Pseudolaric acid B induced autophagy, but not apoptosis, in MRC5 human fibroblast cells

Abstract

Autophagy and apoptosis are closely associated. In previous studies, pseudolaric acid B (PAB), a diterpene acid isolated from the root and trunk bark of Pseudolarix kaempferi Gordon (Pinaceae), was demonstrated to induce apoptosis in various cell lines. However, in L929 murine fibrosarcoma and SW579 human thyroid squamous cell carcinoma cells, only autophagy was induced. In the present study, another cell line, MRC5 human lung fibroblast cells, was identified in which PAB only induced autophagy. The relationship between apoptosis and autophagy subsequent to PAB treatment in MRC5 cells was explored. When autophagy was inhibited by 3-methyladenine (3MA), apoptosis was induced in the PAB-treated MRC5 cells. To study the mechanism for the promotion of apoptosis by 3MA in the PAB-treated cells, the expression of members from the apoptotic signal pathways was assessed. As Bcl-2, Bcl-2 associated X and pro-caspase-9 expression following PAB treatment was not affected by 3MA treatment, it was determined that apoptosis was induced independent of the mitochondrial pathway of apoptosis. As Fas and pro-caspase-8 expression following PAB treatment were not altered by 3MA, it was further determined that the death receptor pathway was not induced. However, the phosphorylation of c-Jun-N-terminal kinase and the expression of pro-caspase-3 were upregulated, and the phosphorylation of extracellular signal-regulated kinase downregulated, by the combination of PAB and 3MA treatment compared with PAB alone. It was also observed that 3MA did not affect the microtubule aggregation ability of PAB. Therefore, inhibiting autophagy in MRC5 cells did not affect the role of PAB in microtubule aggregation, while apoptosis was induced. This may present a strategy to enhance the anti-tumor effects of PAB.

Pseudolaric acid B induces apoptosis in human rhabdomyosarcoma RD cells

“…PAB exhibits potent antitumor effects on human breast cancer MCF-7, cervical cancer HeLa and melanoma A375 cells by inducing apoptosis ( 1 – 5 ), and on murine fibrosarcoma L929, human thyroid squamous cell carcinoma SW579 cells and human lung fibroblasts MRC5 by inducing autophagy ( 6 – 8 ). In the present study, the effects of PAB were examined on human rhabdomyosarcoma RD cell proliferation.…”

“…The fluorescent compound MDC has been proposed as a tracer for autophagic vacuoles ( 6 – 8 ). RD cells (2×10 5 cells/well) were cultured in 6 wells plates for 24 h. 24 h later, RD cells were treated with 4 µM PAB for another 24 h and incubated with 0.05 mM MDC at 37°C for 1 h. Following incubation, the cells were washed once with PBS.…”

“…Pseudolaric acid B (PAB) is a diterpene-type acid isolated from the root and trunk bark of Pseudolarix kaempferi Gordon of the Pinaceae family, also termed ‘Tu-Jin-Pi’ in Traditional Chinese Medicine, which has been used to treat dermatological fungal infections ( 1 , 2 ). PAB exerts potent inhibition of cell proliferation in vitro in various tumor cell lines by inducing cell cycle arrest and apoptosis in human melanoma cell line A375-S2 and human breast cancer cell line MCF-7 ( 1 – 5 ) or autophagy in murine fibrosarcoma cell line L929 and human thyroid squamous cell line SW579 ( 6 – 8 ). Rhabdomyosarcoma is the most common type of soft tissue sarcoma in children worldwide and represents a high-grade neoplasm of skeletal myoblasts, this tumor accounts for 5–10% of all childhood tumors since 2002 ( 9 ).…”

“…Under certain circumstances, this process may promote cell death and morbidity; however, under the majority of circumstances, autophagy promotes survival by adjusting the cellular response to stress conditions ( 16 , 17 ). Following the induction of apoptosis or autophagy, cell cycle arrest is observed ( 6 – 8 , 18 , 19 ). Cell cycle arrest is a tumor-suppressive mechanism, as the cells do not enter the next phase of the retarded phase ( 4 , 7 , 12 ) and thus, cell proliferation is inhibited.…”

Effect of ingredients from Chinese herbs on enterovirus D68 production

“…Then, it was investigated that G2/M phase‐related drugs inhibited EVD68 production, it was found that pseudolaric acid B (Song et al, ; Wang et al, ), oridonin (Zheng et al, ), and erianin (Wang et al, ) did not affect intracellar viral protein expression although they had the trend to decrease the expression, therefore, it was speculated that the decreased trend was due to decreased viral mRNA level by pseudolaric acid B, oridonin, and erianin, respectively. Meanwhile, only oridonin affected supernatant viral protein expression, but all of them decreased supernatant viral genome level, and the fold difference in the TCID50/ml and the supernatant viral genome level between the control‐treated and pseudolaric acid B‐treated cells was 2.34 × 10 2 and 11.31, oridonin was 5.10 × 10 3 and 54.05, and erianin was 1.23 × 10 3 and 108.70, therefore, pseudolaric acid, oridonin, and erianin inhibiting viral genome packaged into virions partly explained the decreased production.…”