[Projects to accelerate the practical use of innovative medical devices to collaborate with TWIns, Center for Advanced Biomedical Sciences, Waseda University and School of Engineering, The University of Tokyo].

Division of Medical Devices has been conducting the projects to accelerate the practical use of innovative medical devices to collaborate with TWIns, Center for Advanced Biomedical Sciences, Waseda University and School of Engineering, The University of Tokyo. The TWIns has been studying to aim at establishment of preclinical evaluation methods by "Engineering Based Medicine", and established Regulatory Science Institute for Medical Devices. School of Engineering, The University of Tokyo has been studying to aim at establishment of assessment methodology for innovative minimally invasive therapeutic devices, materials, and nanobio diagnostic devices. This report reviews the exchanges of personnel, the implement systems and the research progress of these projects.

[Identification of osteogenic signal and the development of artificial bones].

Mammalian skeletons are formed through two distinct processes, intramembranous ossification and endochondral ossification. During the endochondral ossification, indian hedgehog (Ihh) /parathyroid hormone-related protein (PTHrP) negative-feedback loop regulates the distance between epiphysis and the layer of hypertrophic chondrocytes. Ihh also stimulates bone formation. Based on such knowledge, bioactive molecule-loaded artificial bones are expected to achieve efficient bone regeneration without cell transplantation.

Ultrasound-mediated gene transfection in vitro: effect of ultrasonic parameters on efficiency and cell viability.

Ultrasound (US)-mediated gene transfection in the presence of microbubbles is a recently developed and promising non-viral gene delivery method. Optimising the parameters used in ultrasonic transfection is urgently required in order to realise higher transfection efficiencies in clinical settings. This study examined the effect of ultrasound exposure parameters on plasmid DNA transfection in mouse embryonic fibroblast cell lines using perfluorobutane bubbles. Variations in US intensity (0-11 W/cm2), pulse repetition frequency (PRF, 50-50,000 Hz), duty ratio (10 to 50%), exposure time (0-120 s) and microbubble volume concentration (0 to 10%) were tested, and the microbubble volume concentration was also monitored during exposure. Through the experiments, the mechanism of how variations in parameters influence US-mediated gene transfection was discussed, which can provide a basis for future applications of ultrasound mediated transfection.

[Custom-made artificial bones fabricated by an inkjet printing technology].

Although current treatment modalities for bone defects include autograft, allograft, and artificial bone substitutes, they have problems concerning invasiveness, safety, and performance, respectively, calling for development of innovative artificial bones with better handling and mechanical strength, better control of external and internal structures, and better biodegradability and osteo-inductive ability. We propose to fabricate novel high performance artificial bones using 3D inkjet printer based on the image data of bone deformity. Shape precisely fitting to the deformity, internal structure facilitating cell invasion, and good biodegradability are achieved. Bioactive substances can be incorporated by printing in combination with drug delivery system to induce bone regeneration at desired locations. These osteo-inductive artificial bones will help efficiently treat various types of bone deformity in a less invasive and safe manner.

[Optimization of signaling pathways for bone regeneration].

We have developed an ES cell-based monitoring system for osteogenic differentiation and identified a potent combination of osteogenic genes. ES cells were isolated from mice carrying a transgene expressing GFP driven by the 2.3 kb fragment of rat type I collagen alpha1 promoter. We have screened cDNA libraries and combinations of major osteogenesis-related genes and identified that the combination of constitutively active activin receptor-like kinase 6 (caALK6) and runt-related transcription factor 2 (Runx2) was the minimal unit that induced GFP. These combinations of genes would be a potent and ideal tool for bone regeneration.

[Regulation of chondrogenesis by PTH/PTHrP signaling].

During endo chondral bone development, parathyroid hormone-related peptide(PTHrP)is expressed by the perichondrial cells and proliferating chondrocytes in the periarticular region. PTHrP mRNA expression is increased by Indian Hedgehog(Ihh), which is expressed by prehypertrophic and hypertrophic chondrocytes. PTHrP acts directly on columnar proliferating chondrocytes to prevent hypertrophic differentiation. This prevention indirectly results in less Ihh signaling by decreasing the number of prehypertrophic and hypertrophic chondrocytes expressing Ihh. This negative feedback loop serves to regulate the balance of differentiation and proliferation of growth plate chondrocytes. Gs, cAMP, Sox9 and CREB seem to act downstream of PTHrP, while Patched, smoothened, Gli appear to act downstream of Ihh.