Effect Of Yellow Food Colorants On Proliferation And Differentiation Of Small Intestinal Cells

Synthetic colorants are widely used in food manufacture.Compared to natural colorants,the safety of synthetic colorants is more controversial.In brain,kidney,liver etc,synthetic colorants have been linked to some adverse health effects such as neurotoxicity,genotoxicity,carcinogenicity and allergenicity little is known how colorants impact on intestinal heath.This study evaluated the effects of the different yellow food colorants on the growth of intestinal organoids and explored the possible mechanisms.With the in vivo dextran sulphate sodium?DSS?-induced inflammatory model,the effect of colorants on inflammation responses was explored.The main research contents and results are as followed:?1?The small intestinal organoids were used as the in vitro model,they were exposed to different colorants for two generations to mimic the chronic exposure that could result from daily consumption of food.With the measurement of organoids growth rates and the assessment of cell viability,the results showed that after cultured with 2?g/mL curcumin,lutein,hydroxysafflor yellow A?the main active ingredient in carthamin?,lemon yellow and quinoline yellow respectively,the budding rates,expansion rates,number of crypts per organoids and the cell viability of organoids had no significantly difference between control group,which showed that these colorants had no effect on the growth of small intestinal organoids.Meanwhile,SY exhibited an effect on the secondary organoids,that remarkably inhibited the growth of organoids with decreasing the budding rate,expansion rate,numbers of crypts per organoid and the cell viability?p?0.001?.?2?Evaluated the effect of sunset yellow on proliferation and differentiation of intestinal cells by cell lineage analysis and hormone detection.The cell lineage assessment at gene levels showed that with the treatment of 2?g/mL sunset yellow for two generations,compared to control group,the expression of stem cell markers and the secretory cell makers in sunset yellow treated group decreased significantly,the expression of absorptive cells makers increased remarkably in sunset yellow treated group.The immunofluorescence results were matched with the qRT-PCR results,that in sunset yellow treated group,the numbers of stem cells marked with Olfm4 and the secretory cells makers marked with Chromogranin A?ChgA?,Lysozyme?Lyz?and Mucin2?Muc2?decreased significantly,and the number of absorptive cells marked with Villin increased significantly,comparing to control group.At the same time,the amount of glucagon-like peptide 1?GLP-1?detected in culture medium was significantly lower compared to control group?p<0.05?.These results proved that sunset yellow can disturbe the homeostasis of the intestinal epithelium by inhibiting the proliferation and affecting the differentiation of intestinal cells,which in turn disrupted hormone secretion.?3?To explore the mechanism of effect of sunset yellow on proliferation and differentiation of intestinal cells,we evaluated the level of endoplasmic reticulum stress and oxidative stress in sunset yellow treated organoids.In sunset yellow treated group,the expression of unfolded protein response targets Xbp1s,Atf4 and Grp78 gene elevated?p?0.001?,and the expression of Chop had no significantly difference with it in control group,indicating that the endoplasmic reticulum stress level raised and it tended to survival but not apoptosis.Meanwhile in sunset yellow treated organoids,the amount of reactive oxygen species?ROS?,malondialdehyde?MDA?and hydrogen peroxide?H₂O₂?increased?p?0.05?,the activity of superoxide dismutase?SOD?reduced?p?0.05?,illustrating that the level of oxidative stress raised.As the changes of sunset yellow treated organoids were closed to the changes of organoid under endoplasmic reticulum stress,and endoplasmic reticulum stress and oxidative stress can interact with each other.Therefore,it might be endoplasmic reticulum stress and oxidative stress that give rise to the effect of sunset yellow on proliferation and differentiation of intestinal epithelium cells.?4?The DSS induced inflammation mouse model was used to evaluated the effect of sunset yellow on inflammation responses.After treated with 2.5%DSS,compared to the mice fed with normal diet,the mice fed with 40mg/kg diet sunset yellow had significantly higher disease activity index values?p?0.01?,shorter colonic length?p?0.01?,reduced height of villi and crypts in ilea?p?0.001?,higher expression level of IL-1?and TNF-??p?0.01?,indicating that sunset yellow improved the DSS-induced inflammation level in mice.The results of this study indicated that curcumin,lutein,hydroxysafflor yellow A,tartrazine and quinoline yellow had no effect on the growth of small intestinal organoids at low concentration.Meanwhile,the treatment of 2?g/mL sunset yellow can inhibit proliferation and affect the differentiation of intestinal cells,which can disturb the intestinal epithelial homeostasis.The effect of sunset yellow on proliferation and differentiation of intestinal cells may be caused by endoplasmic reticulum stress and oxidative stress.In addition,long-term consumption of sunset yellow may increase the risk of intestinal inflammation.