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This study was carried out at the Microbial Genetic laboratory, at Microbial Genetics Department, Genetic Engineering and Biotechnology Research Division, National Research Center (NRC), Giza, Egypt, and the Molecular Genetics Laboratories of the Department of Genetics, Faculty of Agriculture, Ain Shams University, Egypt, during the period from 2014 to 2019.
The main results of this study were:
1. Selected Knock-out haploid yeast strains were treated with different concentrations of the three food additives (MSG, SB and Saffron) under study for 24-48 hrs. YKO strains confirmed the occurrence of cancer formation due to administration of these food additives that could be caused according to different genotypes. this might give an overview of the relationship between those additives and cancer formation.
2. Comet assay was performed to assess DNA damage in yeast deletion strains after treatment by recommended concentrations of food additives to YKO strains exhibited a significant DNA damage (P < 0.05) that was observed by an increase in tail length, tail DNA% and tail moment as compared to the normal control strains.
3. Four different types of human cell lines; namely, colon carcinoma (Caco-3), breast carcinoma (MCF7), lung carcinoma (A549) and normal lung cell line (Wi38) were treated with the same concentrations of food additives under study for 24 hrs. Viability in shapes of the cells showed considerable variations between control and treatment by NRU assay and effect of food additives on them and confirmed the carcinogenic effect of these additives.
4. Cytotoxicity and programmed cell death (apoptosis) of food additives were tested on in vitro human cell growth. Cell cycle arrest and apoptosis-related genes of the CoCa-3 cell line were also evaluated. The results revealed that food additives stimulated cytotoxicity by decreased cell viability in cancer and normal cell lines. Cell cycle blocking at the G2/M phase was evident.
5. Real-Time PCR of cancer-related genes of cell line homologous with selected genotypes were done. Results of gene expression of selected genotypes of human genes showed that the effect of additives could inflict deleterious and pronounced health hazards in human including cancer.
6. Yeast haploid (knockout) strains genotypes were selected according to alignment between human and yeast genes sequence homology results (E-value and Query covery). Genes alignment between human genes and yeast genes in Clustal Omega web site were carried out.
7. Prediction of protein-protein interaction was done by Gene MANIA web site. Gene MANIA indicate the predictive value of each selected data set for the query. Two organisms are currently supported (Homo sapiens and Saccharomyces cerevisiae) as well or better than other proteins function prediction methods on yeast and human. Gene MANIA showed that the resulting networks are different, with different interactions and four different predicted proteins in yeast and four different predicted in human. The network between proteins in yeast which are related to cancer and then homology in human proteins.
8. In conclusion, These studies provided evidence that food additives can inhibit cell proliferation in the human cell lines and also shed light on dose-dependent cellular cytotoxicity consequences of exposure to this food additives. Our findings detected the toxic and apoptotic effects of food additives in human cell lines. Thus, the use of MSG, SB and Saffron could present negative health biohazards to human beings. This calls for strict control of the threshold levels that should be permitted for the industrial products containing food additives as it could reflect biosafety hazards to human and environment.