The study of the interactions between nutrients and intestine requires in vitro models mimicking as close as possible both the morphology and the physiology of the human intestinal epithelium. Many experimental difficulties hampered in establishing a physiological long-term experimental model starting from primary cultures of normal small intestinal and colon cells. For this reason, a Caco2/HT-29 (70/30) co-culture was set up in our lab starting from the differentiated parental cell populations. Both cell lines originate from a human colon adeno carcinoma but, when differentiated, Caco2 cells are mostly absorptive and do not secrete mucus, while HT-29 cells are a heterogeneous population, comprising scattered enterocyte elements and mucus secreting cells. The co-culture represents an in vitro model of the small human intestine as it concerns final digestion and absorption of digested foods. This experimental setting allowed also studying the effects induced by an excess of nutrients by changing the frequency of the medium administration. Two parallel experimental groups were cultured: the standard group (ST) and the excess group (EX). In ST group the culture medium was changed every four days, whilst in EX group on alternate days from confluence (T0). Co-culture was harvested at T0 and at 3, 7, and 15 days post-confluence (T3, T7, and T15, respectively). In comparison with the ST group, the EX group revealed a maintenance in the number of microvilli, an increase in follicle like-structures and mucus production, and a decrease in the number of tight junction. The specific activity of Alkaline Phosphatase, Aminopeptidase N, and of Dipeptidyl Peptidase-IV, known markers of intestinal and enterocyte differentiation, progressively increased. At T15 in the EX group an increased permeability to large molecules evidenced by the Transepithelial Electrical Resistance and the Lucifer yellow permeability was evident. At the same time, the level of ROS and NO production increased as well as of interleukin-6 and interleukin-8, all markers representing a low grade of inflammation. These results agree with the morpho-functional features associated to the intestine of overweight/obese animals. The two in vitro intestinal models represent the possibility to study the patho-physiology of the interactions between nutrients and human gut. Next step will be the completeness of the co-culture models with the microbiota.

Morpho-physiological correlations in presence of an excess of nutrients in a Caco2/HT29 co-culture / E. Donetti, A. Ferraretto. ((Intervento presentato al 93. convegno National Congress of the Italian Society of Experimental Biology (SIBS) tenutosi a online-Palermo nel 2021.

Morpho-physiological correlations in presence of an excess of nutrients in a Caco2/HT29 co-culture

E. Donetti;A. Ferraretto
2020

Abstract

The study of the interactions between nutrients and intestine requires in vitro models mimicking as close as possible both the morphology and the physiology of the human intestinal epithelium. Many experimental difficulties hampered in establishing a physiological long-term experimental model starting from primary cultures of normal small intestinal and colon cells. For this reason, a Caco2/HT-29 (70/30) co-culture was set up in our lab starting from the differentiated parental cell populations. Both cell lines originate from a human colon adeno carcinoma but, when differentiated, Caco2 cells are mostly absorptive and do not secrete mucus, while HT-29 cells are a heterogeneous population, comprising scattered enterocyte elements and mucus secreting cells. The co-culture represents an in vitro model of the small human intestine as it concerns final digestion and absorption of digested foods. This experimental setting allowed also studying the effects induced by an excess of nutrients by changing the frequency of the medium administration. Two parallel experimental groups were cultured: the standard group (ST) and the excess group (EX). In ST group the culture medium was changed every four days, whilst in EX group on alternate days from confluence (T0). Co-culture was harvested at T0 and at 3, 7, and 15 days post-confluence (T3, T7, and T15, respectively). In comparison with the ST group, the EX group revealed a maintenance in the number of microvilli, an increase in follicle like-structures and mucus production, and a decrease in the number of tight junction. The specific activity of Alkaline Phosphatase, Aminopeptidase N, and of Dipeptidyl Peptidase-IV, known markers of intestinal and enterocyte differentiation, progressively increased. At T15 in the EX group an increased permeability to large molecules evidenced by the Transepithelial Electrical Resistance and the Lucifer yellow permeability was evident. At the same time, the level of ROS and NO production increased as well as of interleukin-6 and interleukin-8, all markers representing a low grade of inflammation. These results agree with the morpho-functional features associated to the intestine of overweight/obese animals. The two in vitro intestinal models represent the possibility to study the patho-physiology of the interactions between nutrients and human gut. Next step will be the completeness of the co-culture models with the microbiota.
Settore BIO/16 - Anatomia Umana
Settore MED/49 - Scienze Tecniche Dietetiche Applicate
Italian Society of Experimental Biology (SIBS)
Morpho-physiological correlations in presence of an excess of nutrients in a Caco2/HT29 co-culture / E. Donetti, A. Ferraretto. ((Intervento presentato al 93. convegno National Congress of the Italian Society of Experimental Biology (SIBS) tenutosi a online-Palermo nel 2021.
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/2434/839825
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