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The use of spironolactone, the most commonly used antimineralocorticoid compound, is limited by the occurrence of sexual endocrine effects. New antagonists are therefore required which lack these unwanted effects. Three 9 alpha,11 alpha-epoxy-derivatives of known aldosterone antagonists (spironolactone, prorenone and mexrenone) have been characterised in vitro and in vivo. In each experiment spironolactone was run as a reference. The introduction of the epoxy-group only marginally affected the binding affinity of these compounds for the mineralocorticoid receptor, whereas it caused a decrease for the androgen and progesterone receptors of between 10- and 500-fold. In vivo, all three epoxy-derivatives (3 mg/kg) were potent aldosterone antagonists, 1 to 2 times the potency of spironolactone in the rat. Parallel to the decreased affinity for the androgen and progesterone receptor in vitro, there was a 3- to 10-fold decrease of the antiandrogenic and progestagenic effect compared to spironolactone in the rat and in the rabbit, respectively. Virtually no disturbance of the vaginal or ovulatory cycle was observed with either epoxymexrenone or epoxyprorenone, although epoxyspironolactone caused a 20% decrease in ovulation. It appears therefore that the 9 alpha, 11 alpha-position of the steroid structure is a site of the molecule which can be modified to improve the specificity of aldosterone-antagonists not only in vitro, but also in vivo.
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The In Vivo Therapeutics (IVT) team maintains and distributes on-site breeding colonies of specialized immunocompetent (BoyJ, C57BL/6, BoyJ/C57 F1 cross) and immunodeficient (NSG and NRG) mouse strains.
As requested and coordinated by research programs, we support the set up and validation of syngeneic mouse tumor models and development of patient-derived human xenografts.
We manage all aspects of the IVIS SpectrumCT optical imaging system to evaluate tumor response in mouse models via bioluminescence and fluorescence modalities.
The IVT team also manages the radiation facility and performs irradiation services needed for in vitro and in vivo experiments, including hematopoietic stem-cell transplantations.
For more information, contact IVT manager Tony Sinn at alsinn@iupui.edu or 317-274-8811.
The core is funded, in part, by grants and contracts of the investigators who utilize the facility. Due to the variety of in vivo models and drug regimens requested, the specific costs of a study will be calculated once the study design is finalized.
NanoLuc luciferase provides a new reporter option for advanced in vivo imaging applications. Its small size is ideal for engineering into compact genomes, and ATP-independence allows in vivo monitoring of both intracellular and extracellular events. Substrate specificity makes NanoLuc luciferase an ideal complementary reporter to firefly luciferase for an improved dual-luciferase in vivo imaging solution.
The Nano-Glo Fluorofurimazine In Vivo Substrate (FFz) is an optimized reagent designed specifically for in vivo detection of NanoLuc luciferase, NanoLuc fusion proteins or reconstituted NanoBiT luciferase. This optimized, aqueous-soluble, in vivo detection reagent provides increased substrate bioavailability for bright, stable signals and offers flexible delivery options with handling requirements compatible with in vivo workflows.
The Nano-Glo Fluorofurimazine In Vivo Substrate can be delivered via multiple routes of administration. Each method offers different signal intensity and kinetics, allowing flexibility between experimental systems.
NanoLuc signal of 4T1 primary tumors in BALB/c mice. Cells expressing NanoLuc luciferase were orthotopically implanted into the #3 mammary fat pad of female BALB/c mice, and tumors were allowed to grow for 22 days. Mice were injected with FFz via i.p. (top image; circles) or i.v. injection (bottom image; squares). Bioluminescence imaging was done at the University of Wisconsin Small Animal Imaging and Radiotherapy Facility.
In vivo whole-body imaging of mice injected with AAV9-NanoLuc-HaloTag viral particles. Mice were injected with increasing levels of AAV9-NanoLuc-HaloTag viral particles. Nano-Glo FFz (0.44mol) was injected 7- and 13-days post-transduction followed by in vivo imaging. Tissue tropism starts to appear at 7 days, and a stronger signal and more extensive tissue tropism is visible after 13 days. Bioluminescence imaging was done at the University of Wisconsin Small Animal Imaging and Radiotherapy Facility.
NanoLuc reporters can be multiplexed with firefly reporters to enable two-population bioluminescent imaging in animals. This publication used a NanoLuc reporter to track tumor size and a firefly substrate to visualize CAR-T cells in the same animal over multiple days:
BY USE OF THIS PRODUCT, RESEARCHER AGREES TO BE BOUND BY THE TERMS OF THIS LIMITED USE LABEL LICENSE. If researcher is not willing to accept the terms of this label license, and the product is unused, Promega will accept return of the unused product and provide researcher with a full refund.
For uses of Nano-Glo-branded reagents intended for energy transfer (such as bioluminescence resonance energy transfer) to acceptors other than a genetically encoded autofluorescent protein, researchers must:
(a) use NanoBRET-branded energy acceptors (e.g., BRET-optimized HaloTag ligands) for all determinations of energy transfer activity by this product; or
(b) contact Promega to obtain a license for use of the product for energy transfer assays to energy acceptors not manufactured by Promega.
With respect to any uses outside this label license, including any diagnostic, therapeutic, prophylactic or commercial uses, please contact Promega for supply and licensing information. PROMEGA MAKES NO REPRESENTATIONS OR WARRANTIES OF ANY KIND, EITHER EXPRESSED OR IMPLIED, INCLUDING FOR MERCHANTABILITY OR FITNESS FOR A PARTICULAR PURPOSE, WITH REGARD TO THE PRODUCT. The terms of this label license shall be governed under the laws of the State of Wisconsin, USA. 152ee80cbc
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