Investigating the effects of lactobacillus isolated from traditional dairy of Gilan province and its extract on the expression of AKT, mTOR, PTEN, CASP۳ genes in HT-۲۹ clone cancer cells

BACKGROUND AND OBJECTIVESTreating diseases with the help of microorganisms is a new and developing method that uses beneficial bacteria as therapeutic agents in immune disorders and infectious diseases. Probiotics are non-pathogenic microorganisms that, when prescribed in appropriate amounts, can prevent or improve some diseases by improving the health status. Therefore, the aim of this study is to investigate the effects of probiotics isolated from dairy products on HT۲۹.MATERIALS AND METHODSDairy samples from different regions of Gilan province were randomly purchased and collected in sterile containers, then MRS broth was used to isolate lactobacilli. Biochemical and microbial tests were used for the phenotypic identification of lactobacilli. Also, for the genotypic identification of Lactobacillus, using specific primers for the molecular identification of ۱۶SrRNA of Lactobacillus, DNA extraction kit and PCR of Lactobacillus bacteria were used to identify them. Sequencing was done. After preparation and cultivation of HT-۲۹ cells, HT-۲۹ were treated with Lactobacillus extract by MTT assay. HT-۲۹ cells were treated with Lactobacillus extract identified by MTT assay in different dilutions and the best effective dilution on the cells was determined. Finally, the expression of AKT, mTOR, PTEN and CASP۳ genes in cancer cell line was investigated by Real Time PCR.RESULTS AND DISCUSSIONDairy samples including cow's milk, cheese and local yogurt were collected from different regions of Gilan province. And using MRS-Agar medium, lactobacilli were isolated. DNA extraction was done using the kit. The results of investigating the treatment of HT-۲۹ cells with Lactobacillus extract by MTT assay. First, the range of IC۵۰ was determined. HT۲۹ cancer cells were treated with Lactobacillus helveticus extract at different concentrations of ۰.۵, ۱, ۱.۵ and ۲ μg/ml using the MTT test within ۲۴ hours, and the results of decreasing cell viability at high concentrations were reported as ۹۲.۲۴, ۸۵.۲۴, ۶۷.۲۴ and ۵۵.۲۴, respectively. Also, the results showed that the extract of Lactobacillus helveticus bacteria at a concentration of ۲ ug/ml has a ۵۰% killing effect. In order to determine the amount of apoptosis induced in HT-۲۹ cancer cell line by the extract of Lactobacillus helveticus bacteria, it was recorded as ۲.۴۷ for the CASP۳ gene, ۰.۷۶ for the AKT gene, ۲.۴۳ for the PTEN gene, and ۰.۹۸ for the mTOR gene, and this value was also recorded as ۱ for the GAPDH control gene. According to the results of the effect of Lactobacillus helveticus extract on HT۲۹ cancer cells, the expression of both genes increased and this effect was more on the expression of fas gene, which indicates an increase in apoptosis.CONCLUSIONThe results of this study showed that Lactobacillus, especially Lactobacillus helveticus, has a high potential to prevent cancer and can be used as an adjuvant treatment. They were also used in the dairy food industry to improve food quality.

کلیدواژه ها:

Lactobacillus ، traditional dairy products of Gilan province ، expression of AKT ، mTOR ، PTEN ، CASP۳ genes ، HT-۲۹ cells