Visualization of the Dynamics of Gene Expression in the Living Mouse

Ryan, Amy; Scrable, Heidi

doi:10.1162/153535004773861705

Cited by 13 publications

(15 citation statements)

References 16 publications

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“…In M mice, MGF was expressed using the HD locus promoter which was modified to include lac operators and in R mice, the human β-actin promoter was used to drive the Lac-I gene expression. Both transgene constructions have been previously utilized in our studies [17, 18] when overexpressed MGF was identified in spinal cord and hippocampal neurons (Fig. 1b and c).…”

Section: Methodsmentioning

confidence: 99%

Mechano growth factor, a splice variant of IGF-1, promotes neurogenesis in the aging mouse brain

et al. 2017

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Mechano growth factor (MGF) is a splice variant of IGF-1 first described in skeletal muscle. MGF induces muscle cell proliferation in response to muscle stress and injury. In control mice we found endogenous expression of MGF in neurogenic areas of the brain and these levels declined with age. To better understand the role of MGF in the brain, we used transgenic mice that constitutively overexpressed MGF from birth. MGF overexpression significantly increased the number of BrdU+ proliferative cells in the dentate gyrus (DG) of the hippocampus and subventricular zone (SVG). Although MGF overexpression increased the overall rate of adult hippocampal neurogenesis at the proliferation stage it did not alter the distribution of neurons at post-mitotic maturation stages. We then used the lac-operon system to conditionally overexpress MGF in the mouse brain beginning at 1, 3 and 12 months with histological and behavioral observation at 24 months of age. With conditional overexpression there was an increase of BrdU+ proliferating cells and BrdU+ differentiated mature neurons in the olfactory bulbs at 24 months when overexpression was induced from 1 and 3 months of age but not when started at 12 months. This was associated with preserved olfactory function. In vitro, MGF increased the size and number of neurospheres harvested from SVZ-derived neural stem cells (NSCs). These findings indicate that MGF overexpression increases the number of neural progenitor cells and promotes neurogenesis but does not alter the distribution of adult newborn neurons at post-mitotic stages. Maintaining youthful levels of MGF may be important in reversing age-related neuronal loss and brain dysfunction.Electronic supplementary materialThe online version of this article (doi:10.1186/s13041-017-0304-0) contains supplementary material, which is available to authorized users.

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Section: Methodsmentioning

confidence: 99%

Mechano growth factor, a splice variant of IGF-1, promotes neurogenesis in the aging mouse brain

et al. 2017

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“…The present study has tested a modified bacterial lac operon system, developed originally by Scrable and colleagues [6]–[8], to reversibly control gene expression in the vasculature in vivo. We show that this system can be used to reliably repress gene expression selectively in the vascular endothelium, through the use of the modified endothelial specific promoter TIE2 [11].…”

Section: Discussionmentioning

confidence: 99%

“…Mice from this strain were developed to be homozygous for a transgene that expresses the LacI R inhibitor protein under control of the ubiquitous β-actin promoter [8]. The heterozygous LacI R strain was a generous gift from Professor Heidi Scrable, University of Virginia, Charlottesville, Virginia, USA.…”

Section: Methodsmentioning

confidence: 99%

“…TIE2LacO-Cx40Tg-EGFP/LacI R mice were generated by crossing the TIE2Lac O-Cx40Tg-EGFP mice with homozygous LacI R mice (Figure 1B) [8].…”

Section: Methodsmentioning

confidence: 99%

“…With the intention of creating an alternative system which utilises an innocuous inducer, Scrable and colleagues have modified the bacterial lac operator system for use in mammalian systems and reported reversible control of the tyrosinase gene and eye pigmentation, as well as the Huntington promoter driving a luciferase reporter, using the lactose analogue, Isopropyl β-D-1-thiogalactopyranoside (IPTG) [6]–[8]. Their system relied on the introduction into the tyrosinase and Huntington promoters of Lac operon ( Lac O) sequences at specific distances spanning the transcription start site (see also [3]); a procedure which did not interfere with promoter function and specificity.…”

Section: Introductionmentioning

confidence: 99%

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Regulation of Endothelial-Specific Transgene Expression by the LacI Repressor Protein In Vivo

et al. 2014

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Genetically modified mice have played an important part in elucidating gene function in vivo. However, conclusions from transgenic studies may be compromised by complications arising from the site of transgene integration into the genome and, in inducible systems, the non-innocuous nature of inducer molecules. The aim of the present study was to use the vascular system to validate a technique based on the bacterial lac operon system, in which transgene expression can be repressed and de-repressed by an innocuous lactose analogue, IPTG. We have modified an endothelium specific promoter (TIE2) with synthetic LacO sequences and made transgenic mouse lines with this modified promoter driving expression of mutant forms of connexin40 and an independently translated reporter, EGFP. We show that tissue specificity of this modified promoter is retained in the vasculature of transgenic mice in spite of the presence of LacO sequences, and that transgene expression is uniform throughout the endothelium of a range of adult systemic and cerebral arteries and arterioles. Moreover, transgene expression can be consistently down-regulated by crossing the transgenic mice with mice expressing an inhibitor protein LacIR, and in one transgenic line, transgene expression could be de-repressed rapidly by the innocuous inducer, IPTG. We conclude that the modified bacterial lac operon system can be used successfully to validate transgenic phenotypes through a simple breeding schedule with mice homozygous for the LacIR protein.

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Perturbation of chemical coupling by an endothelial Cx40 mutant attenuates endothelium-dependent vasodilation by KCa channels and elevates blood pressure in mice

Chaston

Haddock

Howitt

et al. 2014

Pflugers Arch - Eur J Physiol

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Mutant forms of connexin40 (Cx40) exist in the human population and predispose carriers to atrial fibrillation. Since endothelial expression of Cx40 is important for electrical and chemical communication within the arterial wall, carriers of mutant Cx40 proteins may be predisposed to peripheral arterial dysfunction and dysregulation of blood pressure. We have therefore studied mice expressing either a chemically dysfunctional mutant, Cx40T202S, or wild-type Cx40, with native Cx40, specifically in the endothelium. Blood pressure was measured by telemetry under normal conditions and during cardiovascular stress induced by locomotor activity, phenylephrine or nitric oxide blockade (N(ɷ)-nitro-L-arginine methyl ester hydroxide, L-NAME). Blood pressure of Cx40T202STg mice was significantly elevated at night when compared with wild-type or Cx40Tg mice, without change in mean heart rate, pulse pressure or locomotor activity. Analysis over 24 h showed that blood pressure of Cx40T202STg mice was significantly elevated at rest and additionally during locomotor activity. In contrast, neither plasma renin concentration nor pressor responses to phenylephrine or L-NAME were altered, the latter indicating that nitric oxide bioavailability was normal. In isolated, pressurised mesenteric arteries, hyperpolarisation and vasodilation evoked by SKA-31, the selective modulator of SKCa and IKCa channels, was significantly reduced in Cx40T202STg mice, due to attenuation of the SKCa component. Acetylcholine-induced ascending vasodilation in vivo was also significantly attenuated in cremaster muscle arterioles of Cx40T202STg mice, compared to wild-type and Cx40Tg mice. We conclude that endothelial expression of the chemically dysfunctional Cx40T202S reduces peripheral vasodilator capacity mediated by SKCa-dependent hyperpolarisation and also increases blood pressure.

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Visualization of the Dynamics of Gene Expression in the Living Mouse

Cited by 13 publications

References 16 publications

Mechano growth factor, a splice variant of IGF-1, promotes neurogenesis in the aging mouse brain

Mechano growth factor, a splice variant of IGF-1, promotes neurogenesis in the aging mouse brain

Regulation of Endothelial-Specific Transgene Expression by the LacI Repressor Protein In Vivo

Perturbation of chemical coupling by an endothelial Cx40 mutant attenuates endothelium-dependent vasodilation by KCa channels and elevates blood pressure in mice

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