Apelin, a circulating peptide hormone, is synthesized and secreted by a variety of cell types including those of the cardiovascular, endocrine, gastrointestinal and nervous systems. Apelin was recently identified as the endogenous ligand of the APJ, a formerly orphaned G-protein coupled receptor (GPCR) with a similarly broad distribution of expression[1]. ML221 is a potent apelin receptor (APJ) antagonist with IC50s of 0.70 μM and 1.75 μM in cAMP and β-arrestin assay respectively[2]. Cholangiocarcinoma (CCA) cell line HuH-28 treated with 10 μM of ML221 for 24 h showed significant decreased expression of Ki-67, as well as VEGF-A, VEGF-C, Ang-1 and Ang-2, while these were not observed in SG231 cells. CCA xenograft tumors in mice were treated with ML221 (150 μg/kg) for 3× weekly via tail vein injection for 4 weeks, and tumor volumes were significantly smaller compared to those in the untreated control mice[3].
ML221 细胞实验
Cell Line
Concentration
Treated Time
Description
References
PLC5
100 µM
3 days
Evaluate the effect of ML221 on cell proliferation
Theranostics. 2019 Jul 9;9(18):5246-5260.
HepG2
100 µM
3 days
Evaluate the effect of ML221 on cell proliferation
Theranostics. 2019 Jul 9;9(18):5246-5260.
LO2
100 µM
3 days
Evaluate the effect of ML221 on cell proliferation
Evaluate the effect of ML221 on cell proliferation
Theranostics. 2019 Jul 9;9(18):5246-5260.
HKCI-2
100 µM
3 days
Evaluate the effect of ML221 on cell proliferation
Theranostics. 2019 Jul 9;9(18):5246-5260.
Human hepatic stellate cell lines (HHSteCs)
10 µM
24 hours
To evaluate the effect of ML221 on apelin-induced hepatic stellate cell proliferation and activation, results showed that ML221 pretreatment reduced apelin-induced hepatic stellate cell proliferation and activation.
Hepatology. 2021 Jun;73(6):2411-2428.
Human primary cholangiocyte cell line (HIBEpiCs)
10 µM
24 hours
To evaluate the effect of ML221 on apelin-induced cholangiocyte proliferation, results showed that ML221 pretreatment reduced apelin-induced cholangiocyte proliferation.
Hepatology. 2021 Jun;73(6):2411-2428.
TC-1 cells
10 µM
24 hours
ML221, a functional antagonist of apelin 13, was used to explore whether MLN protected against lung injury by modulating apelin 13 expression. ML221 inhibited the MLN-induced improvement in cell viability, increased ROS production, and blocked the beneficial effects of MLN on mitochondrial ATP content, cell apoptosis, and senescence.
Exp Mol Med. 2019 Jul 4;51(7):1-12.
SG231 cells
10 µM
24 hours
ML221 treatment decreased expression of Ki-67, as well as VEGF-A, VEGF-C, Ang-1 and Ang-2
Cancer Lett. 2017 Feb 1;386:179-188.
HuH-28 cells
10 µM
24 hours
ML221 treatment significantly decreased expression of Ki-67, as well as VEGF-A, VEGF-C, Ang-1 and Ang-2
Cancer Lett. 2017 Feb 1;386:179-188.
Mz-ChA-1 cells
7.5, 10, 15 µM
24 hours
ML221 treatment significantly decreased PCNA and Ki-67 expression
Cancer Lett. 2017 Feb 1;386:179-188.
BEnd.3 cells
0-30 µM
24 hours
ML221 suppressed endothelial cell proliferation dose-dependently by blocking apelin-APJ signaling without affecting VEGF/VEGFR2 expression.
Sci Rep. 2017 Nov 8;7(1):15062.
OVCAR-5
15–50 µM
24–96 hours
ML221 efficiently suppressed increased cell proliferation in APJ-overexpressing cells.
Mol Cancer Res. 2019 Jun;17(6):1378-1390.
Buffalo granulosa cells
10 µM
48 hours
To evaluate the effect of APLN on E2 and P4 secretion, ML221 significantly reduced the secretion of E2 and P4
Front Endocrinol (Lausanne). 2022 Mar 10;13:844360.
Raji cells
5 µM
To investigate the effect of apelin/APLNR axis on B cell migration and immune molecule expression. Results showed that apelin/APLNR promoted B cell migration and inflammatory cytokine expression.
Int J Med Sci. 2025 Jan 1;22(1):197-208.
ML221 动物实验
Species
Animal Model
Administration
Dosage
Frequency
Description
References
Sprague-Dawley rats
Chronic constriction injury (CCI) model
Intrathecal injection
1, 3, 10 and 30 µg
Single injection or daily injections for 3 consecutive days
To evaluate the effect of ML221 on CCI-induced neuropathic pain. Results showed that ML221 alleviated mechanical allodynia and heat hyperalgesia in a dose-dependent manner.
Mol Med Rep. 2017 Aug;16(2):1223-1231
Balb/c nude mice
Subcutaneous xenograft model
Intraperitoneal injection
10 mg/kg
Every other day until the end of the experiment
Evaluate the effect of ML221 on HCC xenograft growth
Theranostics. 2019 Jul 9;9(18):5246-5260.
C57BL/6J mice
DOX-induced myocardial injury model
Tail vein injection
10 mg/kg
Once daily for 7 days
To investigate the protective effect of ELA-11 on DOX-induced myocardial injury, results showed that ELA-11 significantly improved cardiac function and reduced myocardial fibrosis and apoptosis.
Front Pharmacol. 2022 Sep 8;13:873614
Mice
Diabetic db/db mouse model
Intraperitoneal injection
10 mg/kg
For 7 consecutive days
To evaluate the effect of ML221 on the integrity of the blood-testis barrier and sperm quality in diabetic mice, it was found that ML221 significantly improved the integrity of the blood-testis barrier and improved sperm quality.
Nat Commun. 2022 Nov 28;13(1):7335
Mice
Bilateral renal ischemia-reperfusion injury model
Intraperitoneal injection
10 mg/kg
Administered every other day from the day of the injury until the end of the experiment
ML221 blocked the beneficial effects of ELA32 peptide on AKI but had no effect on the combination treatment of nor-NOHA and Paricalcitol
Theranostics. 2023 Jun 4;13(10):3387-3401
Mice
Oxygen-induced retinopathy (OIR) model
Intraperitoneal injection
10 mg/kg/day
Once daily for 5 days (P12 to P16)
ML221 significantly suppressed pathological retinal angiogenesis (reduced neovascular tufts) while promoting revascularization of ischemic areas (reduced avascular area), without altering VEGF/VEGFR2 expression. APJ was highly expressed in abnormal vascular endothelial cells but minimally detected in normal vessels.
Sci Rep. 2017 Nov 8;7(1):15062.
Mice
Bile duct ligation (BDL) model
Tail vein injection
150 μg/kg
3 times per week for 1 week
To evaluate the effect of ML221 on bile duct ligation-induced cholangiocyte proliferation and liver fibrosis, results showed that ML221 treatment reduced cholangiocyte proliferation and liver fibrosis.
ML221 was used to block the protective effect of MLN on lung injury by modulating apelin 13. Mice treated with ML221 showed blocked survival benefits induced by MLN and inhibited upregulation of E-cadherin expression.
Exp Mol Med. 2019 Jul 4;51(7):1-12.
Mice
Myocardial ischemia-reperfusion injury model
Langendorff perfusion
50 nM
Single dose, 20 minutes
To evaluate the protective effects of ML221 on ischemia-reperfusion hearts. Results showed that ML221 treatment partially restored left ventricular function, reduced myocardial infarct size, and synchronized the activation of the PI3K-AKT-mTOR signaling axis.
Front Pharmacol. 2023 Oct 24;14:1145413.
Sprague-Dawley rats
Chronic doxorubicin-induced cardiotoxicity model
Subcutaneous administration via osmotic pump
500 µg/kg b.w./day
Continuous for 28 days
To evaluate the effect of ML221 on doxorubicin-induced electrocardiographic abnormalities. Results showed prolongation of QT and QTc intervals in the ML221 group.
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