Transient activation of notch signaling enhances endogenous stromal cell expansion and subsequent bone defect repair.

BACKGROUND: Following traumatic bone loss or removal of bone tumors, the failure of bone allograft transplantation for large bone defect repair remains a significant problem in orthopedics. Therefore, new strategies that can efficiently enhance allograft healing and long-term incorporation are critically needed. METHOD: In this study, we first injected Notch-activating Jagged1 peptide to mice and then isolated bone marrow tissues and cells for proliferation and differentiation assays. Femur bone allograft surgery was also performed in Jagged1 pre-treated mice, and bone defect healing process were monitored by histology, Micro-CT and biomechanical testing. RESULT: Our results showed that Jagged1 therapeutic injection is sufficient to maximally activate Notch and promote bone marrow stromal cell proliferation in vivo, while no effects on bone structure were observed. More importantly, Jagged1 pre-treatment significantly promoted bone callus formation and increased bone mechanical strength during allograft healing in a femur bone defect mouse model. CONCLUSION: This study reveals that Notch in vivo activation can be induced by injection of Jagged1 peptide for expansion of local native stromal cells that will significantly enhance bone callus formation. THE TRANSLATIONAL POTENTIAL OF THIS ARTICLE: The clinical uses of this therapeutic strategy would be immediately applicable for chronic long bone defect repair. More importantly, this devised strategy for expansion of endogenous BMSCs can also be applied to enhance other tissue and organ repair.

Notch ligand Jagged1 promotes mesenchymal stromal cell-based cartilage repair.

Placenta-derived mesenchymal stromal cells (PMSCs) provide a promising cell source for tissue regeneration. However, rapid induction of PMSC chondrogenic differentiation during therapeutic transplantation remains extremely challenging. Here we undertook a study to determine if Notch inhibition by soluble Jagged1 (JAG1) peptides could be utilized to accelerate PMSC-induced cartilage regeneration in a mouse post-traumatic osteoarthritis (PTOA) model. Our results showed that treatment of PMSCs with soluble JAG1 significantly enhanced chondrogenesis in culture as shown by increased alcian blue staining and decreased Notch target Hes1 expression when compared to those in lgG-treated control cells. Importantly, significantly enhanced cartilage formation and decreased joint inflammation were observed when JAG1-treated PMSCs were injected into mouse PTOA knee joints. Finally, in vivo cell tracing showed that more JAG1-treated PMSCs remained in knee joint tissues and that JAG1-treated PMSCs exhibited greater PMSC chondrogenic differentiation than lgG-treated control PMSCs at 4 weeks after injection. These data indicate that transient Notch inhibition by soluble JAG1 could be used to enhance PMSC survival and chondrogenic differentiation, thereby increasing the therapeutic potential of PMSCs for cartilage regeneration.

Human Mesenchymal Stromal Cell Sheet Enhances Allograft Repair in a Mouse Model.

To determine whether cell sheets generated with long-term passaged (P10) aging human mesenchymal stromal cells (MSCs) could be used for bone tissue regeneration as tissue engineered periosteum in a femoral allograft mouse model similar to fresh passaged (P3) young MSCs. At 3 weeks after transplantation of MSC sheets, results showed more bony callus formed between allograft and host bone ends in both young P3 MSC and aged P10 MSC sheet-wrapped groups when compared to allograft alone. At 6 weeks, while both MSC sheet-wrapped allografts showed more bony callus formation when compared to allograft alone groups, the bony callus size in aged P10 MSC sheet groups was significantly less than young P3 MSC sheet groups. Biomechanical testing confirmed that P3 MSC sheet-grafted femurs had the highest biomechanical strength in the three groups. Histology sections showed that the area of the chondriod callus in the aged P10 MSC sheet groups was significantly larger than in P3 MSC sheet groups. Finally, a significant increase of chondro-osteoclast activity was observed in the P3 MSC sheet-grafted femur. Our data demonstrates that extensive long-term culture-induced MSC aging impaired their osteogenic ability and subsequent bony callus formation, and could be used to induce cartilaginous callus formation.

Oleanolic Acid Enhances Mesenchymal Stromal Cell Osteogenic Potential by Inhibition of Notch Signaling.

Oleanolic acid (OA), a pentacyclic triterpenoid, has been shown to modulate multiple signaling pathways in a variety of cell linages. But the mechanisms underlying OA-mediated mesenchymal stromal cell (MSC) osteogenic differentiation are not known. In this study, we examined effects of OA on cell viability, osteogenic differentiation in MSCs, and the involvement of Notch and BMP signaling. OA induced bone marrow derived MSC differentiation towards osteoprogenitor cells and inhibited Notch signaling in a dose dependent manner. Constitutive activation of Notch signaling fully blocked OA induced MSC osteogenic differentiation. The expression level of early osteogenic marker genes, ALP, Runx2, and type I collagen, which play a critical role in MSC to osteoblast transition and servers as a downstream target of BMP signaling, was significantly induced by OA. Furthermore, BMP2 mediated MSC osteogenic differentiation was significantly enhance by OA treatment, indicating a synergistic effect between BMP2 and OA. Our results suggest that OA is a promising bioactive agent for bone tissue regeneration, and inhibition of Notch signaling is required for its osteogenic effects on MSCs.

Design and implementation of a system-based course in musculoskeletal medicine for medical students.

BACKGROUND: The amount of time devoted to musculoskeletal medicine in the typical undergraduate curriculum is disproportionately low compared with the frequency of musculoskeletal complaints that occur in a general practice. Consequently, whether because of the quantity or quality of the education, the competence level of graduating physicians regarding musculoskeletal problems is inadequate. Our purposes were to design a self-contained, system-based course in musculoskeletal medicine for medical students in the preclinical years and to measure the level of competence achieved by a class of first-year medical students who took the course. METHODS: The course was formulated by faculty from the departments of orthopaedic surgery, anatomy, and rheumatology and included elements of both objectives-based and problem-centered curricular models. The clinical lectures were preceded by pertinent anatomy lectures and dissections to provide a context for the clinical information. The lectures on basic science were designed to rationalize and explicate clinical practices. Small-group activities were incorporated to permit engagement of the students in critical thinking and problem-solving. A general musculoskeletal physical examination was taught in two two-hour-long small-group sessions with the orthopaedic residents serving as instructors. Cognitive competency was evaluated with use of comprehensive anatomy laboratory and written examinations, the latter of which included a validated basic competency examination in musculoskeletal medicine. Process-based skills were evaluated in the small-group meetings and in a timed, mock patient encounter in which each student's ability to perform the general musculoskeletal physical examination was assessed. RESULTS: The course lasted six weeks and consisted of forty-four lecture hours, seventeen hours of small-group meetings, and twenty-eight hours of anatomy laboratory. The average student score on the basic competency examination was 77.8%, compared with 59.6% for a historical comparison group (p < 0.05). Each student demonstrated the ability to adequately perform a general musculoskeletal physical examination in twenty minutes. The survey of student opinion after the course indicated a high level of student satisfaction. CONCLUSIONS: The main features of the course were: (1) an emphasis on both cognitive and process-based knowledge; (2) more contact hours and broader content than in previously described courses in musculoskeletal medicine; (3) the use of small groups to focus on problem-solving and physical examination competencies; (4) basic-science content directly related to clinical goals. These features might be used at other institutions that employ a system-based curriculum for the preclinical years to help improve competence in musculoskeletal medicine.