The Construction Of Micro-nano Bacteria,Virus And Protein Biotherapy Systems And Their Application Research

Cell,bacteria,virus and protein mediated biotherapies are widely used in biomedical fields.However,there are some defects in the process of application,such as insufficient activity,easy degradation,and difficulty in evaluating the distribution in vivo,which greatly reduce the therapeutic effects.The introduction of micro-nano carrier technology can realize the encapsulation of biological agents,avoid their degradation and maintain activity,which can fully compensate for the problems,and is expected to achieve high efficient biotherapy.In this paper,we took cyanobacteria,oncolytic virus and photosensitive protein as examples,micro-nano carrier technology was introduced to construct micro-nano cyanobacteria biotherapy system,nanooncolytic virus biotherapy system and micro-photosensitive protein biotherapy system respectively,and explore their applications in tumor therapy and neuroregulation.The specific research contents are as follows:1.Cyanobacteria bio-oxygen pump mediated biotherapy for enhancing photodynamic therapy of hypoxic tumors.Photodynamic therapy(PDT)can inhibit the occurrence and development of tumor by catalyzing the transformation of oxygen into cytotoxic reactive oxygen species(ROS).However,the hypoxic microenvironment of tumor and oxygen consumption during photodynamic therapy(PDT)limit the therapeutic effects of PDT.In order to relieve hypoxia at the tumor site and improve the effect of PDT,a biooxygen-pump based on synechococcus elongatus UTEX 2973(S.2973)was constructed in this chapter.S.2973 produced oxygen under 640 nm laser irradiation(the oxygen content increased by 26% after irradiation for 5 min).Mesoporous silica nanoparticles loaded indocyanine green(MSN-ICG)and S.2973 were encapsulated in an injectable alginate hydrogel to prepare ALG-MI-S2973.The activity of S.2973 in ALG-MI-S2973 was not affected.Moreover,the dual effects of hydrogel and MSN could realize the slow release of ICG,thus ensuring the long-term treatment effect.In addition,in vitro experiments showed that ALG-MI-S2973 could induce more tumor cell deaths under the co-irradiation of 640 nm and 808 nm lasers(ALG-MIS2973+640+NIR)in comparison with single laser irradiation.Through the in situ injection,the hydrogel immobilized S.2973 at the tumor site without spreading.In addition,the ROS generated during the PDT could kill S.2973,ensuring the safety of this biotherapy.During the treatment process,the use of lasers with different wavelengths ensured the stable operation of the system.In addition,the tumor inhibition rate of ALG-MI-S2973+640+NIR was almost 100%.In this chapter,a new photo-oxygen biotherapy was proposed,which provided a new treatment strategy for PDT of hypoxic tumors.2.Oncolytic adenovirus-mediated biotherapy for anti-tumor research via nanovesiclesOncolytic virus can achieve specific replication in cancer cells and induce infected cells dead,not in normal cells.Oncolytic adenovirus(OA)is a very attractive oncolytic "drug" due to its high stability of viral particles and potential cytolytic activity.The efficiency of intravenous oncolytic viruses is limited by the neutralization with antibodies in the blood.And OA administrated systematically is largely enriched in liver,thus resulting in hepatotoxicity.In order to improve the stability of OA in the blood circulation and reduce its accumulation in the liver,OA was wrapped with calcium phosphate(Cap-OA)through electrostatic interaction,and then encapsulated with liposome outer shell to prepare LC-OA,in which liposome and calcium phosphate were respectively used to achieve escape from the blood circulation and intracellular lysosome.In vitro experiments showed that LC-OA had good stability and viral activity,and it was specifically amplified in tumor cells.Compared with chemical drugs,LCOA had almost no damage to normal cells.Real-time fluorescent quantitative PCR(q PCR)experiments showed that the number of viral gene copies in the tumor tissue of LC-OA was significantly higher than that in the OA group.In addition,the number of viral copies of LC-OA in liver tissue was significantly reduced,thus reducing liver damage.LC-OA injected through the tail vein could reach the tumor tissue through the enhanced permeability and retention(EPR)effect,thereby inhibiting tumor growth(the tumor inhibition rate was about 89.1%).This work provides a new perspective for the combination of oncolytic virus biotherapy and nano carrier technology.3.Mechanoluminescent microdevices-photosensitive protein mediated biotherapy for neuromodulationPhotosensitive protein is a kind of ion channel protein that crosses the cell membrane.Under the action of a specific wavelength laser,the channels open and selectively pass cations or anions to activate or inhibit cell activities.Optogenetics is a science that uses light and genetic engineering to regulate the activity of neurons.This method can stimulate specific types of neurons with high precision,but wired operation limits its further applications.The development of microdevices and micro-nano carrier technology has brought much hope to the development of wireless optogenetics.In this chapter,mechanoluminescent microdevices were constructed based on mechanoluminescent materials,which were combined with photosensitive protein for neuromodulation.Mechanoluminescent materials can convert mechanical energy into light,which could be mixed with dimethylsiloxane(PDMS)to prepare mechanoluminescent microdevices.The microdevices could emit light in response to external wireless magnetic fields.At the same time,the external magnetic field rotation frequency could control the luminous intensity of the microdevices.The external magnetic field is not only a coded signal but also an output signal,which improves the mobility of the system.In vitro experiments showed that the microdevices had good luminescent stability and repeatability.In vivo experiments showed that the microdevice could activate neurons and regulate the rats’ movement behavior under the action of external magnetic field.This mechanoluminescent microdevicesphotosensitive protein-mediated biotherapy can achieve wireless regulation and high sensitive response,and provide an important reference for neuroregulation.In this paper,biotherapy is combined with micro-nano carrier technology.The advantages of the two methods are complementary to better meet the needs of biomedical applications.In this paper,three kinds of micro-nano biotherapy systems were constructed based on cyanobacteria,oncolytic adenovirus and photosensitive protein,which can effectively improve the activity of biological agents and the efficiency of disease prevention and treatment,and provide new ideas for the research and development of biological agents.