TheraCell Inc. proposes to develop a new device for the rapid isolation and electroporation of individual mesenchymal adult stem cells (MSCs), as part of its CellLabTM stem cell product suite, for application in spine fusion and other stem cell-based skeletal regeneration therapies. This stem cell processing device will consist of three critical modules, of which the electroporation module is one, and which is the motivation for this proposal. (The other two critical modules, also employing single cell manipulation techniques, are being developed by TheraCell under separate projects). Adult MSCs are pluripotent cells that can differentiate along osteogenic, adipogenic, myogenic, and other pathways, and represent a promising tool for cell and cell-based tissue engineering. We propose to develop the technology to select the individual MSCs, and then to perform single-cell gene-transfection (via electroporation) for up to ~107 individual MSCs in a very short period of time. The new single-cell electroporation method will result in a significantly more efficient and effective gene-transfection method compared with the bulk electroporation methods currently in use. In Phase I, we will design and fabricate a prototype microfluidic micro-electroporation chip, based on new MEMS (micro-electro-mechanical systems) technology. The chip will contain several test structures, including single-channel versions of the design, as well as a multi-channel version. The design will be developed in such as way as to lend itself easily for scale up to hundreds of channels required for the final product. The prototype micro-electroporation chips will be evaluated for basic fluid flow characteristics using fluorescently labeled microspheres, and then with stem cells that will be electroporated with a fluorescence protein for analysis using a fluorescence microscope. Once the optimum electroporation parameters are determined, the chip will be further evaluated with stem cells that have been electroporated with the BMP-2 protein, and those cells will be analyzed for osteogenic potential. In Phase II we will further optimize the design of the micro-electroporation chip, based on the Phase I results. The number of channels will be scaled up from a few to tens or hundreds of channels, to accommodate the large numbers of cells required for the targeted application. We will build a complete pre-production prototype single-cell micro-electroporation device. At the end of Phase II, we plan to demonstrate a completed prototype that can be rapidly commercialized as a stand-alone product for research applications, and also integrated into a complete stem cell processing device, which includes the other cell manipulation modules, for future clinical applications.
Public Health Relevance: We propose to develop a new device for the electroporation and efficient gene-transfection of individual mesenchymal adult stem cells (MSCs) for application in spine fusion and other skeletal regeneration therapies. Adult MSCs are pluripotent cells that represent a promising tool for cell and cell-based tissue engineering. The new single-cell electroporation method will result in a significantly more efficient and effective gene-transfection method compared with the bulk electroporation methods currently in use, for improved stem cell therapy outcomes.
Public Health Relevance Statement: Project narrative: Microfluidic Device for Efficient Gene-Transfection of Adult Stem Cells We propose to develop a new device for the electroporation and efficient gene-transfection of individual mesenchymal adult stem cells (MSCs) for application in spine fusion and other skeletal regeneration therapies. Adult MSCs are pluripotent cells that represent a promising tool for cell and cell-based tissue engineering. The new single-cell electroporation method will result in a significantly more efficient and effective gene-transfection method compared with the bulk electroporation methods currently in use, for improved stem cell therapy outcomes.
NIH Spending Category: Bioengineering; Biotechnology; Regenerative Medicine; Stem Cell Research; Stem Cell Research - Nonembryonic - Human
Project Terms: 21+ years old; Adipose tissue; Adult; Area; BMP-2; BMP-2A; BMP2; Biological Preservation; Body Tissues; Bone Marrow; Bone Regeneration; Cell Count; Cell Function; Cell Number; Cell Process; Cell Survival; Cell Viability; Cell physiology; Cells; Cellular Function; Cellular Physiology; Cellular Process; Characteristics; Collection; Development; Development and Research; Devices; Electroporation; Fatty Tissue; Fluorescence; Future; Gene Products, RNA; Genes; Glass; Human, Adult; Individual; Injection of therapeutic agent; Injections; Label; Lead; Liquid substance; Marketing; Measures; Mechanics; Medical center; Mesenchymal; Methods; Methods and Techniques; Methods, Other; Microbeads; Microfluidic; Microfluidic Device; Microfluidic Lab-On-A-Chip; Microfluidic Microchips; Microfluidics; Microspheres; Monitor; Mother Cells; Motivation; Organ; Outcomes of Therapy; Pathway interactions; Pb element; Phase; Polymers; Population; Power Sources; Power Supplies; Preparation; Preservation, Biologic; Preservation, Biological; Principal Investigator; Process; Product R; Production; Progenitor Cells; Programs (PT); Programs [Publication Type]; Property; Property, LOINC Axis 2; Protein Analysis; Proteins; Pump; R & D; R&D; RNA; RNA, Non-Polyadenylated; Research; Reticuloendothelial System, Bone Marrow; Ribonucleic Acid; Sampling; Series; Site; Source; Spinal Column; Spinal Fusion; Spine; Spondylosyndeses; Stem cells; Structure; Subcellular Process; Surgeon; Suspension substance; Suspensions; Syringes; System; System, LOINC Axis 4; Techniques; Technology; Testing; Time; Tissue Engineering; Tissue Sample; Tissues; Transfection; UV laboratory microscope; Ultraviolet Microscopes; V (voltage); Vertebral column; Work; adipose; adult human (21+); adult stem cell; backbone; base; bone morphogenetic protein 2; bone repair; cell preparation; clinical applicability; clinical application; design; designing; engineered tissue; fluid; fluid flow; fluorescence microscope; fluorescence/UV microscope; fluorescent microscope; gene product; heavy metal Pb; heavy metal lead; improved; laboratory fluorescence light microscope; liquid; osteogenic; pathway; preservation; programs; prototype; public health relevance; regenerate new tissue; regenerating damaged tissue; research and development; scaffold; scaffolding; scale up; skeletal regeneration; stem cell therapy; suspension; therapy outcome; tissue processing; tissue regeneration; tool; transgene expression; voltage; white adipose tissue; yellow adipose tissue
