Original underlying data for Fig 6.

Correction: Fuling Granule, a Traditional Chinese Medicine Compound, Suppresses Cell Proliferation and TGFβ-Induced EMT in Ovarian Cancer</p

CFG suppresses ovarian cancer cell proliferation <i>in vitro</i>.

<p>SRB assays (A) and MTT experiments (B) to show the inhibitory effect of CFG on HEY and SKOV3 cell growth after treatment with different concentration of CFG. (C) Crystal violet staining to show the colony formation ability of CFG-treated HEY and SKOV3 cells compared with control. Representative images are presented in the panel. The data represent mean ± SD of three experiments.</p

CFG downregultates AKT/GSK3β signal pathway and EMT markers.

Cells were treated with 3 mg/ml CFG only or in combination with 10 ng/ml TGF β1 for 24 h. The cellular proteins, including E-cadherin, N-Cadherin, Vimentin, Fibronectin, β-Catenin, pAKT, AKT, pGSK3β, GSK3β, SNAIL, and SLUG, were detected by Western blot.</p

CFG changes cell cycle distribution and cell cycle-related protein expression in ovarian cancer cell lines <i>in vitro</i>.

<p>HEY (A) and SKOV3 (B) cell lines were treated with CFG 3 mg/ml for 24 h and the cell cycle distribution was analyzed by flow cytometry. (C and D) Quatitative PCR to detect the mRNA expression of cell cycle assoicated protein. HEY cells (C) and SKOV3 cells (D) were treated with 3 mg/ml CFG for 24 h. (E) HEY cells were treated with 3 mg/ml CFG for 24h and 48 h and then stained with immunofluorescen labeled antibodies against Cdt1 (red, G1 phase) and Geminin (green, G2 phase). Representative images are presented. (F) Cell cycle-related proteins were detected by western blot in HEY and SKOV3 cells treated with 3 mg/ml CFG for 24h.</p

Diversity of endophytic bacteria of <i>Dendrobium officinale</i> based on culture-dependent and culture-independent methods

Culture-dependent and culture-independent methods were compared and evaluated in the study of the endophytic diversity of Dendrobium officinale. Culture-independent methods consisted of polymerase chain reaction–denaturing gradient gel electrophoresis (PCR-DGGE) and metagenome methods. According to the results, differences were found between the three methods. Three phyla, namely Firmicutes, Proteobacteria, and Actinobacteria, were detected using the culture-dependent method, and two phyla, Firmicutes and Proteobacteria, were detected by the DGGE method. Using the metagenome method, four major phyla were determined, including Proteobacteria (76.54%), Actinobacteria (18.56%), Firmicutes (2.27%), and Bacteroidetes (1.56%). A distinct trend was obtained at the genus level in terms of the method and the corresponding number of genera determined. There were 449 genera and 16 genera obtained from the metagenome and DGGE methods, respectively, and only 7 genera were obtained through the culture-dependent method. By comparison, all the genera from the culture-dependent and DGGE methods were contained in the members determined using the metagenome method. Overall, culture-dependent methods are limited to ‘finding’ endophytic bacteria in plants. DGGE is an alternative to investigating primary diversity patterns; however, the metagenome method is still the best choice for determining the endophytic profile in plants. It is essential to use multiphasic approaches to study cultured and uncultured microbes.</p

CFG treatment reduced tumor growth and metastasis in vivo xenograft mouse.

<p>(A) The photos of the tumor tissues dissected from the mice on day 44 after injection and with aforementioned treatment. (B) The tumor dynamic growth curves of different groups. (C) Tunel assay to show the apoptotic cells in the tumor tissues with indicated treatment at 200× magnification. Lower layer shows the percentage of apoptotic cells in each group. For A, B, and C, SKOV3: SKOV3 control group of subcutaneous injection; CFG: CFG group of subcutaneous injection; Cisplatin: Cisplatin group of subcutaneous injection. (E) H&E staining showed the lung metastases and immunochemistry staining to show the expression of E-Cadherin and N-Cadherin in each group. SKOV3: SKOV3 control group of tail intravenous injection; CFG: CFG group of tail intravenous injection; Normal lung: Normal group for tail intravenous injection. *<i>p</i> < 0.05 and **<i>p</i> < 0.01</p

CFG inhibits the mobility of HEY and SKOV3 cells.

<p>(A and B) The wound healing assay was used to demonstrate the migration ability of HEY and SKOV3 cells, treated with 3mg/ml CFG only or in combination with 10 ng/ml TGF β1. Cell migration rate was quantitatively evaluated by measuring the cell covered area (C and D). Data are presented as mean ± SD. The experiments were repeated at least three times. *<i>p</i> <0.05 compared with the control. **<i>p</i> <0.01 compared with the control.</p

Fuling Granule, a Traditional Chinese Medicine Compound, Suppresses Cell Proliferation and TGFβ-Induced EMT in Ovarian Cancer

<div><p>The compound fuling granule (CFG) is a traditional Chinese drug which has been used to treat ovarian cancer in China for over twenty years. Nevertheless, the underlying molecular mechanism of its anti-cancer effect remains unclear. In this study, microarray data analysis was performed to search differentially expressed genes in CFG-treated ovarian cancer cells. Several cell cycle and epithelial-mesenchymal transition (EMT) related genes were identified. The microarray analyses also revealed that CFG potentially regulates EMT in ovarian cancer. We also found that, functionally, CFG significantly suppresses ovarian cancer cell proliferation by cell cycle arrest, apoptosis and senescence and the AKT/GSK-3β pathway is possibly involved. Additionally, the invasion and migration ability of ovarian cancer induced by TGFβ is significantly suppressed by CFG. In conclusion, our results demonstrated that CFG suppresses ovarian cancer cell proliferation as well as TGFβ1-induced EMT <i>in vitro</i>. Finally, we discovered that CFG suppresses tumor growth and distant metastasis in vivo. Overall, these findings provide helpful clues to design novel clinical treatments against cancer.</p></div

CFG induces apoptosis and senescence in ovarian cancer cells in vitro.

<p>(A) HEY and SKOV3 cell lines were treated with 3 mg/ml CFG for 24 h. The percentage of apoptotic cells were measured by flow cytometry using the tunnel apoptosis detection assay. (B and C) Apoptosis-related genes, including Bad, Bcl-Xl, Caspase 7 were detected by quantitative PCR and western blot. (D) β-gal Staining was performed to demonstrate the effect of 3 mg/ml CFG treatment on HEY and SKOV3 cell senescence.</p

CFG decreases TGF- β 1-induced invasion and migration of HEY and SKOV3 cells in vitro.

<p>HEY cells (A) and SKOV3 cells (B) were treated with 3mg/ml CFG only or in combination with 10 ng/ml TGF β1 for 24 h prior to use and the invasion and migration assays were then performed. Crystal violet OD values represent the amounts of invated and migrated HEY cells (C) and SKOV3 cells (D). Data is presented as mean ± SD. The experiments was repeated at least three times. * <i>p</i> <0.05 compared with the control. **<i>p</i> <0.01 compared with the control.</p