Relationship between paravertebral muscle function, pelvic incidence, and health-related quality of life in patients with degenerative spinal deformity.

BACKGROUND: Patients with degenerative spinal deformity often experience symptoms that seriously affect their quality of life, such as low back pain and dysfunction. This study aimed to investigate the relationship between paravertebral muscle function and pelvic incidence (PI) and their effect on health-related quality of life (HRQL) in patients with degenerative spinal deformity. METHODS: A total of 112 patients with degenerative spinal deformity in Southwest Hospital (Chongqing, China) were enrolled. They were divided into groups according to PI angle: high (PI > 60°, n = 37), normal (PI 50°-60°, n = 31), and low (PI < 50°, n = 44). Paravertebral muscle strength and endurance were assessed using the prone external fixation test frame. The sagittal vertical axis (SVA) was measured on X-rays of the spine in an anterolateral position, and all subjects were assessed with the Oswestry Disability Index (ODI), Roland-Morris questionnaire (RMQ), and 36-Item Short Form Health Survey (SF-36). Pearson or Spearman coefficients were used to assess the relationship of paravertebral muscle function with SVA, PI, and health-related quality of life. RESULTS: Maximal voluntary exercise (MVE) in the high-PI group was significantly lower than the MVE of both the normal- and low-PI groups (p < 0.05). There was no significant difference in MVE between the normal- and low-PI groups (p > 0.05). There was no significant difference in endurance time, SVA, ODI, RMQ, and SF-36 among the three groups. Paravertebral muscle MVE was negatively correlated with PI, SVA, ODI, and RMQ (r = - 0.193, - 0.210, - 0.283, - 0.277, p < 0.05). Endurance time of paravertebral muscle was also negatively correlated with SVA, ODI, and RMQ (r = - 0.200, - 0.420, - 0.348, p < 0.05) and positively correlated with SF-36 (r = 0.245, p < 0.05). In addition, paravertebral muscle MVE was positively correlated with the physical functioning score of the SF-36 (r = 0.251, p < 0.05), and the endurance time of paravertebral muscle was positively correlated with the physical functioning, physical role, bodily pain, and social function scores of the SF-36 (r = 0.342, 0.230, 0.209, 0.256, p < 0.05). CONCLUSIONS: High PI may serve as a risk factor for decreased paraspinal muscle strength in patients with degenerative spinal deformities. Early and targeted exercises focusing on paraspinal muscle strength and endurance could potentially be of positive significance in slowing down the progression of sagittal imbalance, alleviating functional disorders, and increasing health-related quality of life in patients with degenerative spinal deformity.

Dynamic chromatin accessibility landscapes of osteoblast differentiation and mineralization.

Bone acts as a self-healing organ, which undergoes continuous regeneration process that is tightly regulated by the cooperation of osteoclasts with the capability of bone resorption and osteoblasts with the capability of bone formation. Generally, bone marrow derived mesenchymal stem cells (BMSCs) differentiated to final osteoblasts have been considered as critical role in bone remodeling. In this regard, several transcription factors (TFs) whose binding sites are initially hidden deep within accessible chromatin that participate in modulating osteoblast differentiation and bone matrix mineralization. Then, it is necessary to explore further the dynamic changes about the epigenetic transcription machinery during osteoblastogenesis. Here, we performed the chromatin accessibility and transcriptomic landscape of osteoblast differentiation and mineralization by using transposase-accessible chromatin sequencing (ATAC-seq) and RNA sequencing (RNA-Seq). Our data found that global chromatin accessibility during osteoblastogenesis was extensively improved. Above this, it is shown that key target genes including Col6a3, Serpina3n, Ms4a4d, Lyz2, Phf11b and Grin3a were enriched in differential loci RNA-seq and ATAC-Seq peaks with continuous changed tendency during osteoblasts differentiation and mineralization. In addition, Analysis of Motif Enrichment (AME) was used to elucidate TFs which modulated these target genes. In this study, it was shown for the first time that these important TFs including MEF2A, PRRX1, Shox2 and HOXB13 could alter promoter accessibility of target genes during osteoblastogenesis. This helps us understand how TF binding motif accessibility influences osteoblast differentiation. In addition, it also suggests that modulating the chromatin accessibility of osteogenesis could be developed as the promising strategies to regulate bone regeneration.

The effects of back extensor strength in different body positions on health-related quality of life in patients with degenerative spinal deformity.

BACKGROUND: Degenerative spinal deformity (DSD) is believed to originate from degeneration of the discs and facet joints and vertebral wedging. Currently, the nosogeny of DSD is not yet fully clarified and there has been no systematic study on the impact of their lower back muscle strength on quality of life. OBJECTIVE: To determine the characteristics of back extensor strength (BES) in different body positions and examine their correlations with health-related quality of life (HQOL) in degenerative spinal deformity (DSD) patients. METHODS: Participants comprised 60 DSD patients and 40 healthy volunteers. Maximal isometric BES was evaluated by dynamometers with the subject in three different positions (standing, prone, sitting). The visual analogue scale (VAS) score, Oswestry Disability Index (ODI), Roland-Morris Disability Questionnaire (RMQ), and 36-item Short Form Health Survey (SF-36) score were used to evaluate patient HQOL. Correlations between the BES in different body positions and HQOL were analysed. RESULTS: The BES values in three body positions were significantly smaller in DSD patients than healthy subjects (P< 0.05). The standing BES was found to be negatively associated with ODI and RMQ (R= 0.313, p< 0.05 and R= 0.422, p< 0.01, respectively). A negative relationship between sitting BES and RMQ was also seen (R= 0.271, p< 0.05). In addition, the standing and prone BES were positively correlated with the physical functioning score of the SF-36 (R= 0.471, p< 0.01 and R= 0.289, p< 0.05, respectively), and the sitting BES was positively correlated with the role-physical score of the SF-36 (R= 0.436, p< 0.01). CONCLUSION: The results indicate that the back extensor muscle is compromised in DSD patients and there are differences in predicting the severity of disability and physical HQOL scores with BES in different positions. Standing BES was the most reliable contributor to HQOL among three body positions.

Discovery of multipotent progenitor cells from human induced membrane: Equivalent to periosteum-derived stem cells in bone regeneration.

Background: The periosteum stem cells (PSCs) plays a critical role in bone regeneration and defect reconstruction. Insertion of polymethyl methacrylate (PMMA) bone cement can form an induced membrane(IM) and showed promising strategy for bone defect reconstruction, the underlying mechanism remains unclear. Our study sought to determine whether IM-derived cells(IMDCs) versus PSCs have similar characteristics in bone regeneration. Methods: IM and periosteum were harvested from ten bone defect patients treated with PMMA, the IMDCs and PSCs were isolated respectively. Morphological, functional and molecular evaluation was performed and matched for comparison. Results: Both progenitor-like IMDCs and PSCs were successfully isolated. In vitro, we found IMDCs were similar to PSCs in morphology, colony forming capacity and expression of surface marker(CD90+, CD73+, CD105+, CD34-/CD45-). Meanwhile, these IMSCs displayed multipotency with chondrogenic, adipogenic and osteogenic differentiation, but differed in some IMSCs(3/10) population showing relatively poor osteogenic differentiation. The molecular profiles suggests that cell cycle and DNA replication signaling pathways were associated with these varying osteogenic potential. In vivo, we established a cell-based tissue-engineered bone by seeding IMDSs/PSCs to demineralized bone matrix (DBM) scaffold and demonstrated both IMDSs and PSCs enhanced bone regeneration in SCID mice bone defect model compared with DBM alone. Conclusion: Our data demonstrated IM containing multipotent progenitor cells similar to that periosteum promoting bone regeneration, and indicated the existence of multiple subsets in osteogenic differentiation. Overall, the study provided a cellular and molecular insights in understanding the successful or failed outcome of bone defect healing.The translational potential of this article: This study confirmed IMDCs and PSCs share similar regeneration capacity and inform a translation potential of that cellular therapy applying IMDCs in bone defect repair.

Bile acid metabolism regulatory network orchestrates bone homeostasis.

Bile acids (BAs), synthesized in the liver and modified by the gut microbiota, have been widely appreciated not only as simple lipid emulsifiers, but also as complex metabolic regulators and momentous signaling molecules, which play prominent roles in the complex interaction among several metabolic systems. Recent studies have drawn us eyes on the diverse physiological functions of BAs, to enlarge the knowledge about the "gut-bone" axis due to the participation about the gut microbiota-derived BAs to modulate bone homeostasis at physiological and pathological stations. In this review, we have summarized the metabolic processes of BAs and highlighted the crucial roles of BAs targeting bile acid-activated receptors, promoting the proliferation and differentiation of osteoblasts (OBs), inhibiting the activity of osteoclasts (OCs), as well as reducing articular cartilage degradation, thus facilitating bone repair. In addition, we have also focused on the bidirectional effects of BA signaling networks in coordinating the dynamic balance of bone matrix and demonstrated the promising effects of BAs on the development or treatment for pathological bone diseases. In a word, further clinical applications targeting BA metabolism or modulating gut metabolome and related derivatives may be developed as effective therapeutic strategies for bone destruction diseases.

Osteoclast-derived apoptotic bodies inhibit naive CD8+ T cell activation via Siglec15, promoting breast cancer secondary metastasis.

The bone microenvironment promotes cancer cell proliferation and dissemination. During periodic bone remodeling, osteoclasts undergo apoptosis, producing large numbers of apoptotic bodies (ABs). However, the biological role of osteoclast-derived ABs, which are residents of the bone-tumor niche, remains largely unknown. Here, we discover that AB-null MRL/lpr mice show resistance to breast cancer cell implantation, with more CD8+ T cell infiltrations and a higher survival rate. We uncover that the membranous Siglec15 on osteoclast-derived ABs binds with sialylated Toll-like receptor 2 (TLR2) and blocks downstream co-stimulatory signaling, leading to the inhibition of naive CD8+ T cell activation. In addition, our study shows that treatment with Siglec15 neutralizing antibodies significantly reduces the incidence of secondary metastases and improves the survival rate of mice with advanced breast cancer bone metastasis. Our findings reveal the immunosuppressive function of osteoclast-derived ABs in the bone-tumor niche and demonstrate the potential of Siglec15 as a common target for anti-resorption and immunotherapy.

Modulation of SIRT6 activity acts as an emerging therapeutic implication for pathological disorders in the skeletal system.

The skeletal system is a dynamically balanced system, which undergoes continuous bone resorption and formation to maintain bone matrix homeostasis. As an important ADP-ribosylase and NAD+-dependent deacylase, SIRT6 (SIR2-like protein 6) is widely expressed on various kinds of bone cells, such as chondrocytes, osteoblasts, osteoclasts. The aberration of SIRT6 impairs gene expression (e.g., NF-κB and Wnt target genes) and cellular functions (e.g., DNA repair, glucose and lipid metabolism, telomeric maintenance), which disturbs the dynamic balance and ultimately leads to several bone-related diseases. In this review, we summarize the critical roles of SIRT6 in the onset and progression of bone-related diseases including osteoporosis, osteoarthritis, rheumatoid arthritis, and intervertebral disc degeneration, as well as the relevant signaling pathways. In addition, we discuss the advances in the development of SIRT6 activators and elucidate their pharmacological profiles, which may provide novel treatment strategies for these skeletal diseases.

PTP1B knockdown alleviates BMSCs senescence via activating AMPK-mediated mitophagy and promotes osteogenesis in senile osteoporosis.

The senescence of bone marrow mesenchymal stem cells (BMSCs) is the basis of senile osteoporosis (SOP). Targeting BMSCs senescence is of paramount importance for developing anti-osteoporotic strategy. In this study, we found that protein tyrosine phosphatase 1B (PTP1B), an enzyme responsible for tyrosine dephosphorylation, was significantly upregulated in BMSCs and femurs with advancing chronological age. Therefore, the potential role of PTP1B in BMSCs senescence and senile osteoporosis was studied. Firstly, significantly upregulated PTP1B expression along with impaired osteogenic differentiation capacity was observed in D-galactose (D-gal)-induced BMSCs and naturally-aged BMSCs. Furthermore, PTP1B silencing could effectively alleviate senescence, improve mitochondrial dysfunction, and restore osteogenic differentiation in aged BMSCs, which was attributable to enhanced mitophagy mediated by PKM2/AMPK pathway. In addition, hydroxychloroquine (HCQ), an autophagy inhibitor, significantly reversed the protective effects from PTP1B knockdown. In SOP animal model, transplantation of LVsh-PTP1B-transfected D-gal-induced BMSCs harvested double protective effects, including increased bone formation and reduced osteoclastogenesis. Similarly, HCQ treatment remarkably suppressed osteogenesis of LVsh-PTP1B-transfected D-gal-induced BMSCs in vivo. Taken together, our data demonstrated that PTP1B silencing protects against BMSCs senescence and mitigates SOP via activating AMPK-mediated mitophagy. Targeting PTP1B may represent a promising interventional strategy to attenuate SOP.

The impact of methicillin resistance on clinical outcome among patients with Staphylococcus aureus osteomyelitis: a retrospective cohort study of 482 cases.

This study was designed to evaluate the impact of methicillin resistance on the outcomes among patients with S. aureus osteomyelitis. We reviewed all extremity osteomyelitis patients treated in our clinic center between 2013 and 2020. All adult patients with S. aureus pathogen infection were included. Clinical outcome in terms of infection control, length of hospital stay, and complications were observed at the end of a 24-month follow-up and retrospectively analyzed between populations with/without methicillin resistance. In total, 482 osteomyelitis patients due to S. aureus were enrolled. The proportion of methicillin-resistant S. aureus (MRSA) was 17% (82) and 83% (400) of patients had Methicillin-sensitive S. aureus (MSSA). Of 482 patients, 13.7% (66) presented with infection persistence after initial debridement and antibiotic treatment (6 weeks), needed repeated debridement, 8.5% (41) had recurrence after all treatment end and a period infection cure, complications were observed in 17 (3.5%) patients (pathologic fracture; 4, nonunion; 5, amputation; 8) at final follow-up. Following multivariate analysis, we found patients with S. aureus osteomyelitis due to MRSA are more likely to develop a persistent infection (OR: 2.26; 95% CI 1.24-4.13) compared to patients with MSSA. Patients infected with MRSA also suffered more complications (8.5% vs. 2.5%, p = 0.015) and longer hospital stays (median: 32 vs. 23 days, p < 0.001). No statistically significant differences were found in recurrence. The data indicated Methicillin resistance had adverse clinical implication for infection persistence among patients with S. aureus osteomyelitis. These results will help for patients counsel and preparation for treatment.

Gold-nanosphere mitigates osteoporosis through regulating TMAO metabolism in a gut microbiota-dependent manner.

Osteoporosis (OP) is a metabolic bone disease characterized by decreased bone mass and increased bone fragility. The imbalance of bone homeostasis modulated by osteoclasts and osteoblasts is the most crucial pathological change in osteoporosis. As a novel treatment strategy, nanomedicine has been applied in drug delivery and targeted therapy due to its high efficiency, precision, and fewer side effects. Gold nanospheres (GNS), as a common kind of gold nanoparticles (GNPs), possess significant antimicrobial and anti-inflammatory activity, which have been applied for the treatment of eye diseases and rheumatoid arthritis. However, the effect of GNS on osteoporosis remains elusive. In this study, we found that GNS significantly prevented ovariectomy (OVX)-induced osteoporosis in a gut microbiota-dependent manner. 16S rDNA gene sequencing demonstrated GNS markedly altered the gut microbial diversity and flora composition. In addition, GNS reduced the abundance of TMAO-related metabolites in OVX mice. Low TMAO levels might alleviate the bone loss phenomenon by reducing the inflammation response. Therefore, we investigated the alteration of cytokine profiles in OVX mice. GNS inhibited the release of pro-osteoclastogenic or proinflammatory cytokines including tumor necrosis factor α (TNF-α), interleukin (IL)-6, and granulocyte colony-stimulating factor (G-CSF) in the serum. In conclusion, GNS suppressed estrogen deficiency-induced bone loss by regulating the destroyed homeostasis of gut microbiota so as to reduce its relevant TMAO metabolism and restrain the release of proinflammatory cytokines. These results demonstrated the protective effects of GNS on osteoporosis as a gut microbiota modulator and offered novel insights into the regulation of the "gut-bone" axis.

Long non-coding RNA HCAR promotes endochondral bone repair by upregulating VEGF and MMP13 in hypertrophic chondrocyte through sponging miR-15b-5p.

Endochondral bone formation is an important route for bone repair. Although emerging evidence has revealed the functions of long non-coding RNAs (lncRNAs) in bone and cartilage development, the effect of lncRNAs in endochondral bone repair is still largely unknown. Here, we identified a lncRNA, named Hypertrophic Chondrocyte Angiogenesis-related lncRNA (HCAR), and proved it to promote the endochondral bone repair by upregulating the expression of matrix metallopeptidase 13 (Mmp13) and vascular endothelial growth factor α (Vegfa) in hypertrophic chondrocytes. Lnc-HCAR knockdown in hypertrophic chondrocytes restrained the cartilage matrix remodeling and decrease the CD31hiEmcnhi vessels number in a bone repair model. Mechanistically, we proved that lnc-HCAR was mainly enriched in the cytoplasm using fluorescence in situ hybridization (FISH) assay, and it acted as a molecular sponge for miR-15b-5p. Further, in hypertrophic chondrocytes, lnc-HCAR competitively bound to miR-15b-5p to increase Vegfa and Mmp13 expression. Our results proved that lncRNA is deeply involved in endochondral bone repair, which will provide a new theoretical basis for future strategies for promoting fracture healing.

Klotho upregulates the interaction between RANK and TRAF6 to facilitate RANKL-induced osteoclastogenesis via the NF-κB signaling pathway.

BACKGROUND: α-Klotho (Klotho) plays a wide range of roles in pathophysiological processes, such as low-turnover osteoporosis observed in klotho mutant mice (kl/kl mice). However, the precise function and underlying mechanism of klotho during osteoclastogenesis are not fully understood. Here, we investigated the effects of klotho on osteoclastogenesis induced by receptor activator of nuclear factor kappa-B ligand (RANKL). METHODS: The effects of klotho deficiency on osteoclastogenesis were explored using kl/kl mice both in vivo and in vitro. In in vitro experiments, lentivirus transfection, real-time quantitative PCR (RT-qPCR) analysis, western blot analysis, immunostaining, RNA-seq analysis, differential pathway analysis, Energy-based protein docking analysis and co-immunoprecipitation were used for deeply investigating the effects of klotho on RANKL-induced Osteoclastogenesis and the underlying mechanism. RESULTS: We found that klotho deficiency impaired osteoclastogenesis. Furthermore, in vitro studies revealed that klotho facilitated osteoclastogenesis and upregulated the expression of c-Fos and nuclear factor of activated T cells cytoplasmic 1 (NFATc1) during osteoclastogenesis. Mechanistically, we confirmed that klotho co-localized with nuclear factor kappa B (RANK) and facilitated the interaction between activated RANK and TNFR-associated factor 6 (TRAF6), thus klotho exerts its function in osteoclastogenesis through the activation of the NF-κB signaling pathway. CONCLUSIONS: Klotho promotes RANKL-induced osteoclastogenesis through upregulating the interaction between RANK and TARF6, Targeting on klotho may be an attractive therapeutic method for osteopenic diseases.

Beyond immunosuppressive effects: dual roles of myeloid-derived suppressor cells in bone-related diseases.

Myeloid-derived suppressor cells (MDSCs) are a heterogeneous population of immature myeloid cells (IMCs) with immunosuppressive functions, whereas IMCs originally differentiate into granulocytes, macrophages, and dendritic cells (DCs) to participate in innate immunity under steady-state conditions. At present, difficulties remain in identifying MDSCs due to lacking of specific biomarkers. To make identification of MDSCs accurately, it also needs to be determined whether having immunosuppressive functions. MDSCs play crucial roles in anti-tumor, angiogenesis, and metastasis. Meanwhile, MDSCs could make close interaction with osteoclasts, osteoblasts, chondrocytes, and other stromal cells within microenvironment of bone and joint, and thereby contributing to poor prognosis of bone-related diseases such as cancer-related bone metastasis, osteosarcoma (OS), rheumatoid arthritis (RA), osteoarthritis (OA), and orthopedic trauma. In addition, MDSCs have been shown to participate in the procedure of bone repair. In this review, we have summarized the function of MDSCs in cancer-related bone metastasis, the interaction with stromal cells within the bone microenvironment as well as joint microenvironment, and the critical role of MDSCs in bone repair. Besides, the promising value of MDSCs in the treatment for bone-related diseases is also well discussed.

Connective Tissue Growth Factor From Periosteal Tartrate Acid Phosphatase-Positive Monocytes Direct Skeletal Stem Cell Renewal and Fate During Bone Healing.

The periosteum is critical for bone healing. Studies have shown that the periosteum contains periosteal stem cells (PSCs) with multidirectional differentiation potential and self-renewal ability. PSCs are activated in early fracture healing and are committed to the chondrocyte lineage, which is the basis of callus formation. However, the mechanism by which PSCs are activated and committed to chondrocytes in bone regeneration remains unclear. Here, we show that tartrate acid phosphatase (TRAP)-positive monocytes secrete CTGF to activate PSCs during bone regeneration. The loss function of TRAP-positive monocytes identifies their specific role during bone healing. Then, the secreted CTGF promotes endochondral ossification and activates PSCs in mouse bone fracture models. The secreted CTGF enhances PSC renewal by upregulating the expression of multiple pluripotent genes. CTGF upregulates c-Jun expression through αVß5 integrin. Then, c-Jun transcription activates the transcription of the pluripotent genes Sox2, Oct4, and Nanog. Simultaneously, CTGF also activates the transcription and phosphorylation of Smad3 through αVß5 integrin, which is the central gene in chondrogenesis. Our study indicates that TRAP-positive monocyte-derived CTGF promotes bone healing by activating PSCs and directing lineage commitment and that targeting PSCs may be an effective strategy for preventing bone non-union.

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.

[Effect of demineralized bone matrix modified by laminin α4 chain functional peptide on H-type angiogenesis and osteogenesis to promote bone defect repair].

OBJECTIVE: Based on the cell-extracellular matrix adhesion theory in selective cell retention (SCR) technology, demineralized bone matrix (DBM) modified by simplified polypeptide surface was designed to promote both bone regeneration and angiogenesis. METHODS: Functional peptide of α4 chains of laminin protein (LNα4), cyclic RGDfK (cRGD), and collagen-binding domain (CBD) peptides were selected. CBD-LNα4-cRGD peptide was synthesized in solid phase and modified on DBM to construct DBM/CBD-LNα4-cRGD scaffold (DBM/LN). Firstly, scanning electron microscope and laser scanning confocal microscope were used to examine the characteristics and stability of the modified scaffold. Then, the adhesion, proliferation, and tube formation properties of CBD-LNα4-cRGD peptide on endothelial progenitor cells (EPCs) were detected, respectively. Western blot method was used to verify the molecular mechanism affecting EPCs. Finally, 24 10-week-old male C57 mice were used to establish a 2-mm-length defect of femoral bone model. DBM/LN and DBM scaffolds after SCR treatment were used to repair bone defects in DBM/LN group ( n=12) and DBM group ( n=12), respectively. At 8 weeks after operation, the angiogenesis and bone regeneration ability of DBM/LN scaffolds were evaluated by X-ray film, Micro-CT, angiography, histology, and immunofluorescence staining [CD31, endomucin (Emcn), Ki67]. RESULTS: Material related tests showed that the surface of DBM/LN scaffold was rougher than DBM scaffold, but the pore diameter did not change significantly ( t=0.218, P=0.835). After SCR treatment, DBM/LN scaffold was still stable and effective. Compared with DBM scaffold, DBM/LN scaffold could adhere to more EPCs after the surface modification of CBD-LNα4-cRGD ( P<0.05), and the proliferation rate and tube formation ability increased. Western blot analysis showed that the relative expressions of VEGF, phosphorylated FAK (p-FAK), and phosphorylated ERK1/2 (p-ERK1/2) proteins were higher in DBM/LN than in DBM ( P<0.05). In the femoral bone defect model of mice, it was found that mice implanted with DBM/LN scaffold had stronger angiogenesis and bone regeneration capacity ( P<0.05), and the number of CD31 hiEmcn hi cells increased significantly ( P<0.05). CONCLUSION: DBM/LN scaffold can promote the adhesion of EPCs. Importantly, it can significantly promote the generation of H-type vessels and realize the effective coupling between angiogenesis and bone regeneration in bone defect repair.