Tumor Mouse Model Based On CRISPR/Cas9 Technology And The Role Of Carbon Nanomaterials In The Tumorigenesis Of Glioma

Glioma is the most common primary central nervous system tumor,accounting for about half of all intracranial primary tumors.According to the WHO standard,glioma can be classified into four grades from Ⅰ to Ⅳ as the degree of malignancy increases.More than half of all gliomas are glioblastoma,one of the most malignant human tumors.The survival of patients with glioblastoma is generally only about one year.At present,the treatments for brain tumors mainly include surgery,radiotherapy and chemotherapy,which fail to completely cure glioblastoma and are always harmful to patients.Scientific research has shown that glioma stem cells exist in gliomas,the existence of which is the cause of high recurrence rate and poor prognosis of malignant glioma.Based on this,the treatment strategy for glioma stem cells has become the key to glioma treatment.The establishment of an animal tumor model has brought new hopes for determining the cause and treatment of malignant tumors.The mouse tumor model can provide the growth environment and developmental microenvironment for human malignant tumors,which is of great significance for the study of tumor occurrence and development.The CRISPR/Cas9 is a gene editing system in which RNA directs Cas9 nuclease to specifically modify the DNA of interest.In this thesis,we established a LSL-Cas9-GFP mouse model using CRISPR/Cas9 technology,on which gene editing can be easily and efficiently performed,which therefore could lay a foundation for studying the mechanism of tumor development.Moreover,the application of carbon nanomaterials to study the effects of typical electrodeless nanomaterials on cancer stem cells has explored a new path for the treatment of tumors,providing a new direction for clinical treatment and application.In this thesis,we first constructed a time-controlled and space-controlled Cas9 transgenic mouse.Then,using target gene PCR amplification,stereotactic injection of virus and in vitro culture of stem cells for flow analysis,the mouse model with high expression of LSL system was screened out.Afterwards,The function of transgenic mice was identified by in vivo and in vitro introduction of sgRNA.Finally,we obtained two transgenic mouse models with high gene editing efficiency.Secondly,we used several bio-modified carbon nanomaterials to treat mouse fibroblasts,mouse neural stem cells and mouse glioma stem cells,and then test the activity of these cells.Among the 10 carbon dots,we found that one of them had different effects on the activity of the three cells at the same concentration.In order to understand the mechanism of this phenomenon,we will use cell proliferation,apoptosis,and RNA sequencing for further study.In summary,we used the technology of CRISPR/Cas9 to construct a transgenic mouse model with temporal and spatial regulation.Through screening and functional identification,we obtained a transgenic mouse model with high editing efficiency.We also found a carbon dot that has different effects on mouse neural stem cells and mouse glioblastoma stem cells through treating these cells with different carbon nanomaterials and evaluating their growth,which will provide possibilities for new materials and new ideas for subsequent cancer treatment.