Inhibition of Testudinis Carapax et Plastrum extracts targeting BMP4 pathway on PC12 cell apoptosis / 中草药

ABSTRACT

Objective: To observe the protection of Testudinis Carapax et Plastrum extracts (TCPE) on serum starvation-induced PC12 cell apoptosis and explore its mechanism. Methods: The PC12 apoptosis model was established by serum starvation for 3 d. The cells were randomly divided into four groups control group, model group, low-dose and high-dose (3 and 30 μg/mL) TCPE groups. In the three days of the treatment, cell absorbance was determined by MTT, ratio of cell apoptosis was examined by Annexin V/PI double stain flow cytometry (FCM), Caspase-3, BMP4, BMPR-IA, and p-Smad1/5/8 signaling molecular expression were detected by Western blotting, and the anti-apoptotic effect of TCPE was observed after blocking BMPs signal pathway. Semi-quantitative analysis of bands was carried out by Bio-Rad Quantity One gel analysis system. Results: MTT and FCM analyses demonstrated that TCPE could increase PC12 cell viability and decrease their apoptotic ratios in a dose dependent manner. Western blotting results showed that TCPE could decrease Caspase-3 expression, promote the expression of BMP4, BMPR-IA, and p-Smad1/5/8. There was statistically significant difference between TCPE (3 and 30 μg/mL) groups and model group (P<0.05, P<0.01) in all above results. While TCPE had no effect on the expression of BMP2, BMP7, and BMPR-II. BMPR-IB hadn't been detected. The anti-apoptotic activity was partially mitigated by neutralizing BMP4 antibody. Conclusion: TCPE has the capacity to inhibit the apoptosis of PC12 induced by serum starvation in a dose dependent manner and its mechanism may be associated with partially activating and up-regulating the expression of BMP4 signaling pathway.