Effects of Up-Regulated Expression of Heat Shock Protein 70 on Apoptosis in Intestinal Epithelial Cells of Heat-Stressed Piglets

SUN Huilei; ZHONG Qingzhen; LI Mingye; MOLAWA M N

doi:10.3969/j.issn.1006-267x.2020.04.046

2020 , Vol. 32 >Issue 4: 1867 - 1874

DOI: https://doi.org/10.3969/j.issn.1006-267x.2020.04.046

Effects of Up-Regulated Expression of Heat Shock Protein 70 on Apoptosis in Intestinal Epithelial Cells of Heat-Stressed Piglets

SUN Huilei ,
ZHONG Qingzhen ,
LI Mingye ,
MOLAWA M N

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1. College of Animal Science and Technology, Jilin Agricultural University, Changchun 130118, China;
2. Key Laboratory of Animal Production and Product Quality and Safety, Ministry of Education, Changchun 130118, China;
3. Jilin Province Key Laboratory of Animal Nutrition and Feed Science, Changchun 130118, China

Received date: 2019-09-30

Online published: 2020-04-16

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Abstract

This experiment was used glutamine as a heat shock protein 70 (HSP70) inducer, to investigate the effects of HSP70 on intestinal epithelial cells (IPEC-J2) of heat-stressed piglets through mitochondrial signal transduction pathway of apoptosis. In this experiment, IPEC-J2 cells cultured in vitro were used as the model, and different concentrations (1, 2, 4, 6, 8, 10 mmol/L) of glutamine were added to the culture medium for 24 h, the total RNA and total protein were extracted. The relative expressions of HSP70 mRNA and protein were detected using real-time fluorescent quantitative PCR and Western blot, and the optimal concentration of glutamine induced HSP70 expression was screened. The cells in logarithmic growth period were divided into three groups:control group (Con group, 37℃ cultured 12 h), heat stress group (Hs group, 42℃ cultured 12 h) and glutamine group (Gln group, 6 mmol/L glutamine and 42℃ cultured 12 h). Annexin V/PI double staining was used to detect the level of apoptosis, real-time fluorescence quantitative PCR and Western blot were used to detect the relative expressions of HSP70, apoptotic protease activating facter-1 (Apaf-1), cysteinyl aspartate specific proteinase-9 (Caspase-9), cysteinyl aspartate specific proteinase-3 (Caspase-3), B-cell lymphoma-2 (Bcl-2) mRNA and protein. The results showed that the up-regulated expression of HSP70 by glutamine firstly increased and then decreased, the relative expression of HSP70 mRNA and protein induced by 6 mmol/L glutamine was the highest, and significantly higher than that of other concentration (P<0.05), which served as the induction condition of Gln group in the follow-up experiment. Compared with the Con group, the apoptosis rate of Hs group was significantly increased (P<0.01), the relative expressions of HSP70, Apaf-1, Caspase-3, Caspase-9 mRNA and protein were significantly increased (P<0.01), while the relative expression of Bcl-2 mRNA and protein was significantly decreased (P<0.01). Compared with the Hs group, the apoptosis rate of Gln group was significantly decreased (P<0.01), and still significantly higher than that of the Con group (P<0.01); the relative expression of HSP70 mRNA and protein of Gln group was significantly increased (P<0.01); the relative expressions of Apaf-1, Caspase-3, Caspase-9 mRNA and protein of Gln group were significantly decreased (P<0.01), but significantly higher than those of the Con group (P<0.01); the relative expression of Bcl-2 mRNA and protein of Gln group was significantly increased (P<0.01), but significantly lower than that of the Con group (P<0.01). In conclusion, the up-regulated expression of HSP70 can effectively alleviate the apoptosis of intestinal epithelial cells of piglets induced by heat stress.

Key words： heat shock protein 70; IPEC-J2 cells; apoptosis; mitochondrial pathway; Caspase

Cite this article

SUN Huilei , ZHONG Qingzhen , LI Mingye , MOLAWA M N . Effects of Up-Regulated Expression of Heat Shock Protein 70 on Apoptosis in Intestinal Epithelial Cells of Heat-Stressed Piglets[J]. Chinese Journal of Animal Nutrition, 2020 , 32(4) : 1867 -1874 . DOI: 10.3969/j.issn.1006-267x.2020.04.046

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