"Nano-optogenetic" System In Precisely Regulating Cellular Process And Its Application In Disease Therapy

Optogenetics is a novel interdiscipline which uses light to control biological process.Since firstly proposed in 2005,optogenetics was named "Method of the Year" by journal NATURE and SCIENCE in 2010.Although optogenetics started late in China,it is developing rapidly in recent years,catching up with the world class.At the fifth plenary session of the party’s 19,based on the big background of the COVID-19,the government firstly raised new direction for science and technology innovation: "for people’s life and health" and made a series of new strategic deployments.A series of China’s major scientific research achievements in recent years have been listed,including: "In the genetic components design of optogenetics,China has reached world leading level".Compared with traditional regulative methods using chemical reagents,heat or ultrasound as signal,light has few side effects on tissues and living objects.What’s more,light has accurate spatial-temporal specificity.This unique advantage makes light widely applied in neurosciences,signal pathways,disease therapy and other fields of biological process.However,owing to that external visible light cannot penetrate deep tissues,the light source of optogenetics mainly depends on optical fiber implantation in tissues.It’s difficult for traditional optical fiber to be implanted by operation,which is easily to cause the damage of tissues and even death.These drawbacks deeply limit the in vivo application of optogenetics.How to safely,efficiently and conveniently introduce light into the body determines whether optogenetics can be further transformed into biomedical engineering.In order to extend the in vivo applications of optogenetics and solve the problem of in vivo delivery of light,this project combines nanotechnology with optogenetic tools,establishing three "nano-optogenetic" systems which can be applied under skin,in tumor and in the gut.In these systems,the rare-earth upconversion materials can convert tissue-penetrating near-infrared light to local visible light,to avoid the damage of fiber implantation;To ensure the safety of in vivo application and avoid the toxicity of materials,flexible materials and hydrogel are used to encapsule the upconversion materials.These methods avoided the direct contact of rare-earth materials with tissue to safely,efficiently and conveniently introduce light into the body,including:(1)"nano-optogenetic" system is used to precisely regulate autophagy in vivo.Upconversion nanorod are packed to PET/PDMS basement and a flexible device with the thickness of 0.1-0.3mm are made by using high temperature drying method,which is then subcutaneously embedded to mice.The autophagy-related plasmids p53-Cry2 and NLS-Cib1 were co-transfected into subcutaneous tissues.By analyzing the autophagy level at various depth of tissue,we find that the "nano-optogenetic" system can induce 3.5-fold higher autophagy effect.(2)"nano-optogenetic" system for precise in vivo cancer treatment.The EL222 promoter modified Escherichia coli Nissle 1917 which secretes tumor necrosis factor TNFα upon blue light was injected to mice through the tail vein.The engineered bacteria could spontaneous targeting to the hypoxic region of the tumor and then colonize;Folic acid molecules were modified on the surface of the upconversion nanoparticles and injected into tumor-bearing mice through the tail vein.Experimental results showed that after stimulate by 3 times of the "nano-optogenetic" system,the tumor growth could be effectively inhibited.(3)"nano-optogenetic" system for precisely in vivo "gut-brain" axis regulation.Upon the "gut-brain" axis pathway,blue light-sensitive pDawn promoter was introduced to Lactococcus lactate NZ9000 chassis.Under the induction of blue light,the engineered bacteria could secret therapeutic factors such as GABA,Gcsf or GLP1,respectively.The strains with different functions were transplanted into the intestinal tract of mice;In order to achieve light delivery in the intestinal tract,the upconversion nanorods-encapsulated hydrogel were prepared based on UV curing method,which was then delivered orally to the intestinal tract of mice.After 980 nm near-infrared light stimulation in the gut,the experimental results showed that the "nano-optogenetic" system could precisely control the production of engineered bacteria in the gut,which then accurately regulate anxiety,Parkinson’s disease and the vagus nerve of mice.