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Excessive l-cysteine induces vacuole-like cell death by activating endoplasmic reticulum stress and mitogen-activated protein kinase signaling in intestinal porcine epithelial cells

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Abstract

High intake of dietary cysteine is extremely toxic to animals and the underlying mechanism remains largely unknown. This study was conducted to test the hypothesis that excessive l-cysteine induces cell death by activating endoplasmic reticulum (ER) stress and mitogen-activated protein kinase (MAPK) signaling in intestinal porcine epithelial cells. Jejunal enterocytes were cultured in the presence of 0–10 mmol/L l-cysteine. Cell viability, morphologic alterations, mRNA levels for genes involved in ER stress, protein abundances for glucose-regulated protein 78, C/EBP homologous protein (CHOP), alpha subunit of eukaryotic initiation factor-2 (eIF2α), extracellular signal-regulated kinase (ERK1/2), p38 MAPK, and c-Jun N-terminal protein kinase (JNK1/2) were determined. The results showed that l-cysteine (5–10 mmol/L) reduced cell viability (P < 0.05) and led to vacuole-like cell death in intestinal porcine epithelial cells. These adverse effects of L-cysteine  were not affected by the autophagy inhibitor 3-methyladenine. The protein abundances for CHOP, phosphorylated (p)-eIF2α, p-JNK1/2, p-p38 MAPK, and the spliced form of XBP-1 mRNA were enhanced (P < 0.05), whereas those for p-ERK1/2 were reduced (P < 0.05). Collectively,  excessive  l-cysteine induces vacuole-like cell death via the  activation of ER stress and MAPK signaling in small intestinal epithelial cells. These signaling pathways may be potential targets for developing effective strategies to prevent the toxicity of dietary cysteine.

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Abbreviations

CHOP:

C/EBP homologous protein

DDIT3:

DNA damage-inducible transcript 3

eIF2α:

Alpha subunit of eukaryotic initiation factor-2

ER:

Endoplasmic reticulum

ERK1/2:

Extracellular signal-regulated kinase

FBS:

Fetal bovine serum

GRP78:

Glucose-regulated protein 78

HSPA5:

Heat shock 70 kDa protein 5

HRP:

Horseradish peroxidase

IPEC-1:

Intestinal porcine epithelial cell line 1

JNK1/2:

c-Jun N-terminal protein kinase

MAPK:

Mitogen-activated protein kinase

PERK:

Protein kinase RNA-like endoplasmic reticulum kinase

UPR:

Unfolded protein response

XBP1s:

Spliced X-box binding protein 1

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Acknowledgments

This work was supported by the National Basic Research Program of China (No. 2013CB127302), the National Natural Science Foundation of China (No. 31172217, 31272450, 31272451), the Chinese Universities Scientific Fund (2013RC002), the Program for New Century Excellent Talents in University (NCET-12-0522), and the Program for Beijing Municipal Excellent Talents, the Agriculture and Food Research Initiative Competitive Grant from the USDA National Institute of Food and Agriculture (No. 2014-67015-21770), and Texas A&M AgriLife Research (H-8200).

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Authors and Affiliations

  1. State Key Laboratory of Animal Nutrition, Department of Animal Nutrition and Feed Science, China Agricultural University, 100193, Beijing, China

    Yun Ji, Zhenlong Wu, Zhaolai Dai, Kaiji Sun, Qing Zhang & Guoyao Wu

  2. Department of Animal Science, Texas A&M University, College Station, TX, 77843, USA

    Guoyao Wu

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  1. Yun Ji
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  2. Zhenlong Wu
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  3. Zhaolai Dai
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Correspondence to Zhenlong Wu.

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Ji, Y., Wu, Z., Dai, Z. et al. Excessive l-cysteine induces vacuole-like cell death by activating endoplasmic reticulum stress and mitogen-activated protein kinase signaling in intestinal porcine epithelial cells. Amino Acids 48, 149–156 (2016). https://doi.org/10.1007/s00726-015-2071-5

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