Transcriptional and phenotypic comparisons of Ppara knockout and siRNA knockdown mice

Souza, Angus T. De; Dai, Xudong; Spencer, Andrew; Reppen, Tom W.; Menzie, A.M.; Roesch, Paula L.; He, Yudong; Caguyong, M.; Bloomer, Sherri; Herweijer, Hans; Wolff, Jon A.; Hagstrom, James E.; Lewis, David L.; Linsley, Peter S.; Ulrich, Roger G.

doi:10.1093/nar/gkl609

Cited by 103 publications

(74 citation statements)

References 28 publications

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“…These data suggest that Ppara siRNA effectively downregulates transcription of the targeted gene, Ppara, and its known downstream genes in vivo. The genomewide expression profiles in livers from the Ppara siRNAtreated mice also revealed an extensive off-target effect when compared with the profiles in livers from the Ppara knockout mice, as reported separately [26]. A large number of genes that were downregulated in the Ppara siRNA-treated liver profiles either were not downregulated or were upregulated in the Ppara knockout liver profiles, and vice versa.…”

Section: Author Summarysupporting

confidence: 52%

PPARα siRNA-Treated Expression Profiles Uncover the Causal Sufficiency Network for Compound-Induced Liver Hypertrophy

Dai¹,

Souza²,

Dai³

et al. 2005

PLoS Comput Biol

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Uncovering pathways underlying drug-induced toxicity is a fundamental objective in the field of toxicogenomics. Developing mechanism-based toxicity biomarkers requires the identification of such novel pathways and the order of their sufficiency in causing a phenotypic response. Genome-wide RNA interference (RNAi) phenotypic screening has emerged as an effective tool in unveiling the genes essential for specific cellular functions and biological activities. However, eliciting the relative contribution of and sufficiency relationships among the genes identified remains challenging. In the rodent, the most widely used animal model in preclinical studies, it is unrealistic to exhaustively examine all potential interactions by RNAi screening. Application of existing computational approaches to infer regulatory networks with biological outcomes in the rodent is limited by the requirements for a large number of targeted permutations. Therefore, we developed a two-step relay method that requires only one targeted perturbation for genome-wide de novo pathway discovery. Using expression profiles in response to small interfering RNAs (siRNAs) against the gene for peroxisome proliferator-activated receptor a (Ppara), our method unveiled the potential causal sufficiency order network for liver hypertrophy in the rodent. The validity of the inferred 16 causal transcripts or 15 known genes for PPARa-induced liver hypertrophy is supported by their ability to predict non-PPARa-induced liver hypertrophy with 84% sensitivity and 76% specificity. Simulation shows that the probability of achieving such predictive accuracy without the inferred causal relationship is exceedingly small (p , 0.005). Five of the most sufficient causal genes have been previously disrupted in mouse models; the resulting phenotypic changes in the liver support the inferred causal roles in liver hypertrophy. Our results demonstrate the feasibility of defining pathways mediating drug-induced toxicity from siRNA-treated expression profiles. When combined with phenotypic evaluation, our approach should help to unleash the full potential of siRNAs in systematically unveiling the molecular mechanism of biological events.

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Section: Author Summarysupporting

confidence: 52%

PPARα siRNA-Treated Expression Profiles Uncover the Causal Sufficiency Network for Compound-Induced Liver Hypertrophy

Dai¹,

Souza²,

Dai³

et al. 2005

PLoS Comput Biol

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“…Genetic ablation via homologous recombination through the germline, leading to complete loss of function from the outset in development, may be partially compensated for by functional orthologues, whereas RNAi-mediated efficient knockdown, occurring rapidly and at a defined developmental stage, may not be permissive for compensation. 23 In a recent study, T␤4 was described as dispensable for murine cardiac development and function after both global and cardiac-specific knockout. 24 This contrasts with our findings describing partially penetrant vascular phenotypes after complete loss of T␤4; however, we acknowledge that over successive generations the incidence of embryonic lethality decreased from 40% at the outset of the study (60% survival) to a current level of 20% loss in utero (80% survival) due to presumptive modifier effects.…”

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confidence: 99%

Essential Role for Thymosin β4 in Regulating Vascular Smooth Muscle Cell Development and Vessel Wall Stability

Rossdeutsch

Smart

Dubé

et al. 2012

Circulation Research

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Rationale: Compromised development of blood vessel walls leads to vascular instability that may predispose to aneurysm with risk of rupture and lethal hemorrhage. There is currently a lack of insight into developmental insults that may define the molecular and cellular characteristics of initiating and perpetrating factors in adult aneurismal disease.Objective: To investigate a role for the actin-binding protein thymosin ␤4 (T␤4), previously shown to be proangiogenic, in mural cell development and vascular wall stability. Methods and Results:Phenotypic analyses of both global and endothelial-specific loss-of-function T␤4 mouse models revealed a proportion of T␤4-null embryos with vascular hemorrhage coincident with a reduction in smooth muscle cell coverage of their developing vessels. Mechanistic studies revealed that extracellular T␤4 can stimulate differentiation of mesodermal progenitor cells to a mature mural cell phenotype through activation of the transforming growth factor-beta (TGF␤) pathway and that reduced TGF␤ signaling correlates with the severity of hemorrhagic phenotype in T␤4-null vasculature.Conclusions: T␤4 is a novel endothelial secreted trophic factor that functions synergistically with TGF␤ to regulate mural cell development and vascular wall stability. These findings have important implications for understanding congenital anomalies that may be causative for adult-onset vascular instability. (Circ Res. 2012; 111:e89-e102.) Key Words: thymosin Ⅲ vasculature Ⅲ mural cell Ⅲ aorta Ⅲ mouse Ⅲ mouse mutants Ⅲ smooth muscle differentiation Ⅲ vascular biology Ⅲ vascular smooth muscle T he development of a functional vasculature is an essential process during embryogenesis, perturbations in which result in fetal lethality or vascular disease after birth. The formation of systemic blood vessels occurs in a stereotypical fashion -endothelial tubes form through a number of mechanisms (angiogenesis, vasculogenesis, or intussusception). 1 Endothelial cells then recruit mural cells comprising the subsets of vascular smooth muscle cells (VSMCs) and pericytes to the external wall of the vessel. [2][3][4] These mural cells are required to provide structural support for the blood vessel and probably play a role in maintaining endothelial health and integrity. The establishment of a vessel wall is accomplished either through the differentiation of de novo mural cells from precursor populations or recruitment from a proliferating pool of mature cells. The former is thought to occur chiefly through the actions of endothelial secreted transforming growth factor-beta (TGF␤) and the latter through paracrine platelet-derived growth factor-B (PDGF-B). 4 Typically, in the embryo, mural cells originate from the in situ differentiation of mesodermal tissues, which surround endothelial tubes. 3,5 The exception to this is in the central nervous system, where blood vessels recruit to their outer layer via the migration of neurectodermal-derived mature mural cells, as typified by the development of the postnatal retinal vasc...

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“…HTV delivery of siRNA to knockdown peroxisome proliferator-activated receptor alpha (Ppara) resulted in a transcript profile and metabolic phenotype that is comparable to those of Ppara (À/À) knockout mice. 68 Ppara knockdown animals developed hypoglycemia and hypertriglyceridemia, phenotypes also observed in Ppara (À/À) mice. In contrast to Ppara (À/À) mice, fasting was not required to uncover these phenotypes.…”

Section: Study Of the Immune Response By Htv Deliverymentioning

confidence: 70%

Gene therapy progress and prospects: Hydrodynamic gene delivery

2006

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Over the last few years, hydrodynamic tail vein delivery has established itself as a simple, yet very effective method for gene transfer into small rodents. Hydrodynamic delivery of plasmid DNA expression vectors or small interfering RNA allows for a broad range of in vivo experiments, including the testing of regulatory elements, antibody generation, evaluation of gene therapy approaches, basic biology and disease model creation (non-heritable transgenics). The recent development of the hydrodynamic limb vein procedure provides a safe nucleic acid delivery technique with equally high efficiency in small and large research animals and, importantly, the prospects for clinical translation. Gene Therapy (2007) Hydrodynamic tail vein (HTV) delivery is highly efficient at transferring nucleic acids to the liver of mice and rats. This simple non-viral gene transfer procedure entails the rapid delivery of naked plasmid DNA (pDNA) in a relatively large volume of physiological solution. In a typical mouse, weighing 20 g, the pDNA is delivered in a total volume of 2.0 ml over a period of 5-7 s. These seemingly harsh conditions actually cause very little harm and are well tolerated. After the first description of this technique in 1999, there are now hundreds of published studies applying HTV. Although most studies have focused on pDNA delivery, it is clear that any naked polynucleotide can be delivered effectively, including small interfering RNA (siRNA). Here, we will highlight important recent research findings related to hydrodynamic delivery, including mechanistic studies, applications and the development of the clinically relevant hydrodynamic limb vein (HLV) delivery procedure.

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Transcriptional and phenotypic comparisons of Ppara knockout and siRNA knockdown mice

Cited by 103 publications

References 28 publications

PPARα siRNA-Treated Expression Profiles Uncover the Causal Sufficiency Network for Compound-Induced Liver Hypertrophy

PPARα siRNA-Treated Expression Profiles Uncover the Causal Sufficiency Network for Compound-Induced Liver Hypertrophy

Essential Role for Thymosin β4 in Regulating Vascular Smooth Muscle Cell Development and Vessel Wall Stability

Gene therapy progress and prospects: Hydrodynamic gene delivery

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