EVALUATION AND MODULATION OF GRN GENE EXPRESSION VIA THE CRISPR/dCas9 SYSTEM IN iPSC-DERIVED ASTROCYTES AND iMGs FROM SCHIZOPHRENIC PATIENTS

Schizophrenia is a disease that affects about 23.6 million people worldwide. It is a multifactorial disorder, with genetic and environmental factors acting together, making individuals prone to develop this disease. Previous studies suggest that the GRN gene, which encodes a protein known as Progranulin, may be associated with an increased risk of developing schizophrenia. This exploratory study aims to evaluate and modulate the expression of the GRN gene using the CRISPR/dCas9 system in astrocytes and induced microglia-like cells (iMGs) derived from healthy (HC) and schizophrenic (SCZ) individuals. Peripheral Blood Mononuclear Cells were isolated from the blood of SCZ and HC individuals and differentiated into iMGs. Astrocytes were differentiated from Neural Stem Cells derived from induced Pluripotent Stem Cells generated from fibroblasts of HC and SCZ donors. To analyze GRN gene expression, iMGs were stimulated with LPS (100 ng/mL) and astrocytes with TNF-α (10 ng/mL) for 24 h, and the levels of Progranulin mRNA were measured via RT-qPCR. We employed both CRISPRa and CRISPRi systems, using the dCas9 protein fused with the transcriptional effectors VPR and KRAB, respectively, to modulate the transcription of GRN gene. Our data shows that iMGs derived from SCZ individuals express lower levels of Progranulin compared to those derived from HC individuals. On the other hand, astrocytes derived from SCZ individuals produced higher levels of Progranulin mRNA when stimulated with TNF-α compared to astrocytes from HC individuals. We also demonstrated the proper functioning of the CRISPR/dCas9 system in astrocytes. In conclusion, we observed that iMGs and astrocytes derived from SCZ individuals exhibit dysregulation in Progranulin expression levels. We can use the transcriptional regulation tools developed in this study to modulate Progranulin expression in cells derived from SCZ individuals, aiming to better understand the importance of this protein in schizophrenia.