Macrophage-Mediated Bone Formation in Scaffolds Modified With MSC-Derived Extracellular Matrix Is Dependent on the Migration Inhibitory Factor Signaling Pathway.

The positive role of macrophages in the osteogenesis of mesenchymal stem cells (MSCs) has been a recent research focus. On the other hand, MSCs could carefully regulate the paracrine molecules derived from macrophages. Human umbilical cord mesenchymal stem cells (hucMSCs) can reduce the secretion of inflammatory factors from macrophages to improve injury healing. hucMSC-derived extracellular matrix (hucMSC-ECM) has the similar effect to hucMSCs, which could combat the inflammatory response of macrophages. Additionally, MSC-derived extracellular matrix also enhanced bone regeneration by inhibiting osteoclastic differentiation of monocyte/macrophage lineage. However, whether hucMSC-ECM could improve bone formation by guiding macrophage-induced osteogenic differentiation of MSCs is unknown. Here, we present decalcified bone scaffolds modified by hucMSC-derived extracellular matrix (DBM-ECM), which maintained multiple soluble cytokines from hucMSCs, including macrophage migration inhibitory factor (MIF). Compared with DBM, the DBM-ECM scaffolds induced bone formation in an improved heterotopic ossification model of severe combined immunodeficiency (SCID) mice in a macrophage-dependent manner. Macrophages cocultured with DBM-ECM expressed four osteoinductive cytokines (BMP2, FGF2, TGFß3 and OSM), which were screened out by RNA sequencing and measured by qPCR and western blot. The conditioned medium from macrophages cocultured with DBM-ECM improved the osteogenic differentiation of hBMSCs. Furthermore, DBM-ECM activated CD74/CD44 (the typical MIF receptors) signal transduction in macrophages, including phosphorylation of P38 and dephosphorylation of c-jun. On the other side, the inhibitory effects of the DBM-ECM scaffolds with a deficient of MIF on osteogenesis in vitro and in vivo revealed that macrophage-mediated osteogenesis depended on MIF/CD74 signal transduction. The results of this study indicate that the coordinated crosstalk of macrophages and MSCs plays a key role on bone regeneration, with an emphasis on hucMSC-ECM constructing a macrophage-derived osteoinductive microenvironment.

Corrigendum to 'Phosphorylation inhibition of protein-tyrosine phosphatase 1B tyrosine-152 induces bone regeneration coupled with angiogenesis for bone tissue engineering' [Bioactive Mater. Vol. 6, Issue 7, Pages 2039-2057].

[This corrects the article DOI: 10.1016/j.bioactmat.2020.12.025.].

Laminin alpha 4 promotes bone regeneration by facilitating cell adhesion and vascularization.

Selective cell retention (SCR) has been widely used as a bone tissue engineering technique for the real-time fabrication of bone grafts. The greater the number of mesenchymal stem cells (MSCs) and endothelial progenitor cells (EPCs) retained in the scaffold, the better the osteoinductive and angiogenic properties of the scaffold's microenvironment. Improved bioscaffold properties in turn lead to improved bone graft survival, bone regeneration, and angiogenesis. Laminin plays a key role in cell-matrix adhesion, cell proliferation, and differentiation. We designed a collagen-binding domain (CBD) containing the core functional amino acid sequences of laminin α4 (CBD-LN peptide) to supplement the functional surface of a collagen-based decalcified bone matrix (DBM) scaffold. This scaffold promoted MSCs and EPCs early cell adhesion through up-regulating the expression of integrin α5ß1 and integrin αvß3 respectively, thus accelerated the following cell spreading, proliferation, and differentiation. Interestingly, it promoted the retention of MSCs (CD90+/CD105+ cells) and EPCs (CD31+ cells) in the scaffold following the use of clinical SCR technology. Furthermore, the DBM/CBD-LN scaffold induced the formation of type H vessels through the activation of the HIF-1α signaling pathway. The DBM/CBD-LN scaffold displayed rapid bone formation and angiogenesis in vivo, suggesting that it might be used as a new biomaterial in bone tissue engineering. STATEMENT OF SIGNIFICANCE: Selective cell retention technology (SCR) has been utilized in clinical settings to manufacture bioactive bone grafts. Specifically, demineralized bone matrix (DBM) is a widely-used SCR clinical biomaterial but it displays poor adhesion performance and angiogenic activity. In this work, we designed a collagen-binding domain (CBD) containing the core functional amino acid sequences of laminin α4 to supplement the functional surface of a collagen-based DBM scaffold. This bioscaffold promoted SCR-mediated MSCs and EPCs early cell adhesion, thus accelerated the following cell spreading, proliferation, and differentiation. Our results indicate this bioscaffold greatly induced osteogenesis and angiogenesis in vivo. In general, this bioscaffold has a good prospect for SCR application and may provide highly bioactive bone implant in clinical environment.

Phosphorylation inhibition of protein-tyrosine phosphatase 1B tyrosine-152 induces bone regeneration coupled with angiogenesis for bone tissue engineering.

A close relationship has been reported to exist between cadherin-mediated cell-cell adhesion and integrin-mediated cell mobility, and protein tyrosine phosphatase 1B (PTP1B) may be involved in maintaining this homeostasis. The stable residence of mesenchymal stem cells (MSCs) and endothelial cells (ECs) in their niches is closely related to the regulation of PTP1B. However, the exact role of the departure of MSCs and ECs from their niches during bone regeneration is largely unknown. Here, we show that the phosphorylation state of PTP1B tyrosine-152 (Y152) plays a central role in initiating the departure of these cells from their niches and their subsequent recruitment to bone defects. Based on our previous design of a PTP1B Y152 region-mimicking peptide (152RM) that significantly inhibits the phosphorylation of PTP1B Y152, further investigations revealed that 152RM enhanced cell migration partly via integrin αvß3 and promoted MSCs osteogenic differentiation partly by inhibiting ATF3. Moreover, 152RM induced type H vessels formation by activating Notch signaling. Demineralized bone matrix (DBM) scaffolds were fabricated with mesoporous silica nanoparticles (MSNs), and 152RM was then loaded onto them by electrostatic adsorption. The DBM-MSN/152RM scaffolds were demonstrated to induce bone formation and type H vessels expansion in vivo. In conclusion, our data reveal that 152RM contributes to bone formation by coupling osteogenesis with angiogenesis, which may offer a potential therapeutic strategy for bone defects.

Comparison of Individual Tissue-Engineered Bones and Allogeneic Bone in Treating Bone Defects: A Long-Term Follow-Up Study.

The treatment of bone defects has always been a challenge for orthopedic surgeons. The development of tissue engineering technology provides a novel method for repairing bone defects and has been used in animal experiments and clinical trials. However, there are few clinical studies on comparing the long-term outcomes of tissue-engineered bones (TEBs) and other bone grafts in treating bone defects, and the long-term efficiency of TEBs remains controversial. Therefore, a study designed by us was aimed to compare the long-term efficacy and safety of individual tissue-engineered bones (iTEBs) and allogeneic bone granules (ABGs) in treating bone defects caused by curettage of benign bone tumors and tumor-like lesions. From September 2003 to November 2009, 48 patients who received tumor curettage and bone grafting were analyzed with a mean follow-up of 122 mo (range 60 to 173 mo). Based on implant style, patients were divided into groups of iTEBs (n = 23) and ABGs (n = 25). Postoperatively, the healing time, healing quality, incidence of complications, and functional scores were compared between the two groups. The Musculoskeletal Tumor Society functional evaluation system and Activities of Daily Living Scale scores were significantly improved in both groups with no significant difference. The average healing time of ABGs was longer than that of iTEBs (P < 0.05). At the final follow-up, iTEBs had a better performance in the bone healing quality evaluated by modified Neer classification (P < 0.05). In the group of iTEBs, the complication and reoperation rate was lower than that in the group of ABGs, with no tumorigenesis or immune rejection observed. In summary, for treating bone defects caused by tumor curettage, iTEBs were safe, effective, and tagged with more rapid healing speed, better healing outcome, and lower complication and reoperation rate, in comparison with ABGs.

Engineered scaffolds based on mesenchymal stem cells/preosteoclasts extracellular matrix promote bone regeneration.

Recently, extracellular matrix-based tissue-engineered bone is a promising approach to repairing bone defects, and the seed cells are mostly mesenchymal stem cells. However, bone remodelling is a complex biological process, in which osteoclasts perform bone resorption and osteoblasts dominate bone formation. The interaction and coupling of these two kinds of cells is the key to bone repair. Therefore, the extracellular matrix secreted by the mesenchymal stem cells alone cannot mimic a complex bone regeneration microenvironment, and the addition of extracellular matrix by preosteoclasts may contribute as an effective strategy for bone regeneration. Here, we established the mesenchymal stem cell/preosteoclast extracellular matrix -based tissue-engineered bones and demonstrated that engineered-scaffolds based on mesenchymal stem cell/ preosteoclast extracellular matrix significantly enhanced osteogenesis in a 3 mm rat femur defect model compared with mesenchymal stem cell alone. The bioactive proteins released from the mesenchymal stem cell/ preosteoclast extracellular matrix based tissue-engineered bones also promoted the migration, adhesion, and osteogenic differentiation of mesenchymal stem cells in vitro. As for the mechanisms, the iTRAQ-labeled mass spectrometry was performed, and 608 differentially expressed proteins were found, including the IGFBP5 and CXCL12. Through in vitro studies, we proved that CXCL12 and IGFBP5 proteins, mainly released from the preosteoclasts, contributed to mesenchymal stem cells migration and osteogenic differentiation, respectively. Overall, our research, for the first time, introduce pre-osteoclast into the tissue engineering of bone and optimize the strategy of constructing extracellular matrix-based tissue-engineered bone using different cells to simulate the natural bone regeneration environment, which provides new sight for bone tissue engineering.

Modification of PLGA Scaffold by MSC-Derived Extracellular Matrix Combats Macrophage Inflammation to Initiate Bone Regeneration via TGF-ß-Induced Protein.

The immunologic response toward chronic inflammation or bone regeneration via the accumulation of M1 or M2 macrophages after injury could determine the fate of biomaterial. Human umbilical cord mesenchymal stem cells (hUCMSCs) have a pivotal immunomodulatory property on directing macrophage behaviors. Herein, for the first time, 3D-printed poly(lactide-co-glycolide) (PLGA) scaffolds modified with hUCMSC-derived extracellular matrix (PLGA-ECM) are prepared by a facile tissue engineering technique with physical decellularization and 2.44 ± 0.29 mg cm-3 proteins immobilized on the PLGA-ECM contain multiple soluble cytokines with a sustainable release profile. The PLGA-ECM not only attenuates the foreign body response, but also improves bone regeneration by increasing the accumulation of M2 macrophages in an improved heterotopic transplantation model of SCID mice. Furthermore, the PLGA-ECM scaffolds with the knockdown of transforming growth factor-ß-induced protein (TGFßI/ßig-H3) demonstrate that M2 macrophage accumulation improved by the PLGA-ECM could be attributed to increasing the migration of M2 macrophages and the repolarization of M1 macrophages to M2 phenotype, which are mediated by multiple integrin signaling pathways involving in integrin ß7, integrin α9, and integrin ß1 in a TGFßI-dependent manner. This study presents an effective surface modification strategy of polymeric scaffolds to initiate tissue regeneration and combat inflammatory response by increasing M2 macrophage accumulation.

Individual Tissue-Engineered Bone in Repairing Bone Defects: A 10-Year Follow-Up Study.

Bone defects caused by various causes remain a major problem in orthopedic clinics. A number of different treatments have been developed and proposed, but until now, none has proven to be completely satisfactory. For 26 patients with bone defects but limited autologous bone source or allogeneic bone graft failure, we used individual tissue-engineered bones (iTEBs) for repairing, which were constructed by autologous bone marrow mesenchymal stem cells and allogenic decalcified bone matrix (DBM) scaffolds. The clinical outcomes, including efficacy and safety, were evaluated by radiological examinations, postoperative function recovery score and laboratory tests. Twenty-six patients, including 18 men and 8 women, were followed up for an average of 10 years to analyze the long-term outcome. The mean healing time for patients with lacunar bone defects was 3.87 ± 2.01 months (range, 2-9 months) and that for structural bone defects was longer than 12 months. The Musculoskeletal Tumor Society functional evaluation system and the Barthel Index scores were significantly improved during the long-term follow-up. The white blood cell, erythrocyte sedimentation rate, C reactive protein, complement, immunoglobulins, and liver and renal functions were not significantly affected by bone grafting. One patient with bone cyst relapsed at 3 years postoperatively and achieved bone healing after re-transplantation. No tumorigenesis, tumor metastasis, or blood transmissible disease was found in the whole process. The results demonstrated that iTEBs were effective and safe for repairing bone defects in the long period, especially for those with lacunar bone defects and limited autograft source. Impact statement Currently, controversies exist about the long-term safety and effectiveness of the clinical application of tissue-engineered bones (TEBs) due to potential tumorigenesis, immune rejection, disease transmission, and others. In this study, we show that individual TEBs constructed by autologous MSCs and allogenic decalcified bone matrix are reliable for repairing bone defects in regard to its long-term safety and effectiveness. Our study provides experience and basis about the clinical application of TEBs in the treatment of bone defects.

A Standardized and Quality-Controllable Protocol of Constructing Individual Tissue-Engineered Grafts Applicable to Treating Large Bone Defects.

Patient-specific individual tissue-engineered bones (iTEBs) have been recognized as a promising strategy for treating large bone defects. However, current construction protocols of iTEBs vary between lots and lack standardization and quality control, hampering further research and application. This study was aimed to detail a standardized constructing protocol for iTEBs, which can be used for both clinical and experimental purposes. The procedure was designed and described as follows: scaffold preparation, cell isolation and culture, and fabrication of iTEBs. Manipulation and caution points in each section were detailed. A series of scales on the quality control and safety monitoring was developed. The effectiveness and safety of iTEBs were evaluated. Eventually, the preparing portion, from cell culture to scaffold treatment, usually required 21 days. Generally, the fabrication section took 5 days. The main advantage of this protocol was that each step was standardized and quality controlling and safety monitoring were performed throughout the process to ensure the homogeneity, reliability, and safety. The resulting iTEBs were effective and applicable to both clinical and experimental purposes. Thus, we have established a refined and standardized protocol detailing the construction process of patient-specific iTEBs that comply with strict quality control and safety criteria. This protocol is relatively easy for graduate students or staff working in the field of bone tissue engineering to implement.

Multiple integrin ligands provide a highly adhesive and osteoinductive surface that improves selective cell retention technology.

Among various bone tissue engineering strategies, selective cell retention (SCR) technology has been used as a practical clinical method for bone graft manufacturing in real time. The more mesenchymal stem cells (MSCs) are retained, the better the osteoinductive microenvironment provided by the scaffold, which in turn promotes the osteogenesis of the SCR-fabricated bone grafts. Integrin receptors are crucial to cell-matrix adhesion and signal transduction. We designed a collagen-binding domain (CBD)-containing IKVAV-cRGD peptide (CBD-IKVAV-cRGD peptide) to complement the collagen-based demineralized bone matrix (DBM) with a functionalized surface containing multiple integrin ligands, which correspond to the highly expressed integrin subtypes on MSCs. This DBM/CBD-IKVAV-cRGD composite exhibited superior in vitro adhesion capacity to cultured MSCs, as determined by oscillatory cell adhesion assay, centrifugal cell adhesion assay and mimetic SCR. Moreover, it promoted the retention of MSC-like CD271+ cells and MSC-like CD90+/CD105+ cells in the clinical SCR method. Furthermore, the DBM/CBD-IKVAV-cRGD composite induced robust MSC osteogenesis, coupled with the activation of the downstream FAK-ERK1/2 signaling pathway of integrins. The SCR-prepared DBM/CBD-IKVAV-cRGD composite displayed superior in vivo osteogenesis, indicating that it may be potentially utilized as a biomaterial in SCR-mediated bone transplantation. STATEMENT OF SIGNIFICANCE: Selective cell retention technology (SCR) has been utilized in clinical settings to manufacture bioactive bone grafts. Specifically, demineralized bone matrix (DBM) is a widely-used SCR clinical biomaterial but it displays poor adhesion performance and osteoinduction. Improvements of the DBM that promote cell adhesion and osteoinduction will benefit SCR-prepared implants. In this work, we developed a novel peptide that complements the DBM with a functionalized surface of multiple integrin ligands, which are corresponding to integrin subtypes available on human bone marrow-derived mesenchymal stem cells (MSCs). Our results indicate this novel functionalized bioscaffold greatly increases SCR-mediated MSC adhesion and in vivo osteogenesis. Overall, this novel material has promising SCR applications and may likely provide highly bioactive bone implants in clinical settings.

IGFBP3 deposited in the human umbilical cord mesenchymal stem cell-secreted extracellular matrix promotes bone formation.

The extracellular matrix (ECM) contains rich biological cues for cell recruitment, proliferationm, and even differentiation. The osteoinductive potential of scaffolds could be enhanced through human bone marrow mesenchymal stem cell (hBMSC) directly depositing ECM on surface of scaffolds. However, the role and mechanism of human umbilical cord mesenchymal stem cells (hUCMSC)-secreted ECM in bone formation remain unknown. We tested the osteoinductive properties of a hUCMSC-secreted ECM construct (hUCMSC-ECM) in a large femur defect of a severe combined immunodeficiency (SCID) mouse model. The hUCMSC-ECM improved the colonization of endogenous MSCs and bone regeneration, similar to the hUCMSC-seeded scaffold and superior to the scaffold substrate. Besides, the hUCMSC-ECM enhanced the promigratory molecular expressions of the homing cells, including CCR2 and TßRI. Furthermore, the hUCMSC-ECM increased the number of migrated MSCs by nearly 3.3 ± 0.1-fold, relative to the scaffold substrate. As the most abundant cytokine deposited in the hUCMSC-ECM, insulin-like growth factor binding protein 3 (IGFBP3) promoted hBMSC migration in the TßRI/II- and CCR2-dependent mechanisms. The hUCMSC-ECM integrating shRNA-mediated silencing of Igfbp3 that down-regulated IGFBP3 expression by approximately 60%, reduced the number of migrated hBMSCs by 47%. In vivo, the hUCMSC-ECM recruited 10-fold more endogenous MSCs to initiate bone formation compared to the scaffold substrate. The knock-down of Igfbp3 in the hUCMSC-ECM inhibited nearly 60% of MSC homing and bone regeneration capacity. This research demonstrates that IGFBP3 is an important MSC homing molecule and the therapeutic potential of hUCMSC-ECM in bone regeneration is enhanced by improving MSC homing in an IGFBP3-dependent mechanism.

TGFß3 recruits endogenous mesenchymal stem cells to initiate bone regeneration.

BACKGROUND: The recruitment of a sufficient number of endogenous mesenchymal stem cells (MSCs) is the first stage of in-situ tissue regeneration. Transforming growth factor beta-3 (TGFß3) could recruit stem or progenitor cells and endothelial cells to participate in tissue regeneration. However, the mechanism of TGFß3 recruiting MSCs toward bone regeneration has remained obscure. METHODS: We estimated the promigratory property of TGFß3 on human bone marrow MSCs (hBMSCs) cocultured with the vascular cells (human umbilical artery smooth muscle cells or human umbilical vein endothelial cells) or not by Transwell assay. After the addition of the inhibitor (SB431542) or Smad3 siRNA, the levels of MCP1 and SDF1 in coculture medium were tested by ELISA kit, and then the migratory signaling pathway of hBMSCs induced by TGFß3 was investigated by western blot analysis. In vivo, a 2-mm FVB/N mouse femur defect model was used to evaluate chemokine secretion, endogenous cell homing, and bone regeneration induced by scaffolds loading 1 µg TGFß3 through qPCR, immunofluorescent staining, immunohistochemical analysis, and Micro-CT, compared to the vehicle group. RESULTS: TGFß3 (25 ng/ml) directly showed a nearly 40% increase in migrated hBMSCs via the TGFß signaling pathway, compared to the vehicle treatment. Then, in the coculture system of hBMSCs and vascular cells, TGFß3 further upregulated nearly 3-fold MCP1 secretion from vascular cells in a Smad3-dependent manner, to indirectly enhance nearly more than 50% of migrated hBMSCs. In vivo, TGFß3 delivery improved MCP1 expression by nearly 7.9-fold, recruited approximately 2.0-fold CD31+ vascular cells and 2.0-fold Sca-1+ PDGFR-α+ MSCs, and achieved 2.5-fold bone volume fraction (BV/TV) and 2.0-fold bone mineral density, relative to TGFß3-free delivery. CONCLUSIONS: TGFß3, as a MSC homing molecule, recruited MSCs to initiate bone formation in the direct-dependent and indirect-dependent mechanisms. This may shed light on the improvement of MSC homing in bone regeneration.

A High-Adhesive Lysine-Cyclic RGD Peptide Designed for Selective Cell Retention Technology.

Cell adhesion is an important property of biomaterials used in selective cell retention (SCR) technology, which fabricates bone grafts rapidly in clinical settings. This could be improved by physical and biologic manipulations. To facilitate retention of the cells on the scaffold, especially osteoprogenitors from bone marrow in the convenient SCR procedure, a lysine-cyclic RGD (LcRGD) peptide was here designed to coordinate positively charged amino acids and the RGD sequence to enhance the adhesion performance of the scaffold. Demineralized bone matrix (DBM) is an important therapeutic resource, but its cell adhesion ability and osteoinductive capacity are low because of its processing. These capabilities can be increased to enhance the performance of DBM when used in SCR technology. Here, LcRGD peptide was used to modify DBM and produce a DBM/LcRGD composite. This composite exhibited enhanced adhesion performance on cultured human bone marrow-derived mesenchymal stem cells and retained more osteoprogenitors from bone marrow than other materials did. The DBM/LcRGD composite displayed a preferable osteoinduction in vitro and osteogenic capacity in vivo. Thus, LcRGD peptide as a commendable modifier of DBM applied in SCR technology can improve bone transplantation.

Sustained release of bioactive protein from a lyophilized tissue-engineered construct promotes the osteogenic potential of mesenchymal stem cells.

Tissue-engineered constructs (TECs) seeded with mesenchymal stem cells (MSCs) represent a therapy for large bone defects. However, massive cell death in TECs in the early postimplantation period prompted us to investigate the osteoinductive mechanism of TECs. Previous studies demonstrated that stem cell extracts retained equivalent levels of bioactive proteins and exhibited an osteoinductive nature similar to that of intact cells. These data led us to hypothesize that despite the massive cell death in TECs, devitalized MSC-derived proteins remain on the scaffolds and are released to improve cell function. Here, TECs were prepared using demineralized bone matrix seeded with human umbilical cord Wharton's jelly-derived MSCs (hWJMSCs), and the cells seeded in TECs were devitalized by lyophilizing the TECs. Scanning electron microscopy, BCA protein assays, quantitative cytokine array analysis and immunofluorescent staining indicated that approximately 3 mg/cm(3) of total protein and 49 types of cytokines derived from hWJMSCs were preserved in the lyophilized TECs (LTECs). The sustainable release of total protein and cytokines from LTECs lasted for more than 2 weeks. The released protein improved the osteogenic behavior of and gene expression in MSCs. Furthermore, the lyophilized hWJMSC-derived proteins had immunoregulatory properties similar to those of live MSCs in mixed lymphocyte reactions. Collectively, we present a novel perspective on the osteoinductive mechanism of TECs and introduce LTECs as new systems for delivering multiple cytokines to enhance MSC behavior.

Pitavastatin attenuates monocyte activation in response to orthopedic implant-derived wear particles by suppressing the NF-κB signaling pathway.

Aseptic loosening secondary to particle­induced periprosthetic osteolysis is considered to be the primary cause of long­term implant failure in orthopedic surgery. Implant­derived wear particles activate and recruit macrophages and osteoclasts, which cause a persistent inflammatory response with bone destruction that is followed by a loosening of the implant. Thus, strategies for inhibiting macrophage and osteoclast function may provide a therapeutic benefit for preventing aseptic loosening. The aim of the present study was to determine the effects of pitavastatin on the activation and cytokine response of polymethyl methacrylate (PMMA) particle­induced monocytes. Peripheral blood monocytes were obtained and treated with PMMA and pitavastatin. ELISA demonstrated that pitavastatin inhibited mRNA and protein expression of interleukin (IL)­1ß, IL­6 and tumor necrosis factor­α. Western blot analysis and immunofluorescence staining demonstrated that pitavastatin downregulated inhibitor of κB phosphorylation and degradation, and nuclear factor κ­light­chain­enhancer of activated B cells (NF­κB) p65 translocation. Together, these results indicate that pitavastatin may attenuate monocyte activation in response to orthopedic implant wear particles by suppression of the NF­κB signaling pathway.

Surgical treatment of lumbosacral tuberculosis by one-stage debridement and anterior instrumentation with allograft through an extraperitoneal anterior approach.

BACKGROUND: This study was aimed to investigate the clinical outcome of lumbosacral tuberculosis treatment by one-stage radical debridement with bone allograft reconstruction and anterior instrumentation via a retroperitoneal approach. METHODS: We retrospectively analyzed a series of 43 patients with lumbosacral tuberculosis in whom the lumbosacral junction was exposed via an anterior midline retroperitoneal approach. After radical debridement, two parallel tricortical iliac crest bone allografts were placed to reconstruct the anterior column, and then anterior fixation was performed. RESULTS: The mean follow-up period was 34 months (range, 24-91 months), during which no obvious loss of correction was observed. No case experienced recurrence, tuberculous peritonitis, erectile dysfunction, or retrograde ejaculation. CONCLUSIONS: The midline retroperitoneal approach provides direct and safe access to lesions of lumbosacral tuberculosis. Two parallel structural iliac crest allografts and anterior instrumentation effectively stabilize the lumbosacral junction.

Improving the osteogenic efficacy of BMP2 with mechano growth factor by regulating the signaling events in BMP pathway.

Local application of bone morphogenetic protein 2 (BMP2) is known to promote large bone defect healing and BMP2-initiated bone regeneration could be enhanced by an additional mechanical stimulation. The C-terminal 24-a.a. peptide of mechano growth factor (MGF24E), a mechanical-sensitive molecule, has been demonstrated to promote bone healing. Here, we propose a hypothesis that MGF24E could also improve the osteogenic efficacy of BMP2 by regulating the signaling events in the BMP pathway. To confirm the hypothesis, the potentials of MGF24E, BMP2 and BMP2/MGF24E combination treatments on the phosphorylation of Smad 1/5/8, the downstream osteogenesis-related gene expression and osteoblasts mineralization, are investigated with or without the blocking of Smad 5 siRNA. Furthermore, 15-mm rabbit radial bone defects were healed with the cytokine treatments and then evaluated by radiographic examination, histological assessment and immunohistochemical analysis. MGF24E could enhance the BMP2-induced Smad signaling pathway by upregulating the p-Smad protein expression and the downstream osteogenic gene expression. An amount of 5 nM BMP2 in a sub-25 nM concentration of MGF24E medium achieved a higher expression for ALP mRNA and a greater calcium mineral content compared with BMP2 alone. Nevertheless, the inhibition of the MGF24E-regulated BMP pathway could block osteogenesis induced by the dual treatment. In vivo, MGF24E treatment upregulated the endogenous BMP2 expression and the addition of MGF24E into the BMP2 treatment remarkably enhanced the bone mineral density (BMD), the radiographic scores and the histological restoration of the regenerated tissue against BMP2 treatment, suggesting a new strategy for BMP2 in bone defect healing.

The osteogenetic efficacy of goat bone marrow-enriched self-assembly peptide/demineralized bone matrix in vitro and in vivo.

In clinical practice, the prolonged duration, high cost, critical technique requirements, and ethical issues make the classical construction method of tissue-engineered bones difficult to apply widely. The major essentials in tissue engineering strategies include seed cells, growth factors, and scaffolds. This study aimed to incorporate these factors in a rapid and cost-effective manner. A self-assembly peptide/demineralized bone matrix (SAP/DBM) composite was artificially established and used for bone marrow enrichment via a selective cell retention approach. Then, goat mesenchymal stem cells (gMSCs) were seeded onto the SAP/DBM or DBM. The proliferation status of gMSCs in different scaffolds was analyzed, and the osteogenetic efficacy was evaluated after osteogenic induction. Bilateral critical-sized femoral defects (20-mm in length) were created in goats, and then the defects were implanted with the postenriched composite or DBM. Then, bone scan imaging, micro-computed tomography (CT) analysis and histological examination were performed to assess the reparative effects of the different implants. Compared with the DBM scaffolds, the growth of gMSCs in the postenriched SAP/DBM composite was faster and the expression levels of the osteo-specific genes (i.e., alkaline phosphatase, osteocalcin, osteopontin, and runt-related transcription factor 2) were significantly higher after 14 days of osteogenic induction. More importantly, the postenriched SAP/DBM composite significantly enhanced bone metabolic activity in the defect area compared with DBM at 2 and 4 weeks postoperation. Moreover, bone reconstruction was complete in marrow-enriched SAP/DBM composite, but not in the DBM. In addition, all of the osteo-related parameters, including the ratio of bone volume to total bone volume, bone mineral density, new trabecular number, and new trabecular thickness, were significantly higher in the marrow-enriched SAP/DBM than those in the DBM. These results indicated that the SAP/DBM composite held great potential for clinical applications; immediate implantation after marrow enrichment could be a new and effective strategy for treating bone defect.