淫羊藿调节BMP-7预防糖皮质激素致大鼠骨损害的 研究

目的观察醋酸泼尼松和淫羊藿对大鼠胫骨近端上段骨组织形态计量学参数、股骨组织形态学和股骨骨形态发生蛋白-7(BMP-7)表达的影响,探讨淫羊藿预防糖皮质激素致骨损害的BMP-7机制。方法 3月龄SD大鼠24只,随机分为正常对照组、糖皮质激素模型组和淫羊藿防治组,灌胃给药90 d。测量胫骨近端上段骨组织形态计量学参数,苏木素-伊红染色法观察股骨组织形态学,免疫组化法检测股骨BMP-7的表达。结果醋酸泼尼松可导致大鼠胫骨近端上段标记周长百分数(%L.Pm)和骨形成率(BFR/TV、BFR/BV和BFR/BS)减少(P0.05);股骨骨小梁钙化减少,成软骨化,纤维组织增生,骨小梁变窄、数量减少;股骨BMP-7表达减少(P0.05)。与泼尼松组比较,淫羊藿组胫骨近端上段骨小梁面积百分数(%Tb.Ar)和骨形成率(BFR/BS)增加(P0.05),股骨无明显病理损害,股骨BMP-7表达增多(P0.05)。结论淫羊藿能通过上调骨组织BMP-7的表达,预防糖皮质激素所致的骨损害。     

The effects of recombinant human bone morphogenetic protein-2, recombinant human bone morphogenetic protein-12, and adenoviral bone morphogenetic protein-12 on matrix synthesis in human annulus fibrosis and nucleus pulposus cells

BACKGROUND CONTEXT: Bone morphogenetic proteins (BMPs) are potential therapeutic factors for degenerative discs, and BMP-12 does not have the osteogenic potential of BMP-2, making it better suited for intradiscal injection. However, no reports have compared the actions of BMP-2 and -12 on human annulus fibrosus (AF) and nucleus pulposus (NP) cells nor evaluated adenoviral-mediated gene therapy in human AF cells. PURPOSE: To evaluate and compare the effects of recombinant human (rh) BMP-2, rhBMP-12, and adenoviral BMP-12 (Ad-BMP-12) on nucleus pulposus and annulus fibrosis cell matrix protein synthesis. STUDY DESIGN: In vitro study using rhBMP-2 and -12 and adenoviral BMP-12 with human intervertebral disc (IVD) cells. METHODS: Human NP and AF IVD cells were isolated, maintained in monolayer, and incubated with BMP-2 or -12 for 2 days. AF and NP cells were transduced with Ad-BMP-12, pellets formed, and incubated for 6 days. Growth factor-treated cells were labelled with either 35-S or 3H-proline to assay matrix protein synthesis. RESULTS: rhBMP-2 increased NP proteoglycan, collagen, and noncollagen protein synthesis to 355%, 388%, and 234% of control. RhBMP-12 increased the same NP matrix proteins' synthesis to 140%, 143%, and 160% of control. Effects on AF matrix protein synthesis were minimal. Ad-BMP-12 significantly increased matrix protein synthesis and DNA content of AF and NP cells in pellet culture. NP synthesis of all matrix proteins and AF synthesis of proteoglycans was increased when the data were normalized to pellet DNA. AF synthesis of noncollagen protein and collagen was not modulated by Ad-BMP-12 if the data are normalized to pellet DNA content. CONCLUSIONS: Both rhBMP-2 and -12 increase human NP cell matrix protein synthesis while having minimal effects on AF cells. However, Ad-BMP-12 did increase matrix protein synthesis in both NP and AF cells, making it a potential therapy for enhancing matrix production in the IVD. These responses plus the proliferative action of Ad-BMP-12 seen in the current studies, and the lack of an osteogenic action noted in other studies justifies future studies to determine if gene therapy with BMP-12 could provide protective and/or reparative actions in degenerating discs.     

重组BMP-4对兔骨髓基质干细胞生物学行为的影响

目的应用原核细胞基因工程方法生产出有活性的骨形态发生蛋白-4(bonemorphoge-neticprotein-4,BMP-4),观察其对骨髓基质干细胞生物学行为的影响。方法利用RT-PCR技术,从成熟的人胎盘组织中扩增出长0.34kb编码人BMP-4成熟肽的基因序列,装入表达载体pET-22b( ),转化大肠杆菌BL-21菌株,并诱导目的蛋白表达,SDS-PAGE检测表达蛋白。获得的蛋白制品经小鼠异位成骨测活证实后,诱导培养的兔骨髓基质干细胞,观测细胞形态变化、碱性磷酸酶和骨钙素的含量。结果大肠杆菌中目的蛋白表达量达菌体蛋白的15%,该蛋白可诱导骨髓基质干细胞向成骨细胞分化,形成钙结节,碱性磷酸酶和骨钙素的含量也明显增加。结论大肠杆菌可表达出有活性的BMP-4,该蛋白可诱导骨髓基质干细胞向成骨细胞分化。     

The effect of bone morphogenetic protein-2 expression on the early fate of skeletal muscle-derived cells

The identification of bone morphogenetic proteins (BMPs) has stimulated intense interest in BMP delivery approaches. Ex vivo BMP-2 gene delivery has recently been described using skeletal muscle-derived cells. Skeletal muscle-derived cells, because of proven efficient transgene delivery and osteocompetence, represent an attractive cell population on which to base ex vivo BMP-2 gene delivery. However, the early in vivo fate of BMP-2-expressing muscle-derived cells is unknown. This study investigates the in vivo effects of BMP-2 secretion on skeletal muscle-derived cells in terms of cell survival and cell differentiation. The first experiment compared survival of BMP-2-expressing cells with control cells during the first 48 h after in vivo implantation. The results demonstrate that BMP-2 secretion did not adversely affect cell survival 8, 24, or 48 h after intramuscular implantation. The second experiment histologically compared the fate of BMP-2-expressing muscle-derived cells to the same cells not expressing BMP-2. The results show that BMP-2 expression prevented in vivo myogenic differentiation and promoted osteogenic differentiation of the transduced cells. This study further supports the existence of osteoprogenitor cells residing within skeletal muscle. Moreover, it is demonstrated that BMP-2 secretion does not adversely affect early cell survival of muscle-derived cells. These data are important for future investigations into BMP-2 gene delivery approaches to the musculoskeletal system.     

Effect of bone morphogenetic protein-12 gene transfer on mesenchymal progenitor cells

Recombinant adenovirus mediated human bone morphogenetic protein-12 gene transfer induced tendon and cartilage-like tissue formation in vivo. The recombinant adenovirus with the human bone morphogenetic protein-12 gene was constructed, and mature human bone morphogenetic protein-12 expression mediated by adenovirus gene transfer was detected by specific antibody. Unlike bone morphogenetic protein-2 gene transfer, bone morphogenetic protein-12 gene transferred mesenchymal progenitor cell line C3H 10T1/2 showed no change of alkaline phosphatase activity, which is the mark of cell differentiation into osteoblastic phenotype. Injection of bone morphogenetic protein-12 gene transferred C3H 10T1/2 cells into nude mice thigh muscles induced tendon and cartilage-like tissue formation. The results indicate bone morphogenetic protein-12 has different effects on mesenchymal progenitor cell differentiation, and it may influence the cell differentiation into a nonosteoblast lineage.     

骨形态发生蛋白-9研究现状

骨形态发生蛋白(BMP)具有多种生物学功能,BMP-9是骨骼系统已知BMP家族中成骨活性最强的成员之一,其骨诱导活性不会受到免疫反应的较大抑制,故也是该家族中唯一能在有免疫能力的动物体内显著诱导骨形成的成员.目前研究证实,动物模型中BMP-9可有效地修复骨缺损和诱导脊柱融合,此外对神经系统、肝脏等骨外器官和组织也具有生物学作用.     

聚乙二醇化骨形态发生蛋白-2基因转染兔骨髓间充质干细胞及其表达测定

目的：聚乙二醇化骨形态发生蛋白-2基因（PEG/BMP-2）纳米颗粒转染兔骨髓间充质干细胞（rBMSCs）,检测骨形态发生蛋白-2（BMP-2）在靶细胞中的表达。方法：原代分离培养rBMSCs,分别采用PEG/BMP-2、脂质体/BMP-2转染细胞,采用流式细胞仪检测转染效率,采用Western Blot和real time RT-PCR方法检测BMP-2表达。结果：成功制备出PEG/BMP-2纳米颗粒并将PEG/BMP-2转染至rBMSCs,转染细胞中BMP-2呈高表达,且与脂质体转染法相比,具有更高的转染效率。结论：PEG/BMP-2纳米颗粒转染rBMSCs可高表达BMP-2,为骨缺损治疗修复提供新的治疗方法。     

The effects of lentiviral gene therapy with bone morphogenetic protein-2-producing bone marrow cells on spinal fusion in rats

STUDY DESIGN: Rat spinal fusion model. OBJECTIVE: This study aimed to assess the ability of rat bone marrow cells (RBMCs) transfected with bone morphogenetic protein (BMP)-2-containing lentivirus to induce a posterolateral spinal fusion in a rat model. SUMMARY OF BACKGROUND DATA: Spinal arthrodesis is a commonly performed spinal procedure and autograft remains the standard for achieving spinal fusion. However, its procurement is associated with significant morbidity, and the rate of pseudoarthrosis has been reported to be 5% to 43%. Nonunion frequently leads to an unsatisfactory resolution of clinical symptoms and usually results in high medical costs and morbidity as well as the need for additional surgeries. These problems have led surgeons to search for alternative solutions to stimulate bone formation. Recombinant BMPs have also been used successfully in clinical trials. However, large doses of BMPs were required to induce adequate bone repair. The development of a regional gene therapy may be a more efficient method to deliver proteins to a specific anatomic site. Furthermore, adeno-BMP-2-producing rat bone marrow-derived cells have been used successfully to induce posterior spinal fusion. Recently, lentiviral vectors on the basis of human immunodeficiency virus have been developed for gene therapy. Lentiviruses are capable of insertion into the host genome, ensuring a prolonged gene expression. However, safety issues are a major concern when adopting these vectors for clinical use. METHODS: In vitro study, we used RBMCs transfected with lentivirus vectors encoding BMP-2 (Lenti-BMP-2), RBMCs transfected with lentivirus vectors encoding the green fluorescent protein (GFP) (Lenti-GFP), and untransfected RBMCs; the latter 2 were used as controls. Alkaline phosphatase (ALP) staining and ALP activity were compared between the groups to assess the ability of the Lenti-BMP-2-transfected RBMCs to stimulate osteoblastic differentiation. In the rat posterolateral spine fusion model, the experimental study comprised 4 groups. Group 1 comprised 6 animals that were implanted with a collagen sponge containing 5 million RBMCs transfected with Lenti-BMP-2. Group 2 comprised 3 animals that were implanted with a collagen sponge containing 5 million RBMCs transfected with Lenti-GFP. Group 3 comprised 6 animals that were implanted with a collagen sponge containing 5 million untransfected RBMCs. Group 4 comprised 3 animals that were implanted with a collagen sponge alone. The rats were assessed by radiographs obtained at 4, 6, and 8 weeks. After death, their spines were explanted and assessed by manual palpation, high-resolution microcomputerized tomography, and histologic analysis. RESULTS: The ALP staining was significantly greater in the Lenti-BMP-2-transfected RBMCs than in the untransfected RBMCs and the Lenti-GFP-transfected RBMCs. The ALP activity was 3-fold greater in the Lenti-BMP-2-transfected RBMCs than in the untransfected RBMCs and the Lenti-GFP-transfected RBMCs. In the rat spine fusion model, radiographic evaluation, high-resolution microcomputerized tomography, and manual palpation revealed spinal fusion in all the rats in Group 1 at 8 weeks. Groups 2, 3, and 4 comprised the control group. None of the rats in the control group (0 of 12) developed fusion at L4-L5. CONCLUSIONS: The present study demonstrated that BMP-2-producing RBMCs, created through lentiviral gene transfer, induced sufficient spinal fusion. The use of lentiviral vectors that contain the cDNA for BMP-2 will be a novel and promising approach for a spinal fusion strategy.     

Increase of bone morphogenetic protein-7 expressing pulmonary resident cells in pneumonectomized rats

Purpose

Compensatory lung growth (CLG) is recognized in rodents subjected to major pulmonary resection; however, the source of cells constituting regenerated tissues during the CLG is still unknown. We investigated the differentiation of lung resident cells and the participation of bone marrow (BM)-derived cells in the remnant lung of pneumonectomized rats.

Methods

After left pneumonectomy, the right remnant lung of Wistar rats was subjected to morphologic and molecular experiments at several time points. We studied the expression of bone morphogenic protein 7 (BMP-7), an accelerator of epithelial differentiation, based on the gene expression profile data of the remnant lung. Next, we evaluated the presence of GFP-positive cells in the remnant lung of Wistar rats that had received BM transplantation from green fluorescent protein (GFP) gene-transgenic Wistar rats prior to left pneumonectomy.

Results

We observed progression of emphysematous change, modulation of gene expression profile, and proliferating cellular nuclear antigen-positive cells in the alveoli of the remnant lungs. BMP-7 protein positive cells were detected in the alveolar septa, which increased significantly over time with the progression of emphysematous change. No bone marrow-derived cells were detected in the right remnant lung of the GFP-BM transferred rats by fluorescence microscopy, immunohistochemistry, or polymerase chain reaction at any time.

Conclusion

Lung resident cells appear to contribute to CLG, possibly via a trans-differentiation pathway.     

骨形态发生蛋白-7基因转染对新生大鼠心肌细胞缺氧-复氧时凋亡的影响

目的 探讨骨形态发生蛋白-7(BMP-7)基因转染对新生大鼠心肌细胞缺氧-复氧时凋亡的影响.方法 体外培养的新生大鼠心肌细胞分为3组:正常对照组(C组)、缺氧-复氧组(HR组)培养的心肌细胞置于95%N2-5%CO2混合气体饱和的缺氧罐中缺氧2 h后,放回孵育箱中继续培养4 h;BMP-7基因转染组(BT组)将含pcDNA3.1-BMP-7基因的质粒在转染试剂作用下转染心肌细胞后72 h再行缺氧2 h,复氧4 h.每组重复8次.计数心肌细胞搏动次数,采用全自动生化分析仪测定乳酸脱氢酶(LDH)活性.倒置显微镜下观察心肌细胞形态.采用流式细胞仪检测心肌细胞凋亡情况,免疫细胞化学方法测定Bcl-2、Bax蛋白的表达.结果 与c组相比,HR组和BT组心肌细胞LDH活性、细胞凋亡率增加,Bax蛋白表达上调,Bcl-2蛋白表达下调(P＜0.05);与HR组相比,BT组心肌细胞LDH活性、细胞凋亡率降低,Bax蛋白表达下调,Bcl-2蛋白表达上调(P＜0.05).结论 BMP-7基因转染可减轻新生大鼠心肌细胞缺氧-复氧损伤,其机制可能与调控Bcl-2、Bax蛋白的表达,抑制心肌细胞凋亡有关.     

Effect of bone morphogenetic protein-2-expressing muscle-derived cells on healing of critical-sized bone defects in mice.

BACKGROUND: Cells that express bone morphogenetic protein-2 (BMP-2) can now be prepared by transduction with adenovirus containing BMP-2 cDNA. Skeletal muscle tissue contains cells that differentiate into osteoblasts on stimulation with BMP-2. The objectives of this study were to prepare BMP-2-expressing muscle-derived cells by transduction of these cells with an adenovirus containing BMP-2 cDNA and to determine whether the BMP-2-expressing muscle-derived cells would elicit the healing of critical-sized bone defects in mice. METHODS: Primary cultures of muscle-derived cells from a normal male mouse were transduced with adenovirus encoding the recombinant human BMP-2 gene (adBMP-2). These cells (5 yen 10(5)) were implanted into a 5-mm-diameter critical-sized skull defect in female SCID (severe combined immunodeficiency strain) mice with use of a collagen sponge as a scaffold. Healing in the treatment and control groups was examined grossly and histologically at two and four weeks. Implanted cells were identified in vivo with use of the Y-chromosome-specific fluorescent in situ hybridization (FISH) technique, and their differentiation into osteogenic cells was demonstrated by osteocalcin immunohistochemistry. RESULTS: Skull defects treated with muscle cells that had been genetically engineered to express BMP-2 had >85% closure within two weeks and 95% to 100% closure within four weeks. Control groups in which the defect was not treated (group 1), treated with collagen only (group 2), or treated with collagen and muscle cells without adBMP-2 (group 3) showed at most 30% to 40% closure of the defect by four weeks, and the majority of the skull defects in those groups showed no healing. Analysis of injected cells in group 4, with the Y-chromosome-specific FISH technique showed that the majority of the transplanted cells were located on the surfaces of the newly formed bone, but a small fraction (approximately 5%) was identified within the osteocyte lacunae of the new bone. Implanted cells found in the new bone stained immunohistochemically for osteocalcin, indicating that they had differentiated in vivo into osteogenic cells. CONCLUSIONS: This study demonstrates that cells derived from muscle tissue that have been genetically engineered to express BMP-2 elicit the healing of critical-sized skull defects in mice. The cells derived from muscle tissue appear to enhance bone-healing by differentiating into osteoblasts in vivo. Clinical Relevance: Ex vivo gene therapy with muscle-derived cells that have been genetically engineered to express BMP-2 may be used to treat nonhealing bone defects. In addition, muscle-derived cells appear to include stem cells, which are easily obtained with muscle biopsy and could be used in gene therapy to deliver BMP-2.     

The effect of the delivery carrier on the quality of bone formed via bone morphogenetic protein-2

Bone morphogenetic protein-2 (BMP-2) can induce bone generation in vivo. Although many studies have demonstrated an increased quantity of regenerated bone after the delivery of BMP-2 using various carriers, little is known about the effect of the carrier type on the quality of the regenerated bone. In this study, we compared the quality of regenerated bone when BMP-2 was delivered with either β-tricalcium phosphate (β-TCP) or heparin-conjugated fibrin (HCF), both of which are shown to be excellent carriers for BMP-2. The profile of the release of BMP-2 was not significantly different between the delivery carriers. However, the alkaline phosphate activity of cultured osteoblasts was significantly higher when BMP-2 was delivered using HCF than when BMP-2 was delivered using β-TCP. To evaluate the quality of the regenerated bone, both types of BMP-2 carriers were implanted into critical-sized calvarial defects in mice. Eight weeks after implantation, the regenerated bone was examined by histomorphometry. Importantly, the treatment using HCF + BMP-2 and β-TCP + BMP-2 resulted in similar bone formation areas. However, the treatment using HCF + BMP-2 resulted in significantly higher bone density than the treatment using β-TCP + BMP-2. This study shows that a BMP-2 delivery carrier can modulate the quality of bone regenerated via BMP-2 delivery.     

Repair of bone allograft fracture using bone morphogenetic protein-2

Long-term clinical data have shown that reconstruction using bone allografts provide adequate function after extensive tumor surgery. Complications such as nonunion of allograft-host interface, infection, and allograft fracture often require major revision surgeries. Allograft fractures usually do not induce the same repair process that is seen in normal fracture healing. The authors did an experimental study to test whether bone morphogenetic protein-2 can induce and achieve osseous repair in an allograft osteotomy model. Recombinant human bone morphogenetic protein-2 was applied at femoral intercalary allograft osteotomy sites in 20 rats. Forty additional rats served as controls (carrier alone and sham). Specimens in all groups were examined histologically and radiographically at 4 and 8 weeks. Specimens in the control groups showed only fibrosis by 8 weeks. In contrast, none of 10 specimens in the experimental group showed radiographic union at 8 weeks. New bone formation and integration with underlying allografts were seen in the experimental group as early as 4 weeks. These data suggest that fracture repair in the allograft bone can be triggered by a biologic regulator that is expressed during normal fracture healing.     

骨形态发生蛋白-7在前列腺癌组织中的表达及意义

目的 研究骨形态发生蛋白-7(BMP-7)在前列腺癌(PCa)组织中的表达及意义.方法 经病理检查确诊PCa组织标本87例.患者平均年龄66(59～78)岁,术前或穿刺前血清总PSA(t-PSA)平均45.7(2.4～138.2)ng/ml.Gleason评分≤6分37例,7分18例,≥8分32例.临床分期:I期(T1a N0 M0)+Ⅱ期(T1b N0M0,T1cN0M0,T2N0M0)20例,Ⅲ期(T3N0M0)20例.Ⅳ期(T4N0M0,TxN1M0,TxN0M1)47例,其中TxN0M1 45例.根据核素骨扫描或正电子发射计算机体层成像CT检查结果分为:PCa无骨转移42例,PCa伴骨转移45例.以30例BPH组织标本为对照,患者平均年龄68(57～88)岁.免疫组织化学PV法检测BMP-7在PCa组织和BPH组织中的表达,统计学分析2组及PCa组内表达差异,PCa组织中BMP-7与血清t-PSA相关性采用Pearson相关性分析. 结果BPH组织中BMP-7表达吸光度A值为70.55±5.41,PCa组织中为70.47±6.18,2者比较差异无统计学意义(P>0.05).PCa无骨转移组BMP-7表达吸光度A值为65.94±1.76,伴有骨转移组为74.80±5.76,2者比较差异有统计学意义(P<0.05).Gleason评分≤6分者吸光度A值为65.96±1.56,7分者为65.83±2.75,≥8分者为78.06±1.39.≤6分和7分者分别与≥8分相比,BMP-7表达A值差异有统计学意义(P<0.05).临床分期编组中Ⅰ期+Ⅱ期BMP-7表达A值为65.86±1.72,Ⅲ期为65.87±1.85,Ⅳ期为74.49±5.83,Ⅰ期+Ⅱ期和Ⅲ期分别与Ⅳ期相比,差异有统计学意义(P<0.05).PCa组织中BMP-7表达A值与血清t-PSA值呈正相关关系(r=0.77,P<0.05).结论 病理Gleason高评分、临床分期、已发生骨转移的PCa组织中高表达BMP-7,BMP-7表达水平与血清t-PSA具有正相关性,提示BMP-7可能是促进PCa细胞发生骨转移的重要细胞因子之一.     

Bone morphogenetic protein-7 protects human intervertebral disc cells in vitro from apoptosis.

BACKGROUND CONTEXT: Disc degeneration includes dysfunction and loss of disc cells leading to a decrease in extracellular matrix (ECM) components. Apoptosis has been identified in degenerated discs. Bone morphogenetic protein-7 (BMP-7) has been reported to stimulate ECM synthesis in the intervertebral disc (IVD), but its effect on disc cell viability is unknown. PURPOSE: To investigate whether BMP-7 can protect disc cells from programmed cell death while enhancing ECM production. STUDY DESIGN: An in vitro study to examine the effect of BMP-7 on apoptosis of IVD cells. METHODS: Human nucleus pulposus (NP) cells were cultured in monolayer, and human recombinant pure BMP-7 (rhBMP-7) was added to the medium when the cells were in the second passage. Thereafter, apoptosis was induced by either tumor necrosis factor-alpha (TNF-alpha) or hydrogen peroxide (H(2)O(2)). Cellular apoptosis was evaluated by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling assay and caspase-3 activity. ECM synthesis was assessed by immunofluorescence for collagen-2 and aggrecan. To study the possibility of bone induction by rhBMP-7 in disc cells, alkaline phosphatase activity and Alizarin red-S staining were evaluated. RESULTS: Apoptosis was induced by both TNF-alpha and H(2)O(2). Addition of rhBMP-7 resulted in inhibition of the apoptotic effects caused by both inducers. Further, BMP-7 decreased caspase-3 activity. In the presence of BMP-7, ECM production was maintained by the cells despite being in an apoptotic environment. No osteoblastic induction of the disc cells was seen. CONCLUSIONS: BMP-7 was demonstrated to prevent apoptosis of human disc cells in vitro. One of the antiapoptotic effects of BMP-7 on NP cells might be a result of its inactivation of caspase-3. Collagen production was maintained by addition of rhBMP-7 in an apoptotic environment.     

Effect of regional gene therapy with bone morphogenetic protein-2-producing bone marrow cells on spinal fusion in rats

BACKGROUND: Bone morphogenetic proteins (BMPs) are now being used as bone-graft substitutes to enhance spinal fusion. However, the large doses of BMP required to induce a spinal fusion in humans suggests that the delivery of these proteins should be improved. We used ex vivo adenoviral gene transfer to create BMP-2-producing bone marrow cells, and these autologous cells were found to induce a posterolateral fusion of the spine in syngeneic rats. METHODS: Intertransverse spinal arthrodesis (L4 and L5) was attempted in ten groups of Lewis rats with 5 x 10 (6) BMP-2-producing rat bone marrow cells (Ad-BMP-2 cells), created through adenoviral gene transfer with guanidine hydrochloride-extracted demineralized bone matrix as a carrier (Group I); 5 x 10 (6) Ad-BMP-2 cells on a collagen sponge carrier (Group II); 10 micro g of recombinant BMP-2 (rhBMP-2) in a guanidine hydrochloride-extracted demineralized bone matrix carrier (Group III); 10 micro g of rhBMP-2 in a collagen sponge carrier (Group IV); autogenous iliac crest bone-grafting (Group V); 5 x 10 (6) beta-galactosidase-producing rat bone marrow cells, created through adenoviral gene transfer with guanidine hydrochloride-extracted demineralized bone matrix as a carrier (Group VI); decortication of the transverse processes alone (Group VII); 5 x 10 (6) uninfected rat bone marrow cells with a guanidine hydrochloride-extracted demineralized bone matrix carrier (Group VIII); guanidine hydrochloride-extracted demineralized bone matrix only (Group IX); or a collagen sponge alone (Group X). Each specimen underwent plain radiography, manual palpation, and histological analysis. RESULTS: All spines in Groups I and II (BMP-2-producing bone marrow cells) and all spines in Groups III and IV were fused at four weeks postoperatively. In contrast, none of the spines in the other groups had fused at a minimum of eight weeks after implantation. Histological analysis of the specimens revealed that the spines that had received BMP-2-producing bone marrow cells (Groups I and II) were filled with coarse trabecular bone postoperatively, whereas those that had received rhBMP-2 (Groups III and IV) were filled with thin, lace-like trabecular bone. All of the other spines, including those that had been treated with autogenous iliac crest bone-grafting (Group V), produced little or no new bone. CONCLUSION: BMP-2-producing bone marrow cells, created by adenoviral gene transfer, produce sufficient BMP to induce an intertransverse fusion in the rat spine model.