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Methionine deficiency and its hydroxy analogue influence chicken intestinal 3-dimensional organoid development.

ABSTRACT: Methionine and its hydroxy analogue (MHA) have been shown to benefit mouse intestinal regeneration. The intestinal organoid is a good model that directly reflects the impact of certain nutrients or chemicals on intestinal development. Here, we aimed to establish a chicken intestinal organoid culture method first and then use the model to explore the influence of methionine deficiency and MHA on intestinal organoid development. The results showed that 125-μm cell strainer exhibited the highest efficiency for chicken embryo crypt harvesting. We found that transforming growth factor-β inhibitor (A8301) supplementation promoted enterocyte differentiation at the expense of the proliferation of intestinal stem cells (ISC). The mitogen-activated protein kinase p38 inhibitor (SB202190) promoted intestinal organoid formation and enterocyte differentiation but suppressed the differentiation of enteroendocrine cells, goblet cells and Paneth cells. However, the suppression of enteroendocrine cell and Paneth cell differentiation by SB202190 was alleviated at the presence of A8301. The glycogen synthase kinase 3 inhibitor (CHIR99021), valproic acid (VPA) alone and their combination promoted chicken intestinal organoid formation and enterocyte differentiation at the expense of the expression of Paneth cells and goblet cells. Chicken serum significantly improved organoid formation, especially in the presence of A8301, SB202190, CHIR99021, and VPA, but inhibited the differentiation of Paneth cells and enteroendocrine cells. Chicken serum at a concentration of 0.25% meets the requirement of chicken intestinal organoid development, and the beneficial effect of chicken serum on chicken intestinal organoid culture could not be replaced by fetal bovine serum and insulin-like growth factor-1. Moreover, commercial mouse organoid culture medium supplemented with A8301, SB202190, CHIR99021, VPA, and chicken serum promotes chicken organoid budding. Based on the chicken intestinal organoid model, we found that methionine deficiency mimicked by cycloleucine suppressed organoid formation and organoid size, and this effect was reinforced with increased cycloleucine concentrations. Methionine hydroxy analogue promoted regeneration of ISC but decreased cell differentiation compared with the results obtained with L-methionine. In conclusion, our results provide a potentially excellent guideline for chicken intestinal organoid culture and insights into methionine function in crypt development.

SUBMITTER: Wang Y

PROVIDER: S-EPMC8669257 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:This study was designed to compare the effect of methionine (Met) sources (DL-methionine [DLM] and DL-2-hydroxy-4-methylthio-butanoic acid [HMTBa]) and their supplementation levels on broiler growth performance and redox state. A 2 × 2 factorial arrangement was used with 2 sources (DLM and HMTBa) and 2 supplementation levels (0.05% and 0.25%) of Met. A total of 480 one-day-old broiler chicks were randomly divided into 4 treatments with 8 replicates per treatment (15 birds per replicate). The experiment lasted for 21 d. Broiler growth performance, redox capacity, redox-related genes expression, and Met transporters in different tissues were tested. Broilers fed high Met supplementation levels had improved (P < 0.05) body weight (BW), average daily gain (ADG) and feed conversion ratio (FCR). Similarly, broilers fed high Met levels had better (P < 0.05) antioxidant abilities in the serum, small intestine, and liver. Whereas, interactive effects (P < 0.05) were also observed between Met sources and levels. Compared with DLM, birds fed HMTBa diets had decreased (P < 0.05) total glutathione (T-GSH) and oxidized glutathione (GSSG) contents in duodenum, ileum, and liver. Similarly, broilers fed HMTBa supplemented diets had increased (P < 0.05) thioredoxin (Trx) gene expression in the duodenum and ileum, but decreased (P < 0.05) glutaredoxin (Grx), glutathione reductase (GSR), and glutathione synthetase (GSS) genes expression. Furthermore, lower gene expression of Na+ and Cl- dependent neutral and cationic amino acid transporter (ATB 0, + ), and Na+ dependent neutral amino acid transporter (B 0 AT) in the duodenum brush border, but higher gene expression of diamine acetyltransferase 1 (SAT1) and Na+-independent branched-chain and aromatic amino acid transporter (LAT1) in the jejunum and ileum basement membrane along with higher expression of the proton dependent monocarboxylate transporter 1 (MCT1) gene in the ileum were detected in birds fed HMTBa diets. In conclusion, DLM can be effectively used in glutathione synthesis to exert antioxidant functions, whereas HMTBa favors S-adenosylmethionine (SAM) synthesis and thus stimulates antioxidant-related genes expression.

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Methionine deficiency and its hydroxy analogue influence chicken intestinal 3-dimensional organoid development.

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