Study Human Pancreatic Differentiation And Diabetes Prevention And Treatment By Genome Editing And Stem Cell Technologies

Diabetes mellitus is a metabolic disease with a high incidence worldwide.The patients suffer from chronic hyperglycemia caused by the deficiency or dysfunction of insulin-secreting pancreatic ? cells.Therefore,the generation of unlimited functional pancreatic ? cells in vitro is critical for pancreatic developmental biology and diabetes treatment.Recent researches have reported numerous promising research protocols,including the directed differentiation of human pluripotent stem cells(hPSCs)into pancreatic ?-like cells.By optimizing the directed differentiation system,we obtain a large number of functional pancreatic ?-like cells.These cells can highly express the pancreatic markers,contain the ultrastructure of insulin secretion vesicles,and secret insulin in response to the change of glucose level.And these pancreatic ? cells have the functions both in vitro and in vivo.Therefore,it can be widely used in disease modeling,drug development and cell therapyPrecise genome editing can help to construct disease modeling efficiently However,specific genome editing in hPSCs is still very time-consuming and labor-intensive.CRISPR-Cpfl is a recently reported genome editing enzyme.It has many unique properties,including shorter crRNA and low off-target rate.Therefore,it is very helpful to develop CRISPR-Cpfl-mediated genome editing in hPSCs.Chemical small molecules are easy to use,highly efficient and non-integrated.Therefore,they can be used to improve genome editing efficiency.Here,we constructed the CRISPR-Cpfl genome editing system in hPSCs,and developed a non-preference drug screening platform.Using this system,we performed a high-throughput chemical screening in hPSCs,and finally found two small molecules VE-822 and AZD-7762 that increased the CRISPR-Cpfl-mediated genome editing efficiency significantly.The combination of the CRISPR-Cpfl system and chemical small molecules provides a simple and effective method for precise and efficient genome editingIn this study,we combined the directed differentiation system of hPSCs into pancreatic ?-like cells and the CRISPR-Cpfl genome editing system to construct a CLEC16A knockout diabetes-related disease model.Next,we found that CLEC16A played a role in the human pancreatic differentiation and affected pancreatic ? cell functions.These results lay the critical foundation for further studying the role of CLEC16A in pancreatic ? cell differentiation and its mechanismsIn summary,the rapid development of stem cell technology,genome editing,biomedical engineering,high-throughput genetic and chemical screening,and bioinformatics holds great potentials in disease modeling,drug development and cell therapy.These technologies will help to answer basic biology questions and to promote the clinical transformation.