Project description:Purpose: PDGF-D is a potent angiogenic and immune-related factor. It has been demonstrated that PDGF-D play roles in the pathological process of ocular angiogenesis and degenerative diseases. In our previous study, we established a retinal pigment epithelial (RPE) cell-specifc AAV-PDGF-D/GFP mouse model to study PDGF-D's role in age-related macular degeneration (AMD). In this study, we utilized the single-cell transcriptiome seqeuncing technique to explore the cell types of immune cells attracted by PDGF-D overepxression using the same animal models, with emphases on the impact of PDGF-D overexpression on individual cell's behaviour.
Project description:Wet age-related macular degeneration (AMD) is a progressive degenerative disease and a leading cause of blindness in elderly population. AMD pathogenesis initiated by several signals generates cellular and molecular cross-talk, including complement pathway-mediated inflammation, macrophage activation, and upregulation of angiogenic and cytokine/chemokine pathways. However, the mechanism by which the complement system is activated in AMD is not well understood, although its components are found in the neovascular lesions of wet AMD patients. Here, we show that increased PDGF-D expression engaged both classical and alternative complement pathways and markedly increased chemokine and cytokine responses to activate macrophages, thereby triggering neuroinflammatory milieu and exacerbating pathological neovascularization. Pharmacological targeting of the complement C3a receptor using SB290157 alleviated neuroinflammation by blocking macrophage polarization and by inhibiting pathological choroidal neovascularization (CNV). Our study thus suggests that therapeutic strategies targeting both PDGF-D and complement-mediated inflammatory signals may open up new possibilities for the treatment of neovascular diseases.
Project description:This is an ATAC sequencing experiment to explore chromatin accessibility change in mouse skeletal muscle treated with either AAV-GFP or AAV-CAAHR (a constitutively active mutant aryl hydrocarbon receptor). Mice received intramuscular injection of the AAV 5 months before the harvest of muscle.
Project description:Transcriptional profiling of human glioma cells comparing control GFP expressing cells with glioma cells transfected with a human PDGF-A gene. The isogenic cell lines were used to study the impact on glioma tumorigenesis and invasion. Goal was to determine the effects of PDGF-A gene transfection on global ES gene expression. Two set of glioma cell lines, LN444 vs LN444/PDGF-A and LN443 vs LN443/PDGF-A. Biological replicates: 2 control replicates, 2 transfected replicates.
Project description:Transcriptional profiling of human glioma cells comparing control GFP expressing cells with glioma cells transfected with a human PDGF-A gene. The isogenic cell lines were used to study the impact on glioma tumorigenesis and invasion. Goal was to determine the effects of PDGF-A gene transfection on global ES gene expression.
Project description:High-throughput sequencing of murine retina and RPE/Choroid (eye cup) tissue following mueller-cell mediated expression of either hVEGFA, hTNFa, or hIl6 by adeno-associated viruses (AAV)
Project description:AAV gene therapy has recently been approved for clinical use and shown to be efficacious and safe in a growing number of clinical trials. However, the safety of AAV as a gene therapy has been challenged by a few studies that documented hepatocellular carcinoma (HCC) after AAV gene delivery in mice. The association between AAV and HCC has been difficult to reconcile and is the subject of intense debate because numerous AAV studies have not reported toxicity. Here, we report a comprehensive study of HCC in a large number of mice following therapeutic AAV gene delivery. Using a sensitive high-throughput integration site-capture technique and global expressional analysis, we found that AAV integration into the Rian locus and the over-expression of a proximal gene, Rtl1, were associated with HCC. In addition, we identify a number of genes with differential expression that maybe useful in the study, diagnosis and treatment of HCC. We demonstrate that AAV vector dose, enhancer-promoter selection, and the timing of gene delivery are the defining factors in AAV-mediated insertional mutagenesis. Our results help explain the AAV-mediated genotoxicity previously observed and have important implications for the design of both safer AAV vectors and gene therapy studies. To investigate the possibility that insertional mutagenesis by AAV contributed to the development of HCC, we collected normal and tumor tissues from adult mouse livers that received AAV injection at a neonatal stage.
Project description:AAV gene therapy has recently been approved for clinical use and shown to be efficacious and safe in a growing number of clinical trials. However, the safety of AAV as a gene therapy has been challenged by a few studies that documented hepatocellular carcinoma (HCC) after AAV gene delivery in mice. The association between AAV and HCC has been difficult to reconcile and is the subject of intense debate because numerous AAV studies have not reported toxicity. Here, we report a comprehensive study of HCC in a large number of mice following therapeutic AAV gene delivery. Using a sensitive high-throughput integration site-capture technique and global expressional analysis, we found that AAV integration into the Rian locus and the over-expression of a proximal gene, Rtl1, were associated with HCC. In addition, we identify a number of genes with differential expression that maybe useful in the study, diagnosis and treatment of HCC. We demonstrate that AAV vector dose, enhancer-promoter selection, and the timing of gene delivery are the defining factors in AAV-mediated insertional mutagenesis. Our results help explain the AAV-mediated genotoxicity previously observed and have important implications for the design of both safer AAV vectors and gene therapy studies. To investigate the possibility that insertional mutagenesis by AAV contributed to the development of HCC, we collected normal and tumor tissues from adult mouse livers that received AAV injection at a neonatal stage.