内质网应激PERK-eIF2α-ATF4信号通路在延缓APP/PS1小鼠移植瘤生长中的作用

doi:10.13481/j.1671-587X.20220208

Abstract

Abstract: Objective

To investigate the inhibitory effects of endoplasmic reticulum stress（ERS） and endoplasmic reticulum autophagy on the growth of transplanted melanoma model in amyloid precursor protein（APP）/presenilin 1（PS1）mice， and to clarify the possible mechanism of its inhibitory effect of protein kinase R-like endoplasmic reticulum（PERK）-eukaryotic translation initiation factor 2α（eIF2α）-activating transcription factor 4（RATF4） pathway.

Methods

The C57BL/6J（C57） and APP/PS1 mice were transplanted with melanoma B16 cells to establish the C57 transplanted tumor and APP/PS1 transplanted tumor models of male mice），and used as C57 transplanted tumor group （n=7） and APP/PS1 transplanted tumor group （n=7）. The tumor appearance time was observed and the tumor volume was calculated of the mice in two groups. The expression levels of glucose regulatory protein 78（GRP78），PERK and lysosomal cathepsin L （cathepsin L） mRNA in the transplanted tumor tissue of the mice in two groups were detected by real-time fluorescence quantitative PCR（RT-PCR）.The expression levels of GRP78， PERK， phos-PERK， eukaryotic translation initiation factor 2α （eIF2α），p-eIF2α， ATF4， and FAM134B in the transplanted tumor tissue of the mice in two groups were detected by Western blotting method. The expression levels of protein disulfide isomerase（PDI） protein in two groups were detected by immunohistochemistry and the adenosine triphoshate（ATP） levels in the transplanted tumor tissue of the mice were determined by fluorescence assay.

Results

Compared with C57 transplanted tumor group， the tumor appearance time of the mice in APP/PS1 transplanted tumor group was late， and the tumor volume was decreased（P<0.05）；the expression levels of GRP78 and PERK mRNA in the transplanted tumor tissue were increased （P<0.05）， and the expression levels of GRP78， p-PERK/PERK， p-eIF2α/eIF2α and ATF4 proteins were increased（P<0.05）. In C57 transplanted tumor group， melanoma granules were found in the cytoplasm of the tumor cells， which was the morphological characteristic of transplanted tumor tissue， and a small amount of brown granules were found， which were PDI positive granules. In APP/PS1 transplanted tumor group， a small amount of melanoma granules and widely expressed brown granules were found in the cytoplasm of tumor cells. Compared with C57 transplanted tumor group， the expression level of FAM 134B protein in the transplanted tumor tissue of the mice in APP/PS1 transplanted tumor group was increased（P<0.05）， the expression level of cathepsin L mRNA was increased（P<0.05），and the ATP level was decreased（P<0.05）.

Conclusion

The growth of tumor in the APP/PS1 transplanted tumor model mice is slow， and its mechanism may be related to the activation of endoplasmic reticulum stress PERK-eIF2α-ATF4 signaling pathway and regulated endoplasmic reticulum autophagy.

Key words: Protein kinase R-like endoplasmic reticulum kinase, Endoplasmic reticulum stress, Endoplasmic reticulum autophagy, Amyloid precursor protein, Presenilin 1, Transplanted melanoma

CLC Number:

R739.5

Ying DONG,Jianyu GUO,Siyi WANG,Dan GUO,Like WANG,Xu WEN,Lifeng LIU,Meng QU,Chunyan YU,Nannan LIU,Dan WANG,Changjie CHEN. Effect of endoplasmic reticulum stress PERK-eIF2α-ATF4 signaling pathway on delaying transplanted tumor growth in APP/PS1 mice[J].Journal of Jilin University(Medicine Edition), 2022, 48(2): 324-330.

Figures/Tables 5

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Tab.2

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Gene	Primer
GRP78	Forward: 5′-CGCGGATCCGAGGAGGAGGACAAG-3′ Reverse: 5′-CCGCTCGAGCTACCTAACATCTTTACC-3′
PERK	Forward: 5′-AGTCCCTGCTCGAATCTTCCT-3′ Reverse: 5′-TCCCAAGGCAGAACAGATATACC-3′
Cathepsin L	Forward: 5′-CGGAGGAGTCTTACCCCTAT-3′ Reverse: 5′-CTACCCATCAATTCACGACA-3′
GAPDH	Forward: 5′-TTGTGATGGGTGTGAACCACGAGA-3′ Reverse: 5′-CATGAGCCCTTCCACAATGCCAAA-3′

Group	GRP78 PERK
C57 transplanted tumor	1.00±0.00 1.00±0.00
APP/PS1 transplanted tumor	1.39±0.29^* 1.26±0.15^*

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Effect of endoplasmic reticulum stress PERK-eIF2α-ATF4 signaling pathway on delaying transplanted tumor growth in APP/PS1 mice

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