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| Targets |
Endothelin-1 (1-31) targets the endothelin receptors, primarily the ET-A receptor (ETAR) and potentially ET-B receptor (ETBR), which are G-protein-coupled receptors (GPCRs). It acts as a potent vasoconstrictor and induces proliferation of human mesangial cells through ET-A receptor-mediated ERK activation. It also mediates chymase-dependent alternative processing pathway of endothelin activation independent of the classical endothelin-converting enzyme (ECE) pathway.
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| ln Vitro |
Endothelin-1(1-31)(Human)(100 pM-100 nM; 24 h) TFA stimulates the growth of human mesangial cells [2]. In humans, endothelin-1(1-31)(100 nM; 0–10 min) Human mesangial cells are activated via ERK by TFA [2].
In vitro, Endothelin-1 (1-31) at concentrations of 100 pM to 100 nM induces proliferation of human mesangial cells in a concentration-dependent manner over 24 hours, as measured by 3H-thymidine incorporation. At 100 nM, it rapidly induces ERK activation within 5-10 minutes, with peak activation (2.45-fold) at 10 minutes in human mesangial cells. This peptide also induces contraction of isolated blood vessels and stimulates growth of various cell types. It is a potent vasoconstrictor and hypertension-inducing agent derived from chymase-mediated hydrolysis of big ET-1. |
| ln Vivo |
The ET-1 (1-31) TFA (100 nM; single dose) causes the mesenteric arteries in mice to constrict. Age may cause contractions to worsen and may be mediated by ETA receptors. There are distinct distinctions between men and women in the contemporary chronic diabetes scenario [1].
In vivo, Endothelin-1 (1-31) (100 nM; single dose) induces contraction in mouse mesenteric arteries, an effect mediated primarily by ET-A receptors and increased with aging. In a streptozotocin (STZ)-induced diabetic mouse model, the contractile response shows sex-specific differences (greater in females than males) and may be altered in chronic diabetic conditions. The compound is a potent vasoconstrictor and blood pressure-elevating agent, making it useful for studying vascular dysfunction in aging, diabetes, and hypertension. |
| Enzyme Assay |
For radioligand binding assays, express recombinant ET-A receptors in CHO or HEK293 cells. Prepare cell membrane homogenates (20-50 ug protein per well). Incubate with 10-50 pM 125I-labeled ET-1 (or ET-1 (1-31)) and varying concentrations of unlabeled Endothelin-1 (1-31) TFA (1 pM-10 uM) in binding buffer (50 mM Tris-HCl, 0.1% BSA, 10 mM MgCl2, pH 7.4) for 2 hours at 25degC. Terminate by vacuum filtration through GF/C glass fiber filters presoaked in 0.3% polyethyleneimine. Wash filters 3 times with cold buffer. Count retained radioactivity in a gamma counter. Perform competition binding analysis using nonlinear regression. For non-radioactive assays, use fluorescence polarization or surface plasmon resonance.
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| Cell Assay |
Cell Proliferation Assay[2]
Cell Types: Human mesangial cells Tested Concentrations: 100 pM-100 nM Incubation Duration: 24 h Experimental Results: Caused an increase in [3H]-thymidine incorporation into the cells in a concentration-dependent manner. Western Blot Analysis[2] Cell Types: Human mesangial cells Tested Concentrations: 100 nM Incubation Duration: 0, 5, 10, 15 and 30 min Experimental Results: ERK activities rapidly increased 2.45-fold at 5 min and peaked at 10 min. The activities of both ERKs rapidly declined, returning to the baseline control value 30 min after stimulation. Culture primary human mesangial cells or cell lines (e.g., human renal mesangial cells) in RPMI-1640 or DMEM with 10-20% FBS, insulin-transferrin-selenium, and 1% penicillin/streptomycin. For proliferation assays (3H-thymidine incorporation), seed cells in 24-well plates (2 × 10^4 cells/well), serum-starve for 24-48 hours, then treat with Endothelin-1 (1-31) TFA (100 pM-100 nM) for 24 hours. Pulse with 1 uCi/mL 3H-thymidine for the final 4-6 hours. Harvest cells onto glass fiber filters and measure incorporated radioactivity by liquid scintillation counting. For ERK activation studies, treat cells with 100 nM peptide for 0, 5, 10, 15, 30 minutes. Lyse cells with RIPA buffer, separate by SDS-PAGE, transfer to PVDF membrane, and blot with phospho-ERK1/2 (Thr202/Tyr204) and total ERK1/2 antibodies. For calcium mobilization, load cells with Fura-2 AM, add peptide (1-100 nM), and measure calcium flux by fluorescence. For contractility assays, use isolated vascular smooth muscle cells and measure cell shortening or force generation. |
| Animal Protocol |
Animal/Disease Models: ICR mice, Streptozocin (HY-13753)-induced diabetic model[1]
Doses: 100 nM Route of Administration: In the organ bath, single dose Experimental Results: In the 1-week control (but not diabetic) group, induced contraction and the contractile response was Dramatically greater in female mice than in male mice, and there was no significant difference in either male or female mice between the age-matched controls and the diabetic mice. In the 8-weeks group, the contraction was or tended to be increased compared with the corresponding 1-week group in all mice. Although in male mice this contraction was not different between control and diabetic groups, it was Dramatically greater in diabetic female mice than in the control female mice and in female diabetic mice than in male diabetic mice. The contraction was inhibited by ETA receptor inhibitor. For isolated vessel contraction studies, use male ICR mice (6-20 weeks old) or STZ-induced diabetic mice. Sacrifice animals, dissect mesenteric arteries (second/third-order branches), carefully remove connective tissue, and cut into 2-3 mm rings. Mount rings in organ bath chambers containing Krebs-Henseleit solution (118 mM NaCl, 4.7 mM KCl, 2.5 mM CaCl2, 1.2 mM MgSO4, 1.2 mM KH2PO4, 25 mM NaHCO3, 11 mM glucose, pH 7.4) bubbled with 95% O2/5% CO2 at 37degC. Equilibrate for 60 minutes under 1 g resting tension. Pre-contract with 60 mM KCl to assess viability. Add Endothelin-1 (1-31) TFA (100 nM, single dose) cumulatively or as a single concentration. Record isometric tension. To assess receptor mediation, pre-incubate with ET-A antagonist BQ-123 (1 uM, 30 minutes) or ET-B antagonist BQ-788 (1 uM) before adding peptide. For in vivo blood pressure studies (anesthetized rats), administer Endothelin-1 (1-31) via intravenous injection at 0.1-10 nmol/kg and monitor mean arterial pressure (MAP). For aging studies, compare contractile responses in young (6 weeks) vs. aged mice (64-100 weeks). For diabetic studies, use STZ-induced diabetic mice (1-week and 8-week post-diabetes induction) and compare responses to age-matched controls. |
| ADME/Pharmacokinetics |
Endothelin-1 (1-31) is a 31-amino acid peptide (MW approximately 3742 for TFA salt). As a peptide fragment, it is subject to rapid enzymatic degradation in plasma (by neutral endopeptidases, ECE, and other proteases), with a half-life expected to be in the range of minutes in circulation. The compound is not orally bioavailable. For in vitro and ex vivo studies, reconstitute in sterile water or PBS with 0.1% BSA to prevent adsorption to plastic surfaces. Storage: lyophilized powder at -80degC (2 years) or -20degC (1 year); in solvent at -80degC (6 months) or -20degC (1 month) in sealed containers, protected from moisture and light, under nitrogen.
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| Toxicity/Toxicokinetics |
The toxicity profile of Endothelin-1 (1-31) is consistent with the pharmacological activity of the endothelin system. Exogenous administration of vasoactive doses (nmol/kg range) can cause acute hypertension, vasoconstriction, ischemia, and organ hypoperfusion. At higher doses, ET-1 (1-31) may induce renal vasoconstriction, reduced glomerular filtration rate, myocardial ischemia, and pulmonary edema. The compound is not for human use and is classified as a research chemical. Standard laboratory precautions (gloves, lab coat, safety glasses) should be used. Avoid inhalation, ingestion, and skin contact. Material safety data sheets indicate potential irritation.
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| References | |
| Additional Infomation |
Endothelin-1 (1-31) Human TFA is a research peptide used to study the chymase-dependent alternative processing pathway of endothelin activation, independent of the classic ECE pathway. The peptide sequence is Cys-Ser-Cys-Ser-Ser-Leu-Met-Asp-Lys-Glu-Cys-Val-Tyr-Phe-Cys-His-Leu-Asp-Ile-Ile-Trp-Val-Asn-Thr-Pro-Glu-His-Val-Val-Pro-Tyr with disulfide bridges Cys1-Cys15 and Cys3-Cys11 (cyclic structure). The TFA salt is the trifluoroacetate form. The compound is also available as the acetate salt. This peptide is exclusively for research use in understanding endothelin biology, vascular dysfunction, hypertension, diabetic vascular complications, and renal fibrosis. It is not approved for diagnostic or therapeutic use in humans.
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| Molecular Formula |
C164H237F3N38O49S5
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| Molecular Weight |
3742.18
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| Related CAS # |
Endothelin-1 (1-31) (Human);133972-52-8;Endothelin-1 (1-31) (Human) acetate
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| Appearance |
Solid powder
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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 (e.g. under nitrogen), avoid exposure to moisture and light. |
| 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) |
May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples
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| Solubility (In Vivo) |
Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.
Injection Formulations
Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)(e.g. IP/IV/IM/SC) *Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution. Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline) Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil) Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals). View More
Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)] Oral Formulations
Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium) Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals). View More
Oral Formulation 3: Dissolved in PEG400 (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 0.2672 mL | 1.3361 mL | 2.6722 mL | |
| 5 mM | 0.0534 mL | 0.2672 mL | 0.5344 mL | |
| 10 mM | 0.0267 mL | 0.1336 mL | 0.2672 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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