Prognostic Selection of Osteogenic Stem Cells Using Label-Free Microfluidic Separation
Abstract
Topic Area(s): Tissue Regeneration – Research into innovative methods for developing biocompatible scaffolds and stem cell therapies for manufacturing and production of tissues. Current treatment options for difficult fractures rely on autografts, allografts, synthetic bone graft substitutes and U.S. Food and Drug Administration (FDA)-approved bone morphogenetic protein (BMP)-2 based product, Infuse. Severe shortage of autografts and allografts, donor site morbidity associated with autograft use, risk of disease transmission through allografts, lack of osteo-inductive potential of synthetic grafts, and reported disappointing outcomes of Infuse therapy, motivate the search for alternative treatments. Growing evidence emerging from preclinical data and clinical trials suggests that the synthetic bone graft substitutes, fortified with osteo-inductive progenitor stem cells, is emerging as the most promising therapeutic alternatives, since they can provide all of the three necessary factors for bone regeneration – mechanical strength, osteo-conductivity, and osteo-inductivity. Adipose derived stem cells (ADSCs) are gaining tremendous attention in this regard, since they can be isolated easily, in abundant numbers, in simple and comparatively less invasive procedures. However, unreliable bone-forming ability of ADSCs, in comparison with reliable osteo-inductive property of bone marrow derived mesenchymal stem cells (MSCs), has created serious hurdles in therapeutic utilization of ADSCs for bone regeneration. ADSCs are a mixture of at least four distinct populations (CD105+CD34-, CD105-CD34-, CD105+CD34+, and CD105-CD34+) that are dissimilar in cell size, shape, and BMP-responsiveness. Expression of CD105 correlates with high BMP-responsive phenotype and larger cell size, while expression of CD34 correlates with low BMP-responsive phenotype and smaller cell size. Contrary to the expectation that CD105+CD34- population displaying maximum BMP-responsiveness in vitro would also induce maximum bone formation in vivo, these cells do not induce any bone formation. Interestingly, a clonal population of MSCs isolated from the same inbred mouse strain induces robust bone formation in vivo, displays CD105-CD34- phenotype corresponding to the small cell sized morphology, and does not respond to BMPs in vitro. This suggests that CD105-CD34-ADSCs with smaller cell size are likely the cell type with true osteogenic potential. Fluorescence activated cell sorting (FACS) and magnetic activated cell sorting (MACS) have traditionally served as the means for stem cell purification. However, they rely on the binding of specific antibodies to the surface receptors of stem cells, which poses a significant danger of alteration of stem cells properties that are relevant for the transplant application. We therefore seek to develop a microfluidic isolation platform for the isolation of osteogenic ADSCs, without the need for surface markers. Microfluidic methods allow for isolation based on the inherent differences in cell properties, such as size, deformability, and subcellular electrophysiology. Aim 1 will optimize the microfluidic platform for isolation of the osteogenic population of ADSCs from Balb/c derived mixed ADSCs. Aim 2 will determine distribution of predictive markers for osteogenic MSCS (CDAM1) in different ADSCs population. To the best of our knowledge, there are no such methodologies available to orthopaedic surgeons and scientists in the country who are interested in using ADSCs therapeutically for bone regeneration and are aware that a mixed population of ADSCs is not known to have desired bone-forming property. Successful completion of this proposed project will create a roadmap for development of a therapeutic option for bone regeneration, utilizing “minimally manipulated,” osteogenic ADSCs. Thus, the project is not only scientifically innovative but also has remarkable therapeutic prospective.
Document Details
- Document Type
- DoD Grant Award
- Publication Date
- Mar 10, 2021
- Source ID
- W81XWH2010094
Entities
People
- Quanjun Cui
Organizations
- United States Army
- University of Virginia