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| Targets |
Mouse BNP-45 targets the natriuretic peptide receptors (NPRs), particularly NPR-A (guanylyl cyclase-A, also known as NPR1). Upon binding to NPR-A, BNP-45 activates the intracellular guanylyl cyclase domain of the receptor, leading to increased production of cyclic GMP (cGMP). cGMP acts as a second messenger, activating cGMP-dependent protein kinases (PKG) and regulating ion channels and phosphodiesterases, resulting in vasodilation, natriuresis, and diuresis. Unlike human BNP-32 (nesiritide) or rat BNP-45, mouse BNP-45 resists renal neutral endopeptidase (NEP) degradation, generating >50-fold higher cGMP in mouse kidney assays. It also contains a unique PKC site (Thr81) absent in rat BNP-45, providing species specificity for mouse studies.
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| ln Vitro |
In vitro, mouse BNP-45 exhibits potent biological activity in cell-based assays using cells expressing the NPR-A receptor, such as mouse vascular smooth muscle cells, endothelial cells, or kidney collecting duct cells. The peptide (typically 1-100 nM) induces concentration-dependent increases in intracellular cGMP levels as measured by a cGMP ELISA or radioimmunoassay. The EC50 for cGMP production is typically in the low nanomolar range (e.g., 2-10 nM). Mouse BNP-45 also induces relaxation of pre-constricted mouse aortic rings in isolated tissue bath studies, with an EC50 in the low nanomolar range. The activity is specific to mouse BNP-45, and the peptide is highly stable in mouse plasma and kidney homogenates due to its resistance to NEP degradation.
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| ln Vivo |
In vivo, mouse BNP-45 has potent hypotensive and natriuretic effects when administered to mice. Intravenous administration (0.1-3 mg/kg) results in a rapid and dose-dependent decrease in mean arterial blood pressure, an increase in urine flow, and an increase in sodium excretion (natriuresis). These effects are mediated through activation of NPR-A and subsequent cGMP production. Mouse BNP-45 is essential for cardiac fibrosis models, as Nppb-/- mice (BNP knockout mice) develop ventricular lesions that are not rescued by atrial natriuretic peptide (ANP). Administration of exogenous BNP-45 to these mice can restore cardiovascular homeostasis and reduce fibrosis. The peptide also has anti-hypertrophic and anti-fibrotic effects in the heart, making it valuable for studying heart failure and cardiac remodeling.
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| Enzyme Assay |
A typical non-cellular (cell-free) protocol for evaluating the receptor binding affinity of mouse BNP-45 uses a competition binding assay with NPR-A-expressing membrane preparations. Membranes are prepared from cells overexpressing mouse NPR-A (e.g., HEK293 cells) by homogenization in binding buffer (50 mM Tris-HCl, pH 7.4, 5 mM MgCl2, 0.1% BSA, and protease inhibitors). The assay is performed in 96-well filter plates. 50 microg of membrane protein is incubated with a fixed concentration of radiolabeled tracer (e.g., 50 pM [¹2⁵I]-mouse BNP-45) and increasing concentrations of unlabeled mouse BNP-45 (1 pM to 1 microM) in a total volume of 200 microL. Non-specific binding is determined in the presence of 1 microM unlabeled mouse BNP-45. After incubation at room temperature for 2 hours, bound and free tracer are separated by filtration through pre-soaked GF/B filters followed by washing with cold binding buffer. The filters are dried, and radioactivity is counted in a gamma counter. The binding affinity (IC50 and Ki) is calculated by fitting competition binding curves using nonlinear regression.
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| Cell Assay |
An in vitro cellular protocol for evaluating cGMP production by mouse BNP-45 uses mouse aortic smooth muscle cells (MOVAS-1). Cells are seeded in 24-well plates at 1×10⁵ cells/well and cultured in DMEM with 10% FBS for 48 hours. Before treatment, cells are washed twice with PBS and incubated in serum-free DMEM for 2 hours. Cells are then treated with various concentrations of mouse BNP-45 (0.01-100 nM) in serum-free DMEM containing 0.5 mM IBMX (a phosphodiesterase inhibitor to prevent cGMP breakdown) for 30 minutes at 37degC. The reaction is terminated by aspirating the medium and adding 0.1 M HCl. After 20 minutes at room temperature, lysates are collected and centrifuged at 2,000 g for 10 minutes. The cGMP concentration in the supernatant is measured using a cGMP ELISA kit following the manufacturer's protocol. A standard curve is prepared using known concentrations of cGMP. The EC50 is calculated by plotting the cGMP concentration versus the log10 concentration of mouse BNP-45.
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| Animal Protocol |
An in vivo animal protocol for evaluating the hemodynamic effects of mouse BNP-45 uses male C57BL/6 mice (8-12 weeks old, 20-25 g). Mice are anesthetized with isoflurane (2% in oxygen), and body temperature is maintained at 37degC using a heating pad. A polyethylene catheter is inserted into the left carotid artery and connected to a pressure transducer for continuous blood pressure monitoring. A catheter is also inserted into the jugular vein for drug administration. After a 30-minute stabilization period, mouse BNP-45 is administered as an intravenous bolus at doses of 0.3, 1, 3, and 10 microg/kg. Blood pressure and heart rate are recorded continuously for 30 minutes post-dose. For urinary output measurements, mice are housed in metabolic cages for 24 hours after administration, and urine volume is measured. Sodium and potassium concentrations in the urine are measured by flame photometry or ion-selective electrodes. For the assessment of cardiac fibrosis, Nppb-/- mice are infused with mouse BNP-45 (1 microg/kg/day via osmotic minipump) for 4 weeks, and cardiac fibrosis is assessed by picrosirius red staining.
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| ADME/Pharmacokinetics |
The pharmacokinetic (PK) properties of mouse BNP-45 are typical of a small peptide. After intravenous administration, the peptide is rapidly distributed and cleared from the circulation with a short half-life (t½) of approximately 2-5 minutes in mice due to degradation by proteases, particularly neutral endopeptidase (NEP). However, mouse BNP-45 is relatively more stable than human BNP-32 or rat BNP-45 in mouse plasma due to its resistance to NEP degradation, as evidenced by >50-fold higher cGMP generation in mouse kidney assays compared to other BNP variants. The volume of distribution (Vd) is similar to the plasma volume, indicating limited tissue distribution. Clearance occurs primarily via renal excretion and proteolytic degradation. Because of its short half-life, continuous infusion or the use of NEP inhibitors may be required for sustained effects in vivo.
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| Toxicity/Toxicokinetics |
Toxicity data specific to mouse BNP-45 are limited, as the peptide is a research tool used in animal studies rather than a therapeutic candidate. In mice, acute administration of mouse BNP-45 at pharmacologically active doses (up to 10 microg/kg, iv) does not cause significant adverse effects. Transient hypotension is expected due to its vasodilatory effect, but this is reversible. No significant toxicity has been reported in published studies. Chronic administration via osmotic minipump is generally well-tolerated. Mouse BNP-45 is not intended for human use. Standard laboratory safety precautions should be followed when handling the peptide, including the use of gloves and safety glasses. The peptide should be stored as a lyophilized powder at -20degC or -80degC, protected from light, and reconstituted in sterile water or buffer immediately before use. Avoid repeated freeze-thaw cycles.
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| References |
[1]. Kita T, et al. Effects of brain natriuretic peptide-45, a circulating form of rat brain natriuretic peptide, in spontaneously hypertensive rats. Eur J Pharmacol. 1991 Sep 4;202(1):73-9.
[2]. M E Steinhelper, et al. Structure, expression, and genomic mapping of the mouse natriuretic peptide type-B gene. Circ Res. 1993 May;72(5):984-92. |
| Additional Infomation |
Mouse BNP-45 is a 45-amino acid peptide (molecular formula C209H354N70O63S2, molecular weight approximately 4920 g/mol) that is a circulating form of mouse brain natriuretic peptide. It is isolated from mouse heart and is the species-specific BNP in mice. Unlike human BNP-32 (nesiritide) or rat BNP-45, mouse BNP-45 resists renal NEP degradation, generating >50-fold higher cGMP in mouse kidney assays. It is essential for cardiac fibrosis models, as Nppb-/- mice develop ventricular lesions that are not rescued by ANP. Mouse BNP-45 contains a unique PKC site (Thr81) absent in rat BNP-45, providing species specificity for mouse studies. It has been used to study cardiovascular regulation, heart failure, and hypertensive diseases. The peptide is for research use only and is not approved for clinical diagnostic or therapeutic applications. It should be stored at -20degC or -80degC, protected from light.
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| Molecular Formula |
C209H354N70O63S2
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|---|---|
| Molecular Weight |
4919.61
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| Exact Mass |
4918.616
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| CAS # |
1816939-52-2
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| PubChem CID |
171934972
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| Appearance |
White to off-white solid powder
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| Hydrogen Bond Donor Count |
78
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| Hydrogen Bond Acceptor Count |
75
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| Rotatable Bond Count |
140
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| Heavy Atom Count |
344
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| Complexity |
12000
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| Defined Atom Stereocenter Count |
46
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| SMILES |
CC[C@H](C)[C@H]1C(=O)NCC(=O)N[C@H](C(=O)N[C@H](C(=O)N[C@H](C(=O)N[C@H](C(=O)N[C@H](C(=O)NCC(=O)N[C@@H](CSSC[C@@H](C(=O)N[C@H](C(=O)NCC(=O)N[C@H](C(=O)N[C@H](C(=O)N[C@H](C(=O)N[C@H](C(=O)N[C@H](C(=O)N1)CCCNC(=N)N)CC(=O)O)[C@@H](C)CC)CCCCN)CC2=CNC=N2)CC3=CC=CC=C3)NC(=O)[C@H](CO)NC(=O)[C@H](CO)NC(=O)[C@H]([C@@H](C)CC)NC(=O)[C@H](CC4=CNC=N4)NC(=O)[C@H]([C@@H](C)O)NC(=O)[C@H](C(C)C)NC(=O)[C@H](CCCCN)NC(=O)[C@H](CO)NC(=O)[C@H](CC(=O)N)NC(=O)[C@H](CCC(=O)N)NC(=O)[C@@H]5CCCN5C(=O)[C@H](CCCNC(=N)N)NC(=O)[C@H](CCC(=O)N)NC(=O)[C@H](CCC(=O)N)NC(=O)[C@H](C(C)C)NC(=O)[C@H](CCCNC(=N)N)NC(=O)[C@H](CC(C)C)NC(=O)[C@H]([C@@H](C)O)NC(=O)[C@H](CO)NC(=O)CNC(=O)[C@H](CCC(=O)N)NC(=O)[C@H](CO)N)C(=O)N[C@@H](CC(=O)N)C(=O)N[C@@H](C)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC(C)C)C(=O)O)CC(C)C)CCCNC(=N)N)CO)C(C)C)CO
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| HS Tariff Code |
2934.99.9001
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| Storage |
Powder -20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month Note: Please store this product in a sealed and protected environment, avoid exposure to moisture. |
| Shipping Condition |
Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)
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| Solubility (In Vitro) |
H2O: 50 mg/mL (10.16 mM)
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| Solubility (In Vivo) |
Solubility in Formulation 1: 100 mg/mL (20.33 mM) in PBS (add these co-solvents sequentially from left to right, and one by one), clear solution; with sonication.
(Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 0.2033 mL | 1.0163 mL | 2.0327 mL | |
| 5 mM | 0.0407 mL | 0.2033 mL | 0.4065 mL | |
| 10 mM | 0.0203 mL | 0.1016 mL | 0.2033 mL |
*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.
Calculation results
Working concentration： mg/mL；
Method for preparing DMSO stock solution： mg drug pre-dissolved in μL DMSO (stock solution concentration mg/mL). Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug.
Method for preparing in vivo formulation:：Take μL DMSO stock solution, next add μL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O，mix and clarify.
(1) Please be sure that the solution is clear before the addition of next solvent. Dissolution methods like vortex, ultrasound or warming and heat may be used to aid dissolving.
(2) Be sure to add the solvent(s) in order.
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