Craniosynostosis and the role of stem/progenitor cells in premature suture fusion and accelerated bone formation
New Faculty II
Craniosynostosis, the premature fusion of one or more cranial sutures, is a relatively common malformation occurring in one in 2,000 live human births and represents a clinical condition where bone formation is accelerated. Restriction of normal brain growth caused by prematurely fused sutures can lead to significant complications, and affected children face complex surgical remodeling of the skull to prevent functional and anatomical deficits. Furthermore, recent evidence suggests current surgical therapies are not entirely effective, thereby necessitating additional surgeries. Suture fusion, including that in nonsyndromic craniosynostosis (NSC), is controlled by a complex interaction between many different types of cells, tissue types, and growth factors. Enhanced bone formation during suture fusion may be related to increased local concentrations of tissue-produced bone-inducing factors (proteins) stemming from deficiencies in the local extracellular matrix (ECM) to appropriately bind and sequester these factors. New treatment strategies designed to control local concentrations of these factors employ the localized application of inhibitors of bone formation, yet this approach lacks long term effectiveness due to difficulties in delivering sufficient concentrations. Coupled with current surgical approaches, this strategy will likely only delay recurrence of suture fusion. Moreover, this approach does not take into consideration the impact of the extracellular matrix (ECM) and its contributions to protein signaling. Therefore, this proposal seeks to generate a carefully engineered, highly tunable ECM capable of binding locally produced bone-inducing proteins while providing a viable graft alternative to native bone tissue. To fulfill this goal, we will complete the following tasks: 1) Characterize the natural ECM produced by mesenchymal stem cells (MSCs) from patients diagnosed with NSC in each fused human suture and, upon comparing it to ECM produced by healthy human subjects, determine targets for engineering this matrix; 2) Modify the composition of ECM produced by MSCs from NSC patients to mimic the ECM composition produced by MSCs of healthy human patients; 3) Determine the response of bone-forming cells from patients affected by NSC to bone-inducing proteins when seeded on engineered graft biomaterials coated with modified ECMs; and 4) Determine the capacity of engineered biomaterials to modulate bone formation in a developing suture using an in vivo bone defect model. Successful completion of these proposed aims will yield several important outcomes that may lead to more effective strategies to reverse the damaging effects of premature fusion, provide new approaches to treat conditions where accelerated bone formation would be advantageous, and greatly facilitate the PI’s long-term goal of developing effective strategies for controlling stem cell fate with instructive biomaterials for tissue repair.
Statement of Benefit to California:
Congenital bone abnormalities, bone loss, and defects resulting from trauma pose a significant health problem that impacts individuals across the lifespan. Conventional therapies for bone-related abnormalities commonly require the grafting of bone segments into the defect (more than 500,000 procedures annually), yet a lack of sufficient material often precludes such therapies. Moreover, greater than 10% of all bone defects are nonhealing, with an even higher prevalence of nonunions in the elderly. Given that at least 20% of California’s population will be over the age of 65 by 2025, it is imperative that new approaches to bone repair are developed. This proposal has two primary goals. First, we seek to develop a novel approach for modulating bone repair using mesenchymal stem cells (MSCs) derived from patients diagnosed with nonsymptomatic craniosynostosis (NCS), a condition resulting in accelerated cranial suture fusion, by modulating the cell-produced local microenvironment. Second, we propose to exploit knowledge gained from the basic studies using these cells, tissues, and engineered biomaterials to control the rate of bone formation in vivo. Successfully achieving these primary goals will benefit the State of California and its citizens in several ways. Our findings may provide a novel means to treat children diagnosed with NSC by developing a novel bone graft. Clinical utilization of this system could markedly lower the recurrence of suture fusion, reduce the need for repeated surgical procedures, and conceivably improve the quality of life for these patients. These approaches could also have value in other health conditions where accelerated bone formation is warranted including osteogenesis imperfecta and osteoporosis. Also, the versatile technology developed here will have applications to other tissue engineering approaches that could benefit the biotechnology companies of California investing in regenerative medicine. Finally, we anticipate that the proposed studies will not only broaden the PI’s current research approaches to tissue repair by enhancing his development as an independent stem cell researcher, but will also directly benefit young scientists and researchers in training among the respective collaborating laboratories. Exposure of students to novel stem cell-related research may provide the greatest benefit to California by inspiring future leaders in science to pursue their research efforts within the state or develop products and therapies at California-based biotechnology companies.
This proposal focuses on craniosynostosis, premature fusion of the cranial sutures in infants (the tissue that holds the bones of the skull together, allowing for growth of the skull). The applicant has identified mesenchymal stem cell interactions with the extracellular matrix, and inappropriate activation signals used in bone formation as potential causes of this pathology. The applicant proposes to isolate and characterize the matrix secreted by mesenchymal stem cells from patients with craniosynostosis vs. normal patients, to manipulate (down-regulate) genes that are potential candidates for the abnormal behavior, and to determine the relative ability of mesenchymal stem cells from craniosynostosis patients vs. normal patients to generate bone on engineered scaffold matrices and in a rodent bone defect model. The major strengths of the proposal were thought to be the novelty of the disease model, and the potential for further understanding the role mesenchymal stem cells play in generating and modulating the extracellular matrix environment in general, a role that has broad implications for mesenchymal stem cell-mediated therapies. The quick transition to an in vivo model was applauded but reviewers suggested that pilot in vivo studies should have been part of the research design before proceeding directly to expensive, large-scale in vivo studies. The mentors named for the project had diverse expertise, including an expert in stem cell biology. However, there was no letter in the application from the stem cell mentor, which would have considerably improved the application. The major weaknesses cited by reviewers were the lack of alternative hypotheses to the mechanism which centered on the signaling pathways discussed in the proposal, the measurement of candidate factors at the message level (analysis of RNA) when the more appropriate measurements may be at the protein level, and the lack of plans for classical stem cell studies including clonal analysis. Reviewers were concerned that only one isolate of the stem cells was presented in the grant, and in this single isolate, the cells were insufficiently characterize to justify their use as the main reagent in the research. Overall there was not sufficient data to determine whether the assays proposed were justified, and reviewers had serious concerns about the project’s feasibility. The applicant brings doctoral training in chemical engineering and post-doctoral training in tissue engineering into stem cell biology, which is a strength of the application. Reviewers noted great potential in bringing this engineering expertise into stem cell biology. The applicant presents a well-developed career development plan. The applicant has not yet had independent first or senior author papers published and so is just at the very start of being an independent investigator. The applicant institution has built a strong stem cell program, and is providing the appropriate support, space, and intellectual resources to assure success of the project.