Proteomic changes by radio-mitigative thrombopoietin receptor agonist romiplostim in the blood of mice exposed to lethal total-body irradiation

Abstract

Purpose

The thrombopoietin receptor agonist romiplostim (RP) is a therapeutic agent for immune thrombocytopenia that can achieve complete survival in mice exposed to a lethal dose of ionizing radiation. The estimated mechanism of the radio-protective/mitigative effects of RP has been proposed; however, the detailed mechanism of action remains unclear. This study aimed to elucidate the mechanism of the radio-protective/mitigative effects of RP, the fluctuation of protein in the blood was analyzed by proteomics.

Materials and Methods

Eight-week-old female C57BL/6J mice were randomly divided into 5 groups; control at day 0, total-body irradiation (TBI) groups at day 10 and day 18, and TBI plus RP groups at day 10 and day18, consisting of 3 mice per group, and subjected to TBI with 7 Gy of ¹³⁷Cs γ-rays at a dose rate of 0.74 Gy/min. RP was administered intraperitoneally to mice at a dose of 50 µg/kg once daily for 3 days starting 2 hours after TBI. On day 10 and day 18 after TBI, serum collected from each mouse was analyzed by liquid chromatography tandem mass spectrometry.

Results

Nine proteins were identified by proteomics methods from 269 analyzed proteins detected in mice exposed to a lethal dose of TBI: keratin, type II cytoskeletal 1 (KRT1), fructose-1, 6-bisphosphatase (FBP1), cytosolic 10-formyltetrahydrofolate dehydrogenase (ALDH1L1), peptidyl-prolyl cis-trans isomerase A (PPIA), glycine N-methyltransferase (GNMT), glutathione S-transferase Mu 1 (GSTM1), regucalcin (RGN), fructose-bisphosphate aldolase B (ALDOB) and betain–homocysteine S-methyltransferase 1 (BHMT). On the 10th day after TBI, KRT1 was significantly increased (p < 0.05) by 4.26-fold compared to the control group in the TBI group and significantly inhibited in the TBI plus RP group (p < 0.05). Similarly, the expression levels of other 8 proteins detected at 18th day after TBI were significantly increased by 4.29 to 27.44-fold in the TBI group, but significantly decreased in the TBI plus RP group compared to the TBI group, respectively.

Conclusion

Nine proteins were identified by proteomics methods from 269 analyzed proteins detected in mice exposed to a lethal dose of TBI. These proteins are also expected to be indicators of the damage induced by high-dose radiation.

Keywords:

• Thrombopoietin receptor agonist
• romiplostim
• radio-protective/mitigative effects
• proteomics

Acknowledgements

The authors are grateful to Miyu Miyazaki in the Scientific Research Facility Center of Hirosaki University Graduate School of Medicine for help with LC-MS/MS analysis.

Disclosure statement

The authors have no potential conflicts of interest to declare.

Additional information

Funding

This work was supported by a KAKENHI Grant-in-Aid for Scientific Research (A) [No. 16H02667 IK] and was partially supported by a KAKENHI Early-Career Scientists [No. 18K18190 MY].

Notes on contributors

Teruki Nishida

Teruki Nishida, M.S. in Health Sciences, is a Researcher of Radiation Biology at the Department of Radiation Science, Hirosaki University Graduate School of Health Sciences, Hirosaki, Aomori, Japan.

Masaru Yamaguchi

Masaru Yamaguchi, Ph.D. in Health Sciences, is an Assistant Professor of Radiation Biology and a Researcher at the Department of Radiation Science, Hirosaki University Graduate School of Health Sciences, Hirosaki, Aomori, Japan.

Yota Tatara

Yota Tatara, Ph.D. in Medical Science, is an Assistant Professor of Molecular Biology and a Senior Researcher at the Department of Glycotechnology, Center for Advanced Medical Research, Hirosaki University Graduate School of Medicine, Hirosaki, Aomori, Japan.

Ikuo Kashiwakura

Ikuo Kashiwakura, Ph.D. in Pharmacy, is a Professor of Radiation Biology and a Senior Researcher at the Department of Radiation Science, Hirosaki University Graduate School of Health Sciences, Hirosaki, Aomori, Japan.