the association between symptomatic LSS and physical performance using magnetic resonance imaging (MRI) in a population-based cohort. DESIGN This cross-sectional study was performed as a part of the research on osteoarthritis/osteoporosis against disability (ROAD) in Japan and 1,009 subjects (335 men, 674 women, mean age 66.3 years, age range 21-97 years) were analyzed. An experienced orthopedic surgeon obtained the medical history and performed the physical testing for all participants. Symptomatic LSS diagnostic criteria required the presence of both symptoms and radiographic LSS findings. A 6-m walking time, chair standing time, and one-leg standing time were obtained from all participants. RESULTS The prevalence of symptomatic LSS was 9.3% (95% confidence interval [CI]: 7.7-11.3) overall, 10.1% (CI: 7.4-13.8) in men and 8.9% (CI: 7.0-11.3) in women. There was a difference in the prevalence with increasing age by gender. The LSS prevalence showed little difference with age greater than 70 years for men, but the LSS prevalence for women was higher with increasing age. Among physical performance measures, 6-m walking time at a maximal pace was significantly associated with symptomatic LSS (P = 0.03). CONCLUSION The prevalence of symptomatic LSS was approximately 10% in a cohort resembling the general Japanese population. A 6-m walking time at a maximal pace was a more sensitive index than walking at a usual pace in assessing decreased physical performance associated with symptomatic LSS.

Effects of basic fibroblast growth factor on spontaneous resorption of herniated intervertebral discs. An experimental study in the rabbit.

STUDY DESIGN Histologic examination was performed on the autologous intervertebral disc material that was removed from the intervertebral space at L1-L2 and then relocated to the L4 posterior epidural space after the addition of basic fibroblast growth factor (bFGF) in a rabbit. OBJECTIVES To evaluate whether basic fibroblast growth factor influences the resorption process of the herniated intervertebral disc through the promotion of angiogenesis and chemotaxis. SUMMARY OF BACKGROUND DATA It has been reported that newly formed vessels, inflammatory cells, and their products may play an important role in the spontaneous resorption process of herniated intervertebral discs. In a rabbit model that mimics the sequestration type of intervertebral disc herniation, it has been reported that the autologous intervertebral disc material that relocated into the epidural space was penetrated by newly formed vessels originating from the epidural fat tissue. Therefore, it is possible that promotion of angiogenesis may influence the resorption of herniated intervertebral discs. Basic fibroblast growth factor is well known as an angiogenesis stimulation factor in vivo. METHODS Thirty-six adult rabbits were divided into three groups. The L1-L2 intervertebral disc was partially incised through a retroperitoneal approach in each rabbit. The harvested disc material, which contained nucleus pulposus and anulus fibrosus, was immersed in one of three kinds of solution before relocation into the posterior epidural space at L4. In the control group, the harvested intervertebral disc was immersed in physiologic saline for 2 hours before relocation. In the group receiving 5 micrograms bFGF, the disc was immersed in 5 micrograms/mL bFGF for 2 hours before the relocation. In the group receiving 20 micrograms bFGF, the disc was immersed in 20 micrograms/mL bFGF for 2 hours before the relocation. Rabbits of each group were killed for histologic examination 1, 2, 4, and 8 weeks after surgery. RESULTS In the bFGF-treated groups, newly formed vessels were observed to be in more numerous than those in the control group, 1 and 2 weeks after surgery. The number of inflammatory cells, including macrophages, lymphocytes, and fibroblasts, also increased in the bFGF-treated groups. The period from the surgery to the degradation of the intervertebral disc in the bFGF-treated groups was shorter than that in the control group, although the resorption process of the relocated discs was also observed in the control group. The size of relocated intervertebral discs in the bFGF-treated groups decreased at a higher rate than in the control group as time progressed. The rate of decrease in the size of discs in the group treated with 20 micrograms bFGF was more than that in the group treated with 5 micrograms. CONCLUSIONS Epidural injection of bFGF facilitated the resorption of the intervertebral disc relocated to the epidural space.

Effect of tail suspension (or simulated weightlessness) on the lumbar intervertebral disc: study of proteoglycans and collagen.

Study Design. An experiment to measure the proteoglycan and collagen content of the lumbar intervertebral discs of rats that had been tail-suspended for up to 4 weeks. Objectives. To determine the effect of tensile force (or simulated weightlessness) on the intervertebral disc. Summary of Background Data. During space flight the intervertebral disc experiences low compressive force (because of so-called “weightlessness”), which, in turn, produces, among other things, low hydrostatic pressure acting on the disc cells. Although disc cells respond (in vitro) to changes in hydrostatic pressure, it is unclear what effect low levels of hydrostatic pressure have in vivo and whether they lead to a degenerative catabolic process. The rat tail-suspension model is appropriate for studying the effects of tensile force on the disc. The disc (especially the anulus) is subjected to tension during various body movements (e.g., bending stretches the posterior anulus, and twisting tensions the whole anulus). Methods. Thirty-two Sprague–Dawley rats were tail-suspended for either 2 weeks (16 rats) or 4 weeks (16 rats). Sixteen other rats were left unsuspended for 4 weeks; these were used as controls. At the end of 2 or 4 weeks, as appropriate, the rats were killed and their lumbar spines were removed. In each rat the six lumbar discs were bisected and the discs (anulus and nucleus together) were carefully removed. The six lumbar discs from one rat were pooled with the six lumbar discs of a second matching rat (i.e., from the same group) to give one sample. The disc samples were then assessed using enzyme-linked immunosorbent assays. Results. There was a 35% statistically significant decrease in proteoglycan content going from the control group down to the 4-week group, but no significant differences between the control group and the 2-week group or between the 2-week group and the 4-week group. There were no statistically significant differences between the three groups for collagen I or collagen II. Conclusions. These findings clearly establish a link between decreased proteoglycan content and tension on the disc, as modeled by the tail-suspended rat.