Final Report Summary - CARBONANOBRIDGE (Neuron Networking with Nano Bridges via the Synthesis and Integration of Functionalized Carbon Nanotubes)
With our work, we gained important insights into interfacing synthetic nano-materials to neuronal networks. Our results show that growing hippocampal neurons on a CNT scaffold increased their global synaptic connectivity. Our findings strongly indicate that true ex-novo formation of synaptic contacts is promoted by CNT, acting as unnatural biomimetic cues in cultured neuronal networks. The CARBONANObridge team demonstrated that CNT at the nanoscale instruct cell-specific behaviours in the hippocampal cultured networks.
We successfully implemented multiple research activities related to the general issue of CNT exploitation for organ repair, these are grouped in four major research streams: i. understanding the ability of CNT to transform spinal neuron development and signalling-we reported how carbon nanotube substrates alter the excitability and synaptic responses of brain neurons in culture. This observation highlighted the exceptional ability of this material in interfering with nerve tissue growth. Here we test the hypothesis that carbon nanotube scaffolds promote the development of immature spinal cord neurons. We performed electrophysiological studies associated, for the first time, to gene expression analysis. Our results indicate that spinal neurons plated on electro-conductive carbon nanotubes show a facilitated development. These changes are accompanied by a selective modulation of gene expression. Hence, future tissue scaffolds blended with conductive nanotubes may be exploited to promote cell differentiation and reparative pathways in neural regeneration strategies. ii. carbon based platforms interfacing anatomically intact structures such as spinal explants interfaced for weeks with carbon nanotube scaffolds to investigate whether and how CNT interactions are translated to nerve tissue explants. Long-term interfacing spinal explants to purified CNT induces two major effects: a. an increase in the number and length of neuronal fibers outgrowing the spinal segment, associated to changes in growth cone activity and in fiber elasto-mechanical properties; b. in spinal networks, synaptic currents display a significant increase in amplitude, indicating an augment in synaptic responses in intact tissue due to cell-substrate interactions. iii. carbon nanotube towards nanoscale interfaces for cardiac muscle cells for constructing nano-bridges for excitable tissues, this new concept is further sustained by an exciting series of experiments. We discovered that cardiac muscle cells improve their proliferative capacity, viability, growth and electrical properties when cultured on carbon nanotubes scaffolds. We further demonstrate by analyzing the gene expression program, that carbon nanotubes interacting with cardiomyocytes have the ability to promote physiological growth and functional maturation. These properties are unique in the current vexing field of tissue engineering, and offer unprecedented perspectives in the development of innovative therapies for cardiac repair.
iv. functionalization of CNT with fluorescing agents and bioactive moieties -our data reveal how CNT are able to translocate across cell membranes of both phagocytic and non-phagocytic cell lines, and, in at least 30-50% of cases, cells uptake CNT through an energy-independent mechanism. This characteristic makes nanotubes loaded with therapeutic or diagnostic cargos extremely interesting as the release of active molecules directly into the cytoplasm increase their biological activity and therapeutic efficacy. Additional studies highlighted how chemical functionalization of CNT provides useful tools in rendering CNT safer via altering their reactivity profile. In fact, tuning surface chemical functionalization of CNTs controls also their organ distribution and clearance in vivo and is crucial to the future design of any CNT based diagnostic or therapeutics.
We successfully implemented multiple research activities related to the general issue of CNT exploitation for organ repair, these are grouped in four major research streams: i. understanding the ability of CNT to transform spinal neuron development and signalling-we reported how carbon nanotube substrates alter the excitability and synaptic responses of brain neurons in culture. This observation highlighted the exceptional ability of this material in interfering with nerve tissue growth. Here we test the hypothesis that carbon nanotube scaffolds promote the development of immature spinal cord neurons. We performed electrophysiological studies associated, for the first time, to gene expression analysis. Our results indicate that spinal neurons plated on electro-conductive carbon nanotubes show a facilitated development. These changes are accompanied by a selective modulation of gene expression. Hence, future tissue scaffolds blended with conductive nanotubes may be exploited to promote cell differentiation and reparative pathways in neural regeneration strategies. ii. carbon based platforms interfacing anatomically intact structures such as spinal explants interfaced for weeks with carbon nanotube scaffolds to investigate whether and how CNT interactions are translated to nerve tissue explants. Long-term interfacing spinal explants to purified CNT induces two major effects: a. an increase in the number and length of neuronal fibers outgrowing the spinal segment, associated to changes in growth cone activity and in fiber elasto-mechanical properties; b. in spinal networks, synaptic currents display a significant increase in amplitude, indicating an augment in synaptic responses in intact tissue due to cell-substrate interactions. iii. carbon nanotube towards nanoscale interfaces for cardiac muscle cells for constructing nano-bridges for excitable tissues, this new concept is further sustained by an exciting series of experiments. We discovered that cardiac muscle cells improve their proliferative capacity, viability, growth and electrical properties when cultured on carbon nanotubes scaffolds. We further demonstrate by analyzing the gene expression program, that carbon nanotubes interacting with cardiomyocytes have the ability to promote physiological growth and functional maturation. These properties are unique in the current vexing field of tissue engineering, and offer unprecedented perspectives in the development of innovative therapies for cardiac repair.
iv. functionalization of CNT with fluorescing agents and bioactive moieties -our data reveal how CNT are able to translocate across cell membranes of both phagocytic and non-phagocytic cell lines, and, in at least 30-50% of cases, cells uptake CNT through an energy-independent mechanism. This characteristic makes nanotubes loaded with therapeutic or diagnostic cargos extremely interesting as the release of active molecules directly into the cytoplasm increase their biological activity and therapeutic efficacy. Additional studies highlighted how chemical functionalization of CNT provides useful tools in rendering CNT safer via altering their reactivity profile. In fact, tuning surface chemical functionalization of CNTs controls also their organ distribution and clearance in vivo and is crucial to the future design of any CNT based diagnostic or therapeutics.