Multi-parameter Cardiomyocyte-based Biosensors And Their Applications In Pharmaceutical Cardiac Action And Toxicity Analysis

Cell-based biosensors use living cells as the sensitive elements, combining a sensor or transducer for detecting the intercellular and extracellular micro-environment and a variety of cellular physiological parameters under the external stimulus signal. Unlike other types of biosensors, the main feature of cell-based biosensor is using the living cells to sensitively sense external environment. Cell-based biosensor can be widely used in the research field of cell physiology, pharmacology, and toxicology with sensitive, dynamic and real-time, non-invasive, and label-free properties.In this dissertation, based on the theories of cell-based biosensor, we combined with information technology, sensor technology, and biotechnology to construct cardimyocyte-based biosensors and detection systems, which are applied in the analysis of drug cardiac safety. In this dissertation, an impedance sensor was designed to simultaneously detect the cardiomyocyte growth and beating status, and also a multi-function cell impedance sensor detection system was built for cardiomyocyte growth and beating status simultaneous detection; besides, an integrated cell-based biosensor was designed for recording extracellular potential and beating status, also an integrated multi-parameter detection system was built for extracellular potential and beating status simultaneous detection. In this study, we also employed these two types of cardimyocyte-based biosensors for the ion channel drug effects validation and related pharmaceutical cardiac safety analysis. Moreover beating pattern function of human induced pluripotent stem cell-derived cardiomyocyte-based biosensor was developed for hERG channel inhibitors specific recognition. Furthermore, the integrated cardiomyocyte-based biosensors was used for drug analysis. With the continuous development of cell-based biosensor technology, the cardiomocyte·based biosensor will be an important tool for the fast high-throughput analysis of drug cardiac safety and toxins.1. Proposing a detection method of cardiomyocyte beating status based on high-speed A/D conversion chip and FPGA signal processing technology, improving the detection speed of cell-based impedance biosensor system 1000 time than the ECIS-16 system, and enhance the accuracy to 0.05% for the simultaneous recording of cardiomyocyte growth and beating status.In vitro cultured cardiomyocytes can attach onto the sensor surface with the behaviors of attachment, growth, beating, death and so on. Traditional cell-impedance sensor can detect the behavior of these cells with low speed and low accuracy, cannot satisfy the demand of simultaneous recording of the cardiomyocyte growth and beating status. This paper designs a new high-speed signal acquisition and processing system, which is employed in monitoring the cardiomyocyte growth and beating status in parallel. By primary neonatal rat cardiomyocyte bioassays, the system is verified to contain the functions of detecting growth and beating status.2. Using microelectrode array and interdigitated electrodes to construct the integrated cell-based biosensor, based on the hardware and software filtering technology, recording of extracellular potential, beating status, and growth status simultaneously and parallelly.Cardiomyocytes have the special electric excitability and mechanical beating. Traditional cell-based biosensor usually has the problem of single parameter, and cannot simultaneously record the growth status, excitation-contraction coupling reaction and electric excitability activities. In order to fully reflect changes of the cardiomyocyte physiological parameters under pharmacology, this thesis designs an integrated multiparameter cell-based biosensor, based on the frequency difference of cardiomyocyte extracellular potential and cell impedance drive signal, employing the hardware and software filtering to realize simultaneously detection functions of extracellular potential, beating status, and growth status. The results determined the feasibility of this method.3. Studying the multiparameter cardiomyocyte-based biosensor for pharmaceutical cardiac action and toxicity assessment methods, verifying the feasibility of this cell-based biosensor.Cardio toxicity of drugs is the main reason for withdrawal and development termination of certain drugs, and thus it is particularly important in the pharmaceutical cardiac safety analysis prior to the clinical application. The traditional drug cardiac safety analysis methods had a contradiction between clinical relevance and experimental throughput. In this dissertation, primary cardiomyocyte and human induced pluripotent stem cell-derived cardiomyocyte build a cardiomyocytes-based biosensor tool for ion channel drug analysis, reflecting the drug effect. Besides, beating pattern function is developed to identify the hERG channel inhibitors, and the multi-parameter joint analysis function of integrated cardiomyocyte-based biosensor was used for cardiac pharmaceutical safety analysis.