The Effects of Ganoderma lucidum Treatment in an Optimised Colorectal Cancer Co-culture Cell Model

Abstract

Background: Ganoderma lucidum (GL) is a mushroom used in traditional Chinese medicine to promote health and longevity of humans. Its major bioactive components, triterpenes and polysaccharides have been associated with cytotoxic and immune boosting effects. The use of GL as an adjunct in colorectal cancer treatments had shown favourable immune-associated gene expression changes in patients. However, treatment with GL that contributes to these changes in the gut have yet to be studied. Objectives: The aim of this study was to investigate whether treatment with GL extracts, in a co-culture cell model utilised to mimic an in vivo setting to allow cross-talk between the gut epithelium and commensal bacteria, would stimulate expression changes in selected immunerelated and anti-cancer genes. Methods: An optimised co-culture cell model was developed with two colorectal carcinoma cell lines HT-29 and Caco-2. Two cytotoxicity assays using water soluble tetrazolium-1 and sulforhodamine B were performed to determine 50% inhibitory concentrations (IC50) of the GL extracts in both cell lines. The growth rate and concentration of the Lactobacillus plantarum ATCC 8014 (L.p.) strain were also determined and added to the model. Lastly, relative quantitative expression of candidate genes in HT-29 and Caco-2 cells were determined using RT-qPCR. Results: Cytotoxicity testing showed that the GL malt whiskey (GL(W)) extract inhibited the proliferation of the colorectal cancer cell lines tested with 3-day IC50s of 8.54 ±0.82 μL/well on HT-29 cells and 30.02 ±2.27 μL/well on Caco-2 cells. The stationary growth phase of L.p. was reached at 12 h post-incubation and a concentration of 109 CFU/well was added into the co-culture model. Inconsistent results in the expression of the COX-2 gene by the GL(W) extract in the HT-29 and Caco-2 cell lines were obtained. In contrast, the GL(W) extract induced an up-regulation of the c-Myc gene on HT-29 cells, whereas the expression of this gene was down-regulated on Caco-2 cells. However, these fold changes for the c-Myc gene were small. Conclusion: Overall, the results obtained in this study suggest that the GL(W) extract had an effect on the COX-2 and c-Myc gene expression in the HT-29 and Caco-2 cell lines. The anticancer effects of GL that are evident in the literature, suggests that the GL(W) extract may exert its effects by means of mediating the gut immune system.