GDF11/BMP11 as a novel tumor marker for liver cancer

Abstract

Growth differentiation factor 11 (GDF11), also known as bone morphogenetic protein 11, a member of the transforming growth factor-β superfamily, has been reported to be involved in colorectal cancer. However, the roles of GDF11 in Chinese patients with liver cancer and the underlying mechanisms have remained elusive. The present study assessed the expression of GDF11 in 10 paired samples of cancerous and normal tissues from Chinese liver cancer patients. The results indicated that the expression of GDF11 was significantly lower in cancerous tissues than in normal tissues. In vitro, the expression of GDF11 was reduced in a panel of liver cancer cell lines compared with that in a normal liver cell line at the mRNA and protein level. Treatment with GDF11 reduced the viability of HepG2 for up to 72 h and GDF11 treatment reduced the viability of SMMC-7721 after 48 and 72 h. Furthermore, GDF11 activated Smad2/3 signaling in HepG2 cells. In conclusion, GDF11 has a tumor suppressor role in liver cancer, exerts its effects through Smad2/3 signaling and may serve as a novel tumor marker in liver cancer diagnosis.

Keywords: Smad2/3; bone morphogenetic protein 11; growth differentiation factor 11; liver cancer; tumor marker.

Figure 1.

The expression of GDF11 is decreased in liver cancer tissue at the mRNA level and is downregulated in liver cancer cells. (A) GDF11 mRNA expression in 10 paired samples of cancerous and normal liver tissue in Chinese patients with liver cancer. Cancer tissues had significantly lower GDF11 mRNA expression than normal tissues. (B) Oncomine data indicating GDF11 expression in normal vs. cancerous liver tissues. (C) GDF11 mRNA expression in a normal liver cell line and liver cancer cell lines (HepG2 and SMMC-7721). (D) GDF11 protein expression in a normal liver cell line and liver cancer cell lines (HepG2 and SMMC-7721). Data are presented as the mean ± standard error of the mean. *P<0.05 vs. normal tissues (n=10); **P<0.01 vs. normal tissues (n=19); ***P<0.001 vs. normal liver cell line. GDF11, growth differentiation factor 11.

Figure 2.

GDF11 increases the level of p-Smad2/3 protein in HepG2 cells. (A) Recombinant GDF11 increased the protein expression of GDF11 in SMMC-7721 and HepG2 cells. Cells in the control group were treated with the same volume (5 µl per cell culture flask) of sterile ultra-pure water. (B) HepG2 cells treated with GDF11 (50 and 100 ng/ml) for 15 min and 1 h. The phosphorylation of Smad2/3 was increased after GDF11 treatment in HepG2 cells. (C) The activation of Smad3 by GDF11 was attenuated in the presence of the Smad3 inhibitor SIS3 (5 µM). β-actin was used as an internal reference. *P<0.05, **P<0.01 and ***P<0.001 vs. CTL group (n=5). CTL group is normalized to 1. GDF11, growth differentiation factor 11; CTL, control; p-Smad3, phosphorylated Smad3.

Figure 3.

Treatment with GDF11 reduced the viability of HepG2 for up to 72 h and GDF11 treatment reduced the viability of SMMC-7721 after 48 and 72 h. (A) HepG2 cells treated with GDF11 (50 and 100 ng/ml) for 24, 48 and 72 h. The cell viability was decreased following 72 h of incubation, while it remained unchanged at 24 and 48 h. (B) SMMC-7721 cells treated with GDF11 (50 and 100 ng/ml) for 24, 48 and 72 h. The cell viability was decreased following 48 and 72 h of treatment, while it remained unchanged for 24 h. ***P<0.001 vs. CTL group (n=24). GDF11, growth differentiation factor 11; CTL, control.

Figure 4.

Schematic of GDF11 signaling in human liver cancer. GDF11 first binds to ActRIIB to recruit the type I ALK receptors ALK4, ALK5 and ALK7. The downstream pathways of GDF11 binding to its receptors include Smad and non-Smad pathways. The Smad pathway is the canonical signaling for transforming growth factor-β superfamily members. GDF11 first activates receptor Smads, including Smad2/3 and Smad1/5/8 signals. After phosphorylation, the receptor Smads are released from the receptor and recruit the common Smad4 to form a complex, which then migrates into the nucleus and activates the transcription of specific target genes that have a role in liver cancer. GDF11, growth differentiation factor 11; p, phosphate; ActRIIB, activin receptor type IIB; ALK, activin receptor-like kinase.