Cytotoxic and Anti-angiogenic Effects of Postbiotics Derived from Pediococcus spp. against CT26 Mouse Colon Carcinoma Cells 

Umi Khalsom Mohd Bajuri¹, Kalavathy Ramasamy¹, Siong Meng Lim^1,*

Abstract

Introduction: The current pharmacological treatments against colorectal cancer (CRC) are often compromised by their side effects and cancer resistance. As the majority of CRC are sporadic in nature and associated with diet, current efforts have been directed towards alternative approaches which include probiotics. There is now evidence indicating the strain-dependent usefulness of probiotic-derived bioactive metabolites (i.e., postbiotics) against CRC. There are also reports on Lactobacillus and Bifidobacterium probiotics that inhibit tumour angiogenesis which drives the growth of CRC. As such, this study aimed at assessing the cytotoxic and anti-angiogenic potentials of cell free supernatant (CFS) fermented by two unique strains of lactic acid bacteria (LAB) isolated from fermented tapioca in vitro. Methods: LAB identification was performed by sequencing the 16S rRNA gene of the LAB and matching the consensus sequence from GeneBank. Cytotoxicity of the LAB were then screened by using Sulforhodamine B (SRB) assay. Immunocytostaining of CT26 cells treated with LAB-derived CFS was performed at their respective highest subtoxic concentration (IC₁₅) for examination of the anti-angiogenicity potential. Lastly, high performance liquid chromatography (HPLC) analyses was undertaken to determine the concentrations of four major short chain fatty acids (SCFA; i.e., acetate, butyrate, lactate and propionate) in the LAB-derived CFS. Results: The LAB were identified as Pediococcus pentosaceus LAB3 and Pediococcus acidilactici LAB4 (99% similarity), respectively. LAB4 emerged as the most potent Pediococcus sp. against CT26 cells with IC₅₀=4.67+0.3%. Immunocytostaining of CT26 cells treated with LAB-derived CFS (especially LAB4) resulted in downregulation (<6%) of pro-angiogenic vascular endothelial growth factor (VEGF) and upregulation (<42%) of anti-angiogenic thrombospondin (TSP-1). HPLC analyses found LAB4-derived CFS to yield the highest concentration for lactate and propionate. Conclusion: The present findings imply the potential strain-dependent cytotoxic and anti-angiogenic properties of LAB-derived postbiotics against CRC. The potent cytotoxic and anti-angiogenic effects of LAB4 may be associated with the production of SCFA. 

Keywords: Lactic acid bacteria, Colorectal cancer, Cytotoxicity, Angiogenesis, Short chain fatty acids

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