Paeoniflorin inhibits tributyltin chloride-induced apoptosis in hypothalamic neurons via inhibition of MKK4-JNK signaling pathway

ETHNOPHARMACOLOGICAL RELEVANCE: Paeoniflorin (PF) exerts a significant protective effect against neurotoxicity and mitochondrial damage in neurons. However, the mechanisms underlying PF-mediated rescue remain elusive. Therefore, we endeavored to further research the molecular mechanisms underlying PF-mediated inhibition of tributyltin chloride (TBTC)-induced apoptosis of neurons.

AIM OF THE STUDY: To investigate the influence and possible mechanism of action of PF in TBTC-induced neurodegenerative disease.

MATERIALS AND METHODS: First, primary hypothalamic neurons were treated with tributyltin chloride (150 μg/L) and PF (25, 50, and 100 μM). 17β-estradiol (1 nM) was used as a positive control. Subsequently, CCK-8 assay was performed. The level of apoptosis was examined by flow cytometry and the function of mitochondria was reflected by MMP levels. The mRNA expression levels of B-cell lymphoma-2 (Bcl-2), together with Bax, were examined using qRT-PCR. The protein levels of mitogen-activated protein kinase kinase 4 (MKK4), c-Jun N-terminal kinase (JNK), Bcl-2, Bax, and Caspase-3 were examined using western blotting. Finally, pretreatment with JNK agonist, anisomycin, was done to observe the change in expressions of MKK4 and JNK.

RESULTS: Paeoniflorin treatment reduced TBTC-induced damage and neuron loss in a dose-dependent manner. Decrease in mitogen-activated protein kinase (MAPK) as well as JNK levels were reversed by treatment with paeoniflorin via inhibition of JNK activation. Furthermore, ratio of levels of Bcl-2/Bax increased while the activation of caspase-3 was suppressed. In addition, pretreatment with JNK agonist, anisomycin effectively suppressed TBTC-induced cytotoxicity in hypothalamic neuron.

CONCLUSIONS: PF can potentially be used to prevent and/or treat neurodegenerative diseases and neural injury by inhibiting MKK4-JNK signaling pathway.