CRISPR-mediated direct mutation of cancer genes in the mouse liver

The study of cancer genes in mouse models has traditionally relied on genetically-engineered strains made via transgenesis or gene targeting in embryonic stem (ES) cells1. Here we describe a new method of cancer model generation using the CRISPR/Cas system in vivo in wild-type mice. We have used hydrodynamic injection to deliver a CRISPR plasmid DNA expressing Cas9 and single guide RNAs (sgRNAs)2–4 to the liver and directly target the tumor suppressor genes Pten5 and p536, alone and in combination. CRISPR-mediated Pten mutation led to elevated Akt phosphorylation and lipid accumulation in hepatocytes, phenocopying the effects of deletion of the gene using Cre-LoxP technology7, 8. Simultaneous targeting of Pten and p53 induced liver tumors that mimicked those caused by Cre-loxP-mediated deletion of Pten and p53. DNA sequencing of liver and tumor tissue revealed insertion or deletion (indel) mutations of the tumor suppressor genes, including bi-allelic mutations of both Pten and p53 in tumors. Furthermore, co-injection of Cas9 plasmids harboring sgRNAs targeting the β-Catenin gene (Ctnnb1) and a single-stranded DNA (ssDNA) oligonucleotide donor carrying activating point mutations led to the generation of hepatocytes with nuclear localization of β-Catenin. This study demonstrates the feasibility of direct mutation of tumor suppressor genes and oncogenes in the liver using the CRISPR/Cas system, which presents a new avenue for rapid development of liver cancer models and functional genomics

Parallel Implementation of a Compressed Image Decoder

. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . viii CHAPTER I. INTRODUCTION . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 A. Previous Work . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 B. Scope . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 II. THE STATISTICALLY LOSSLESS COMPRESSION ALGORITHM . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3 A. Vector Quantization . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3 B. Mean Residual Vector Quantization . . . . . . . . . . . . . . . . . 5 C. Lossless Encoding . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6 D. Implementation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10 E. Application Specific Alterations . . . . . . . . . . . . . . . . . . 11 III. THE DECODING PROBLEM . . . . . . . . . . . . . . . . . . . . . 12 A. Compressed Data Format . . . . . . . . . . . . . . . . . . . . . . ..

Functional beta(1)- and beta(2)-adrenoceptors in the left and right atrium of pre-hypertensive rats

There is a small increase in the functional beta(2)-adrenoceptor response on the spontaneously hypertensive rat (SHR) left atrium in the early stages of hypertension. In the present study, the functional beta(1)- and beta(2)-adrenoceptors of the left and right atrium in SHR pre-hypertension and age-matched (5-week-old) Wistar Kyoto (WKY) rats were characterized. Contractility methods with isoprenaline, T-0509 (a selective beta(1)-adrenoceptor agonist) and procaterol (a selective beta(2)-adrenoceptor agonist) were used. At 5 weeks, the SHRs were pre-hypertensive. Isoprenaline was more potent on the left atrium of 5-week-old SHRs than WKY rats. Bisoprolol, a selective beta(1)-adrenoceptor antagonist, was more potent against isoprenaline and T-0509 on the SHR than WKY rat left atrium. ICI 118,551, a selective beta(2)-adrenoceptor antagonist, was more potent against procaterol and T-0509 on the SHR than WKY rat left atrium. The results with bisoprolol and ICI 118,551 suggest that there are more functional beta(1)- and beta(2)-adrenoceptors on the left atrium of 5-week-old SHRs than WKY rats. Isoprenaline, T-0509 and procaterol were equipotent on the right atrium of 5-week-old WKY rats and SHRs. Bisoprolol was more potent against isoprenaline, T-0509 and procaterol on the SHR than WKY rat right atrium. ICI 118,551 was more potent against T-0509, but not isoprenaline and procaterol, on the SHR than WKY rat left atrium. This suggests there are more functional beta(1)-adrenoceptors, and probably more functional beta(2)-adrenoceptors, on the right atrium of 5-week-old SHRs than WKY rats. These functional differences in beta(1)-and beta(2)-adrenoceptor-mediated responses of the left and right atria of pre-hypertensive SHRs cannot be caused by hypertension, and may be associated with the onset of hypertension