TISSUE TESTING TECHNOLOGIES LLC — Department of Health and Human Services SBIR Phase I: NHLBI

Description

ABSTRACT This proposal focuses on translation of ice free cryopreservation by vitrification employing a novel approach of volumetric heating by nanowarming using Fe nanoparticles in an alternating electromagneticeldVitrificationsub zero storage below the glass transition temperature in aglassyrather than a crystalline frozen phaseis a form of cryopreservation that avoids ice formationVitri cation can be achieved by quickly cooling the material to cryogenic storage temperatureswhere ice cannot formVitri cation can be maintained at the end of the cryogenic protocol by quickly rewarming the tissue to temperatures above the temperatures where ice nucleation may occurThe magnitude of the rewarming rates necessary to maintain vitri cation is much higher than the magnitude of the cooling rates that are required to achieve it in therst placeThe most common approach to achieve the required cooling and rewarming rates is by convection based boundary warming in which the the specimenandapos s surface is exposed to a temperature controlled environmentsuch as a fluid bathDue to the underlying principles of heat transferthere is a size limit in the case of surface boundary heating beyond which crystallization cannot be prevented at the center of the specimenFurthermoredue to the underlying principles of solid mechanicsthere is also a size limit beyond which thermal expansion in the specimen can lead to structural damage and fracturesVolumetric heating by nanowarming during the rewarming phase of the cryogenic protocol can alleviate these size limitationsVitrification is already an important enabling approach for reproductive medicine with the potential to permit storage and transport of cellstissues and organs for a great variety of biomedical usesUnfortunatelypractical application of vitrification has been limited to smaller systems such as cells and thin tissues due to diffusive and phase change limitations that preclude use for blood vesselslarger tissues and organsTo circumvent this problem we demonstrated that nanowarming effectively rewarms blood vessels in our preliminary researchOur experiments demonstrated that this innovative rewarming technique rewarmed vitrified femoral and carotid arteries in volumes ranging fromtomL with retention of cell viability and physiologic functionHoweverwarming of thick arteries was suboptimalWe propose using large animal blood vesselmodels for further optimization and evaluation of nanowarmed vessels using a combination of in vitro and in vivo studiesIn Phasein a single specific aim we will optimize ice free vitrification of thick walled arteriesaorta and pulmonarywith a go no go objective of achieving andgtviability for progression to PhaseIn Phasespecific aimswe propose using porcine vascular models in a combination of ex vivo and in vivo studiesThe magnetic nanoparticles will be distributed around and within the internal spaces of vesselsThe large vessel lumen space makes them a good choice for optimization of vitrification and nanowarmingIn Aimwe will evaluate cryopreserved arteries after real time shippingcomparing methods and validating the transport conditions that are finally approved based upon absence of tissue crackingIn Aimwe will characterize the post ice free cryopreservation state of arteries preserved for at leastyearsIn additionduring this aim we will characterize the chemistry and biomaterial properties of ice free cryopreserved blood vesselsEffective vitrification will be evaluated using cryomacroscopy to detect ice formation and cryoprotectant residuals by Raman spectroscopyIn Aimwe will perform short term transplant studiesdaysin two porcine vascular modelsfemoral and pulmonary artery into the carotid and pulmonaryrespectivelyin order to validate our technology for a future Phase IIb SBIR proposal using clinically relevant preclinical non human primate models and human tissues NARRATIVE There are huge markets for researchdiagnostic and clinical applications of naturally occurring and engineered cellstissues and organsStrategic assessment of the field has identified the need for better preservation methods because freezing methods of cryopreservation have been shown to damage tissues and organs due to ice formationThis proposal focuses on nanowarming technology development for cryopreservation by vitrification of large volume blood vessels samplesNanowarming is required for viablefunctional preservation of blood vesselsThere are significant clinical needs for vascular grafts for dialysis as well as patients requiring peripheral and coronary bypass graftsAdvances in the preservation of tissues are also needed for trauma careparticularly to incorporate regenerative medicine products into strategic national stockpilesThis proposal combines the use of novel cryoprotectant formulations with magnetic nanoparticles and radiofrequencyinduced warming to warm optimally vitrifiedbankedlivingbiological materialsThese technologies could eventually impact hundreds of thousands of patients in North America annually if applied to tissuestissue engineered cellular constructs and one day organs