The correction of severe spinal deformities often requires osteotomy procedures such as laminectomy. This compromises the structural integrity of the spinal posterior column and may lead to complications including spinal instability, spinal cord exposure, and failure of bone graft fusion. A persistent clinical challenge is how to reconstruct the lamina to restore biomechanical stability while simultaneously protecting the spinal cord. Currently, no standardized artificial lamina repair material is available, and traditional metallic or single-component bioabsorbable materials cannot adequately balance biocompatibility, mechanical support, and physiological healing. To address this need, we designed and fabricated a novel multifunctional split-type porous biomimetic absorbable modified PLA-HA lamina (MSPBA mPLA-HA lamina). In a sheep laminectomy model, the implant-bone interface bone volume fraction in the experimental group reached 74.6% and was significantly higher than that in the control group, while the mechanical stiffness of the reconstructed lamina was comparable to that of native sheep lumbar laminae. These findings suggest that the implant can support lamina reconstruction and that its porous architecture may provide a structural basis for reducing local epidural compression. As a bioactive absorbable construct, the MSPBA mPLA-HA lamina may represent a promising strategy for more physiologically compatible spinal posterior column reconstruction.

Objective: Posterior lumbar interbody fusion (PLIF), posterolateral fusion (PLF), and Hybrid fusion are widely used fusion procedures for lumbar degenerative diseases (LDDs). Postoperative complications dominated by cage migration (CM) and adjacent segment degeneration (ASD) remain major challenges. This study aimed to identify and compare the independent risk factors for CM and ASD in PLIF, PLF, and Hybrid fusion, so as to provide evidence-based references for preoperative evaluation, surgical selection, and complication prevention in clinical practice. Methods: A retrospective cohort study was conducted in patients who underwent PLIF, PLF, or Hybrid fusion for LDDs at our institution. Demographic data (age, gender, and body mass index [BMI]), lifestyle factors (smoking and insobriety), comorbidities (hypertension, diabetes, hyperuricemia, osteoporosis, and hypoalbuminemia), surgical parameters (operative time, intraoperative blood loss, fusion segments, and lumbar lordosis angle), radiological indices (Pfirrmann grading of intervertebral disc degeneration and relative disc height), and biological markers (C-reactive protein/lymphocyte ratio [CLR], procalcitonin [PCT], and serum amyloid A [SAA]) were collected. Patients were stratified into complication and non-complication groups based on the occurrence of CM or ASD. Univariate and binary logistic regression analyses were performed to determine independent risk factors for postoperative complications. Results: A total of 203 patients were enrolled, including 80 cases with complications in the PLIF group, 64 in the Hybrid group, and 59 in the PLF group. No significant differences were noted in the distribution of complication types among the three groups (p = 0.179). Univariate analysis revealed that BMI, osteoporosis, the Pfirrmann grading of superior adjacent disc degeneration, lumbar lordosis angle, operative time, and intraoperative blood loss were significantly associated with postoperative complications across all three surgical groups (p < 0.05). Binary logistic regression analysis confirmed that elevated BMI (PLIF: OR = 1.18, 95%CI: 1.05-4.38; PLF: OR = 1.19, 95%CI: 0.76-2.18; Hybrid: OR = 1.14, 95%CI: 1.07-2.54), osteoporosis (PLIF: OR = 6.86; PLF: OR = 7.62; Hybrid: OR = 5.62), advanced superior adjacent disc degeneration (PLIF: OR = 8.04; PLF: OR = 4.49; Hybrid: OR = 2.87), prolonged operative time, and increased intraoperative blood loss were independent risk factors for postoperative complications. In contrast, age, gender, smoking, insobriety, hypertension, diabetes, CLR, PCT, and SAA were not identified as risk factors (p* > 0.05). Conclusions: Elevated BMI, osteoporosis, pre-existing superior adjacent disc degeneration, prolonged operative time, and increased intraoperative blood loss are shared independent risk factors for CM and ASD following PLIF, PLF, and Hybrid fusion for LDDs. Targeted interventions addressing these factors may reduce postoperative complication rates and improve patient outcomes.

Background: Numerous studies have confirmed that both posterior lumbar interbody fusion (PLIF) and posterior lumbar fusion (PLF), have their advantages and disadvantages. However, the inconsistent results of these studies make it difficult to reach a consensus on which fusion method is superior. Objective: To compare the clinical outcomes of PLIF, PLF, and hybrid surgery combining PLIF and PLF in the treatment of lumbar degenerative disease. Methods: A retrospective review was conducted, collecting clinical records and radiological data of patients with lumbar degenerative disease from 2014 to 2022. Patients were divided into 3 groups based on surgical strategy: PLIF group, PLF group, and hybrid group. Clinical data included patient- reported outcomes such as the Japanese Orthopedic Association score, Oswestry Disability Index score, visual analog scale score, 36- item Short Form Health Survey score, and the occurrence of complications. Radiological data included Cobb angle, fusion rate, adjacent segment degeneration (ASDeg), adjacent segment disease (ASDis), and cage subsidence. Results: A total of 378 patients were divided into 3 groups: PLIF group (n = 122), PLF group (n = 126), and hybrid group (n = 130). The baseline characteristics were balanced among the 3 groups. As the follow- up time increased, visual analog scale scores showed varying degrees of improvement (all P measuretime < 0.001), but there were no significant differences observed between the groups (all P measuretime * group > 0.05). Oswestry Disability Index scores improved over time ( F measuretime = 939, P measure time < 0.001), with the hybrid group showing more significant improvement ( F measure time * group = 2.826, P measure time * group = 0.006). The 36- item Short Form Health Survey scores and Cobb angles also improved significantly during the follow- up period, with no significant differences observed among the groups. The overall fusion rates for the hybrid group and PLIF group were 93% and 91%, significantly higher than the fusion rate of the PLF group (84%; P = 0.031). The postoperative complication rate was significantly higher in the PLIF group (24.4%) compared with the PLF group (16.4%) and the hybrid group (12.5%; P = 0.022). There was no significant difference in the overall 5- year ASDeg occurrence rate (38% vs 36%) and ASDis occurrence rate (11.3% vs 8.3%) between the PLIF group and PLF group for single- level fusion (P > 0.05). The occurrence rate of ASDeg for multilevel fusion in the hybrid group was 29%, significantly lower than that in the PLIF group (42%) and PLF group (37%; P = 0.044). The overall 5- year ASDis occurrence rates for multilevel fusion were 12.3%, 9.9%, and 7.6% for the PLIF group, PLF group, and hybrid group, respectively, with no significant statistical difference (P = 0.338). Conclusion: All 3 surgical techniques might improve the clinical symptoms of patients with degenerative lumbar disease effectively. The hybrid technique demonstrated comparable efficacy to PLIF and PLF in increasing fusion rate, reducing complications, and decreasing the occurrence of ASDeg in multilevel fusion cases significantly. Clinical Relevance: This study holds significant clinical relevance as it directly addresses the treatment outcomes of common surgical interventions for lumbar degenerative disease, a condition that significantly impacts patient quality of life and functionality. This study is also crucial for clinicians when selecting the most appropriate treatment strategy for patients with lumbar degenerative disease.

The treatment of intervertebral disc degeneration (IVDD) is still a huge challenge for clinical updated surgical techniques and basic strategies of intervertebral disc regeneration. Few studies have ever tried to combine surgery and cell therapy to bridge the gap between clinical and basic research. A prospective clinical study with a 72-month follow-up was conducted to assess the safety and feasibility of autologous discogenic cells transplantation combined with discectomy in the treatment of lumbar disc herniation (LDH) and to evaluate the regenerative ability of discogenic cells in IVDD. Forty patients with LDH who were scheduled to have discectomy enrolled in our study and were divided into the observed group (transplantation of autologous discogenic cells after discectomy) and control group (only-discectomy). Serial MRI and X-ray were used to evaluate the degenerative extent of index discs, and clinical scores were used to determine the symptomatic improvement. No adverse events were observed in the observed group, and seven patients in the control group underwent revisions. Both groups had significant improvement of all functional scores post-operatively, with the observed group improving more considerably at 36-month and 72-month follow-up. The height and water content of discs in both groups decreased significantly since 36 months post-op with the control group decreased more obviously. Discectomy combined with autologous discogenic cells transplantation is safe and feasible in the treatment of LDH. Radiological analysis demonstrated that discogenic cells transplantation could slow down the further degeneration of index discs and decrease the complications of discectomy.

随着融合器(Cage)在腰椎后路融合术中得到广泛应用,临床上因Cage脱出而导致的严重并发症越来越多.Cage脱出的危险因素有患者、Cage、手术技术、其他等因素,本文针对Cage脱出危险因素的相关研究进展进行综述.

Intervertebral disc degeneration (IDD) is a condition involving disruption of the bone tissue distribution. Nucleus pulposus mesenchymal stem cells (NPMSCs) and Sirtuin 1 (SIRT1) play important roles in bone diseases, therefore the aim of the present study was to evaluate the roles of SIRT1 and NPMSCs in IDD. First, NPMSCs were harvested from patients with IDD. Then, the NPMSCs were treated with a SIRT activator, and monocyte chemoattractant protein 1 (MCP1) and chemokine receptor 2 (CCR2) inhibitors. Indices related to NPMSC growth, proliferation, differentiation and apoptosis were measured. Subsequently, IDD rat models were established and were transfected with NPMSCs overexpressing SIRT1. NPMSC apoptosis and cartilage differentiation were detected in the rat IDD model. SIRT1 expression was found to be decreased, and the expression of MCP1 and CCR2 increased in NPMSCs of patients with IDD. The upregulation of SIRT1 and the downregulation of the MCP1/CCR2 axis promoted cartilage differentiation and reduced the number of apoptotic NPMSCs. Furthermore, MCP1 reversed the progression of the cartilage differentiation of NPMSCs and the inhibition of NPMSC apoptosis induced by SIRT1 overexpression. Moreover, the transplantation of rat NPMSCs overexpressing SIRT1 relieved IDD in rats. Therefore, SIRT1 overexpression improved cartilage differentiation and reduced the apoptosis of NPMSCs by inactivating the MCP1/CCR2 axis, thus attenuating IDD in rats.

Objective To evaluate the therapeutic effects of limited decompression fusion operation based on selective nerve root blocking and long segment decompression and fusion operation in the treatment of multiple segment lumbar degenerative diseases. Methods Patients diagnosed as the multi-segmental lumbar degenerative disease in our hospital were randomly divided into limited decompression fusion group ( Group A) and long segment decompression group ( Group B). Group A was given the limited decompression and fusion to the responsibility segment ensured by the selective nerve root blocking. Group B was given the wide decompression and fusion to long segments existing nerve root compression on imaging. Operation time, intraoperative blood loss, incision drainage volume and drainage tube placement time of the 2 groups were compared. The JOA scores, ODI index and patient satisfaction assessment were compared at 1, 12, 24 months after follow-up. At the same time, follow-up filming, lumbar dynamic X-ray, and lumbar MRI were applied to observe degeneration of lumbar segments. Results Twenty-seven patients in Group A, 29 in Group B were followed up. Results showed that Group A was significantly less than Group B in operation time [ Group A, ( 128 ± 32) min, Group B, ( 152 ± 28) min ], intraoperative blood loss [ Group A, ( 220 ± 52) ml, Group B, ( 320 ± 68) ml ], incision drainage volume [ Group A, ( 330 ± 55) ml, Group B, ( 480 ±82) ml ] and drainage tube placement time [ Group A, ( 2.8 ± 0.8) d, Group B, ( 4.4 ± 1.2) d ]. JOR scores ( Group A, 21.7 ± 6.1, Group B, 22.9 ± 5.7) and ODI indexes of both groups ( Group A, 21.3 ± 7.2, Group B, 20.1 ± 9.2) at two years after operation were significantly better than that before operation [ JOA: Group A, ( 10.6 ± 2.7), Group B, ( 11.2 ± 3.8); ODI: Group A, ( 65.3 ± 22.7), Group B, ( 71.1 ± 18.2) ]. No significant differences were found between the 2 groups. Follow-up to 1 year, the satisfaction rates of surgery in both groups were above 95％, but there were no significant differences between the 2 groups. However, the complaints ratio of surgery area related discomfort in Group B ( 16/29) was significantly higher than Group A ( 6/27). There was no obvious degeneration of X-ray after operation in both groups. MRI scan showed the degenerative changes of lumbar vertebrae, but there were no significant differences in both groups. Conclusions For multi-segmental complex lumbar degenerative diseases, limited decompression fusion operation based on selective nerve root blocking is a surgical treatment with small trauma and definite therapeutic effects. It is worthy of clinical promotion.

目的 探讨用人骨形态发生蛋白7(human bone morphogenetic protein 7,hBMP7)基因修饰的犬髓核细胞与同种异体椎间盘复合后体内原位移植的方法阻止或延缓同种异体椎间盘移植后退变的能力.方法 将18个比格犬椎间盘(L4-5)置于-196℃冻存液中保存2个月,随机分为A、B、C三组,A组以rAAV2-hBMP7转染犬髓核细胞,将能够表达hBMP7蛋白的髓核细胞悬液(5×106个/ml,20μl)注射入复温后的椎间盘中体外培养,B组以相同数量未转染的hBMP7的髓核细胞注入椎间盘后体外培养,C组以20μl DMEM/F12细胞培养液生理盐水注射后体外培养.将培养7 d的3组椎间盘分别移植至15只比格犬的L4-5,在术前、术后即刻及术后1、3、6个月通过X线检测移植椎间盘的愈合情况及其高度变化;通过MRI T2像分析椎间盘的退变情况;术后6个月时处死动物取材,分析腰椎屈伸、侧弯及扭转的生物力学变化;组织学染色检测椎间盘的结构及退变情况;PCR法检测3组髓核组织中hBMP7 mRNA的表达.结果 X线检测显示3组移植椎间盘高度变化指数(DHVI)差异无统计学意义(P>0.05);MRI检测结果显示:在术后12、24周时,B、C两组移植椎间盘的MRI T2像信号灰度比明显低于A组(P<0.05);生物力学检测结果显示:C组的左右扭转度明显大于A、B组(P<0.05),而A、B两组间的差异无统计学意义(P>0.05),而对屈伸及侧弯活动度检测发现3组间差异无统计学意义;PCR检测结果显示:6个月时A组仍可检测到hBMP mRNA的高表达;组织学检测结果显示:移植后6个月时仍有外源性髓核细胞存活,A组相对于B、C两组髓核结构更加完整,所含细胞外基质更多.结论 表达hBMP7的髓核细胞能阻止同种异体椎间盘移植后的退变性.

The main reason for intervertebral disc (IVD) degeneration is the decrease in the quantity and activity of IVD cells with subsequent reduction of the extracellular matrix (ECM). In this study, we investigated a cell-based repair strategy by injecting nucleus pulposus cells (NPCs) transduced with human bone morphogenetic protein (hBMP7) by adeno-associated virus-2 into the canine degenerative IVD to determine whether NPCs expressing hBMP7 could delay the degeneration of the IVD. Fourteen canines received annular punctures to induce disc degeneration. Eight weeks later, saline (group A), allogeneic NPCs (group B), or allogeneic NPCs transduced with hBMP7 (group C) were injected into the degenerative discs. Twelve weeks after the injection, MRI scan showed that the degeneration process of groups C was slower and less severe compared with that of groups B and C. The IVD stability in group C was superior to that in groups A and B in left-right bending and rotation. HE, safranin-O staining, and ELISA indicated that the degenerative degree of the IVD in group C was significantly milder than that in groups A and B. The study demonstrated that the implantation of NPCs-hBMP7 could effectively maintained the structural integrity, ECM, and biomechanical properties of the canine degenerated discs. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:1311-1322, 2017.

Although nucleus pulposus (NP) tissue engineering has achieved tremendous success, researches still face the huge obstacles in maintaining cell survival and function. A novel functional self-assembled peptide RADA-KPSS was constructed by conjugating BMP-7 short active fragment (KPSS) to the C-terminus of RADA16-I that displays anti-inflammatory and anti-apoptosis effects. However, whether this functional self-assembled RADA-KPSS peptide can alleviate inflammation and NPC apoptosis induced by tumor necrosis factor-alpha (TNF-α) has not been studied. Therefore, we cultured NPCs treated with TNF-α for 48 h with the RADA-KPSS peptide, and compared the results to those with RADA16-I peptide. The cell apoptosis rate, inflammatory mediator secretion, expression of matrix-degrading enzymes, and extracellular matrix (ECM) protein levels were evaluated. The expression of nuclear factor-κB-p65 (NF-κB-p65) protein was also tested. TNF-α-treated NPCs cultured with the RADA16-I peptide showed up-regulated gene expression for matrix-degrading enzymes, such as matrix metalloproteinases-3 (MMP-3), MMP-9, and a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS-4), and down-regulated gene expression for ECM proteins such as aggrecan, collagen II, and Sox-9. The RADA-KPSS peptide could attenuate the expression of MMP-3, MMP-9, and ADAMTS-4, promote accumulation of ECM proteins, and increase secretion of glycosaminoglycan as compared with the RADA16-I peptide. Moreover, the TNF-α-damaged NPCs was further demonstrated to inhibit NF-κB-p65, IL-1, IL-6, and prostaglandin E-2 proteins and decrease cell apoptosis in RADA-KPSS peptide. In conclusion, the functional self-assembled RADA-KPSS peptides have anti-inflammatory and anti-apoptotic effects by promoting anabolic processes and inhibiting catabolic processes in intervertebral disk degeneration. These peptides may be feasible for clinical applications in NP tissue engineering. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 1082-1091, 2018.

Low back pain (LBP) is mainly caused by intervertebral disc degeneration (IDD). Recent studies have demonstrated that the transplantation of mesenchymal stem cells (MSCs) can regenerate regions that have undergone degeneration, and the regenerative effect can be enhanced by using a hydrogel carrier. This article describes an injectable functional hydrogel system manufactured by combining RADA16-I and RADA-KPSS (RADA-KPSS was manufactured by conjugating a bioactive motif derived from BMP-7 [KPSS] onto the C terminal of RADA16-I) at a volume ratio of 1:1. This hydrogel system can enhance the proliferation, differentiation, and chemotactic migration of BMSCs. In addition, the encapsulation of BMSCs with this system maintains cell viability for a long period after transplantation into an ex vivo cultured disc model. In conclusion, KPSS-conjugated RADKPS is an ideal encapsulation system for BMSCs in intervertebral disc (IVD) regeneration.

Tissue engineering has shown great success in the treatment of intervertebral disk degeneration (IVDD) in the past decade. However, the adverse and harsh microenvironment associated in the intervertebral disks remains a great obstacle for the survival of transplanted cells. Although increasing numbers of new materials have been created or modified to overcome this hurdle, a new effective strategy of biological therapy is still required. In this study, bone morphogenic protein 7 (BMP7)-based functionalized self-assembling peptides were developed by conjugating a bioactive motif from BMP-7 (RKPS) onto the C-terminal of the peptide RADARADARADARADA (RADA16-I) at a ratio of 1:1 to form a new RADARKPS peptide. Human nucleus pulposus-derived stem cells (NPDCs) were cultured in the presence of RADA-RKPS or RADA16-I in an apoptosis-promoting environment that was induced by tumor necrosis factor-alpha, and cells were cultured with RADA16-I in normal medium that served as the control group. After 48h of apoptosis induction, the viability, proliferation, apoptosis rate, and expression of apoptosis-related genes of NPDCs in the different groups were evaluated, and the differentiation of NPDCs toward nucleus pulposus-like cells was tested. The results showed that the RADA-RKPS peptide could significantly protect the survival and proliferation of NPDCs. In addition, the application of RADA-RKPS decreased the rate of cell apoptosis, as detected by TUNEL-positive staining. Furthermore, our in vitro study confirmed the apoptosis-protecting effects of RADA-RKPS peptides, which significantly reduced the BAX/BCL-2 ratio of NPDCs and upregulated the gene expression of collagen II a1, aggrecan, and Sox-9 after 48h of apoptosis induction. Collectively, these lines of evidence suggest that RADA-RKPS peptides confer a protective effect to NPDCs in an apoptosis environment, suggesting their potential application in the development of new biological treatment strategies for IVDD.

In vivo angiogenesis in a three-dimensional bone graft after the implantation of spherical porous β-tricalcium phosphate scaffolding materials into lumbodorsal fascia of New Zealand rabbits.

Nucleus pulposus (NP) tissue engineering has been demonstrated to be a feasible therapeutic strategy for intervertebral disc regeneration. In this study, we constructed a novel injectable biomaterial by conjugating three different short peptides of BMP7 to the C-terminus of the self-assembling peptide RADA16-I, and we then mixed each of these conjugates with RADA16-I at equal volumes to obtain the novel functionalized peptides RAD-SNV, RAD-KPS, and RAD-KAI. The bioactivities of these functionalized peptides for human degenerated NP cells (hdNPCs) were evaluated in vitro, and the most ideal scaffold was chosen for assessment of its in vivo degradation and the tissue reactions to it. All of the functionalized peptides self-assembled to form hydrogel scaffolds with a nanofibrous structure under physiological conditions. Compared with the RADA16-I and RAD-KAI scaffolds, the RAD-SNV and RAD-KPS scaffolds possessed better bioactivities for hdNPCs, which were characterized by their enhanced proliferation, migration, and ECM (collagen II, aggrecan, and sox-9) secretion. RAD-KPS was chosen over RAD-SNV as the most ideal scaffold material due to the cells' higher rate of expression of aggrecan both at the gene and protein level after 28 days of coculture. Moreover, in vivo analysis demonstrated that subcutaneously injected RAD-KPS degraded in vivo without invoking intense inflammation. Therefore, RAD-KPS is an ideal candidate scaffold for NP tissue engineering and holds great potential for NP regeneration.

STUDY DESIGN:An animal experiment about intervertebral disc allograft.OBJECTIVE:To explore the feasibility to decellularize disc allografts treated by 6°Co Gamma Irradiation, and simultaneously, to assess the possibility to make use of the decellularized natural disc scaffold for disc degeneration biotherapy.SUMMARY OF BACKGROUND DATA:Studies of both animal and human disc allograft transplantation indicated that the disc allograft may serve as a scaffold to undertake the physiological responsibility of the segment.METHODS:Experiment in vitro: 48 discs of beagles were harvested and divided randomly into four groups including a control group and three irradiated groups. Immediate cell viability and biomechanical properties of the discs were checked and comparisons were made among these groups. Experiment in vivo: 24 beagles accepted single-level allografted disc treated with different doses of gamma irradiation. Plain X-rays and MRIs were taken before and after surgery. Then, the spinal columns were harvested en bloc from the sacrificed beagles and were examined morphologically.RESULTS:There were significant differences of both the annulus fibrosus and nucleus pulposus immediate cell viabilities among the various groups. There were no obvious differences of the biomechanical properties among the four groups. The disc height and range of motion decreased significantly in all groups as time went on. The observed indexes in irradiated groups were much smaller than those in the control group, but the indexes in 18-kGy group were larger than those in 25-kGy and 50-kGy groups. Both MRI and macroscopic findings showed that the segmental degeneration in the control and 18-kGy group was less severe than that in 25-kGy and 50-kGy groups.CONCLUSION:Gamma Irradiation can decellularize disc allograft successfully to provide natural scaffold for the study of degenerative disc disease therapy, and also can be used as an effective method to produce adjustable animal models.

Nucleus pulposus (NP) tissue engineering has been proposed as a novel biological treatment for early-stage intervertebral disc degeneration. In this study, a novel functional self-assembling peptide PKP was first designed by linking the short functional motif of bone morphogenetic protein-7 (BMP7) to the C-terminal of RADA16-I, and another new functional self-assembling peptide was obtained by mixing RKP with RADA16-I. Then, the biocompatibilities and bioactivities of RKP and RAD-RKP for human degenerated nucleus pulposus cells (hNPCs) were studied in vitro. Atomic force microscopy and scanning electron microscopy (SEM) confirmed that both RKP and RAD-RKP could self-assemble into three-dimensional (3D) nanofiber hydrogel scaffolds in a culture medium at 37°C. After the hNPCs were cultured in 3D scaffolds, both RKP and RAD-RKP exhibited reliable attachment and extremely low cytotoxicities (<14%), which were verified by SEM and cytotoxity assays, respectively. Our results also showed that the functional-based scaffolds could increase the proliferation and migration of hNPCs after 7 days compared with culture plates and pure RADA16-I. Quantitative real-time polymerase chain reaction demonstrated that the expressions of collagen II α1, Sox-9, and aggrecan were upregulated, while collagen I α1 was downregulated by functional-based scaffolds after 28 days. Furthermore, we also confirmed that RAD-RKP exhibited a higher hNPC proliferation, migration, and expression of Sox-9 and aggrecan compared with pure RKP. Therefore, the results of this study indicated that the BMP7 short motif-designed functional self-assembling peptide nanofiber hydrogels could be used as excellent scaffolds in NP tissue engineering, and RAD-RKP might have further potential application in human mild degenerated NP tissue regeneration.

Objective To investigate the feasibility of constructing the biological intervertebral disc using cryopreserved intervertebral disc with injected nucleus pulposus cells(NPCs) expressing bone morphogenetic protein-7(hBMP7) in vitro.Methods The 48 intervertebral discs stored in liquid nitrogen for 2 months were divided into EGFP control,1×104 NPCs,1×105 NPCs and 1×106 NPCs groups.In EGFP group,20 μL cell suspension including 1×105 NPCs expressing EGFP was injected into the intervertebral disc.In 1×104,1×105 and 1×106 NPCs groups,20 μL cell suspension including 1×104,1×105 and 1×106 NPCs expressing hBMP7 by PKH-26 dying were injected into the intervertebral discs respectively.Intervertebral discs injected were immerged into 30 mL culture medium.The cell survival,cell fluorescence intensity,hBMP7 mRNA expression,PG and collagen contents of the intervertebral discs in each group were assessed at culture days 4,7 and 14.Results: Endogeneous NPCs remained alive at 4th,7th and 14th day of culture.Results showed that cell fluorescence intensity in 1×105 NPCs group was higher than that in 1×106 and 1×104 NPCs groups.The hBMP7 mRNA expression levels in 1×105 NPCs group were higher than those in 1×106 and 1×104 NPCs groups at 7th and 14th day of culture.The hBMP7 mRNA expression in EGFP group was not detectable at any time points.Moreover,the PG and collagen contents in 1×105 NPCs group were higher than those in other groups at 7th and 14th day of culture.Conclusion It is feasible that the biological intervertebral disc is constructed to use cryopreserved intervertebral disc with injected NPCs expressing hBMP7,and then cultured for the suitable time in vitro.

Objective To compare the effect of two decellularizing methods for the preparation of acellular nucleus pulposus,and provide ideal scaffold for tissue engineered intervertebral disc which has potential application in treatment of disc degenerative diseases.Methods Integrated nucleus pulposus was harvested from 1-month-old(mean body weight 0.6 kg) and 1-year-old(mean body weight 7.5 kg) Beagle’s dogs,and 80 of complete structured nucleus organization were collected,which were divided into 4 groups(n = 10) and then decellularized by the method of chemical extraction(immersed in 30 mL/L Triton X-100 and 40 mL/L sodium deoxycholate) or by enzyme digestion(immersed in 0.5 % Trypsin followed by the mixture of 1 × 103 U/mL DNase and RNase) respectively.The degree of decellularization,demyelization and integrity of the scaffold construct were assessed by HE staining and scoring system.The cytocompatibility of acellular nucleus pulposus was measured by methyl-thiazolyl-tetrazolium(MTT) assay.Results The decellularizing effects of 1-month-old nucleus pulposus by the 2 methods were satisfactory,which were(4.32 ± 4.15) / high power field(group chemical extraction) and(4.98 ± 4.52) / high power field(group enzyme digestion).But the scaffold construct were seriously damaged at the same time,and cytotoxicity classification in 1-week were grade 0 and grade 1,respectively.Nucleus pulposus cells of 1-year-old Beagle’s dogs disappeared by the method of enzyme digestion and maintenance of scaffolds were superior over that of chemical extraction method.Conclusion It is demonstrated that decellularized nucleus pulposus of 1-year-old Beagle’s dogs by enzyme digestion maintains normal function without obvious cell toxicity,so it could be a good candidate for natural acellular scaffold based tissue engineering of intervertebral disc.

Human degenerative disc disease (DDD) is characterized by progressive loss of human nucleus pulposus (HNP) cells and extracellular matrix, in which the massive deposition are secreted by HNP cells. Cell therapy to supplement HNP cells to degenerated discs has been thought to be a promising strategy to treat DDD. However, obtaining a large quality of fully functional HNP cells has been severely hampered by limited proliferation capacity of HNP cells in vitro. Previous studies have used lipofectamine or recombinant adeno-associated viral (rAAV) vectors to deliver human telomerase reverse transcriptase (hTERT) into ovine or HNP cells to prolong the activity of nucleus pulposus cells with limited success. Here we developed a lentiviral vector bearing both hTERT and a gene encoding green fluorescence protein (L-hTERT/EGFP). This vector efficiently mediated both hTERT and EGFP into freshly isolated HNP cells. The expressions of both transgenes in L-hTERT/EGFP transduced HNP cells were detected up to day 210 post viral infection, which was twice as long as rAAV vector did. Furthermore, we observed restored telomerase activity, maintained telomere length, delayed cell senescence, and increased cell proliferation rate in those L-hTERT/EGFP transduced HNP cells. Our study suggests that lentiviral vector might be a useful gene delivery vehicle for HNP cell therapy to treat DDD.

ABSTRACTWe have previously explored the possibilities of allogenic intervertebral disc (IVD) curing disc degeneration disease in clinical practice. The results showed that the motion and stability of the spinal unit was preserved after transplantation of allogenic IVD in human beings at 5‐year follow‐up. However, mild degeneration was observed in the allogenic transplanted IVD cases. In this study, we construct the biological tissue engineering IVD by injecting the nucleus pulposus cells (NPCs) expressing human bone morphogenetic protein 7 (hBMP7) into cryopreserved IVD, and transplant the biological tissue engineering IVD into a beagle dog to investigate whether NPCs expressing hBMP7 could prevent the degeneration of the transplanted allogenic IVDs. At 24 weeks after transplantation, MRI scan showed that IVD allografts injected NPCs expressing hBMP7 have a slighter signs of degeneration than IVD allografts with NPCs or without NPCs. The range of motion of left–right rotation in the group without NPCs was bigger than that of two cells injection group. PKH‐26‐labeled cells were identified at IVD allograft. The study demonstrated that NPCs expressing hBMP7 could survive at least 24 weeks and prevent the degeneration of the transplanted IVD. This solution might have a potential role in preventing the IVD allograft degeneration in long time follow‐up. © 2013 Orthopaedic Research Society Published by Wiley Periodicals, Inc. J Orthop Res 31:1366–1373, 2013