Journal of Shandong University (Health Sciences) ›› 2021, Vol. 59 ›› Issue (4): 17-27.doi: 10.6040/j.issn.1671-7554.0.2021.0288

Previous Articles     Next Articles

Quantitative proteomic analysis of epidermal stem cells in oxygen-glucose deprivation conditions

ZHANG Huayu, YIN Siyuan, LIU Jian, MA Jiaxu, SONG Ru, CAO Guoqi, WANG Yibing   

  1. Department of Plastic Surgery, Shandong Provincial Qianfoshan Hospital, Shandong University, Jinan Clinical Research Center for Tissue Engineering Skin Regeneration and Wound Repair, Jinan 250014, Shandong, China
  • Published:2021-04-30

PDF (PC)

95

Abstract: Objective To find the key proteins and pathways leading to the dysfunction of epidermal stem cells in oxygen-glucose deprivation(OGD)conditions and to explore the formation mechanism of chronic hard healing wounds. Methods Epidermal stem cells were extracted from newborn mice and cultured normally to the P3 generation. The cells were cultured under OGD conditions for 0, 3, 6, 9 and 12 h, respectively, and then were divided into five goups: 0 h was taken as the control group, and the other groups were taken as the experimental groups. There are four comparison groups: OGD3_vs_OGD0, OGD6_vs_OGD0, OGD9_vs_OGD0 and OGD12_vs_OGD0. Quantitative proteomics determination was performed on different groups of epidermal stem cells using Tandem mass tag(TMT)technology. The four comparison groups were screened for proteins with significantly differential expression. These differentially expressed proteins(DEPs)were analyzed for Gene Ontology(GO)category, KEGG pathway, protein-protein interaction(PPI), gene overlap, and analysis of pathways and proteins related to mitophagy to find the key proteins and pathways in the dysfunction of epidermal stem cells. Results A total of 4 852 quantifiable proteins were detected in the experiment. In the four comparison groups, that is, OGD3_vs_OGD0, OGD6_vs_OGD0, OGD9_vs_OGD0, and OGD12_vs_OGD0, the number of up-regulated DEPs were 1, 225, 346 and 386, respectively, while the number of down-regulated DEPs were 26, 229, 330 and 462, respectively. In the analysis of the comparison group OGD6_vs_OGD0, key pathways included “ribonucleoprotein complex biogenesis(GO: 0022613)”“chromatin binding(GO: 0003682)”“cytoplasmic ribosomal large subunit(GO:0022625)” and “ribosome(ko03010)”,and the 21 hub proteins screened through PPI analysis were all structural proteins of the ribosome, of which the most important seed protein was the mitochondrial ribosomal protein MRPL24. In the comprehensive analysis of the four comparison groups, it was found that there was a large amount of overlap between the DEPs of each comparison group, and key pathways were all related to protein processing. In the analysis of mitophagy-related pathway and proteins, it was found that the expression of Tbc1d15, Rab7a, which were related to lysosomal maturation and p53, continued to be up-regulated, while the expression of LC3 related to the level of autophagy flow did not show a definite trend. Conclusion The key protein “MRPL24” obtained through TMT quantitative proteomics analysis and the pathways related to protein processing are the key to the dysfunction of epidermal stem cells in OGD conditions. They will be the breakthrough points for further research on the formation mechanism of chronic hard healing wounds.

Key words: Chronic hard healing wounds, Epidermal stem cells, Oxygen-glucose deprivation, Mitophagy, Proteomic analysis

CLC Number: 

  • R574
[1] 陈端凯,单云龙,唐乾利. 从细胞外微环境探讨MEBT/MEBO对慢性难愈合创面的修复作用[J]. 中国烧伤创疡杂志, 2019, 31(4): 236-239. CHEN Duankai, SHAN Yunlong, TANG Qianli. Study on the effect of MEBT/MEBO in repairing chronic refractory wounds from the perspective of extracellular microenvironment[J]. The Chinese Journal of Burns Wounds and Surface Ulcers, 2019, 31(4): 236-239.
[2] Eming SA, Martin P, Tomic-Canic M. Wound repair and regeneration: mechanisms, signaling, and translation[J]. Sci Transl Med, 2014, 6(265): 265sr6. doi: 10.1126/scitranslmed.3009337.
[3] Han G, Ceilley R. Chronic wound healing: a review of current management and treatments[J]. Adv Ther, 2017, 34(3): 599-610.
[4] Sorg H, Tilkorn DJ, Hager S, et al. Skin wound healing: an update on the current knowledge and concepts[J]. Eur Surg Res, 2017, 58(1-2): 81-94.
[5] 张建中,高兴华.皮肤性病学[M]. 北京:人民卫生出版社, 2015: 3-4.
[6] Lopez-Paniagua M, Nieto-Miguel T, de la Mata A, et al. Consecutive expansion of limbal epithelial stem cells from a single limbal biopsy[J]. Curr Eye Res, 2013, 38(5): 537-549.
[7] Pikula M, Kondej K, Jaskiewicz J, et al. Flow cytometric sorting and analysis of human epidermal stem cell candidates[J]. Cell Biol Int, 2010, 34(9): 911-915.
[8] 程飚,付小兵. 微环境控制是实现创面完美修复的必由之路[J].中华烧伤杂志, 2020, 36(11): 1003-1008. CHENG Biao, FU Xiaobing. Microenvironment control is the only way to achieve perfect wound repair[J]. Chinese Journal of Burns, 2020, 36(11): 1003-1008.
[9] Lemasters JJ. Selective mitochondrial autophagy, or mitophagy, as a targeted defense against oxidative stress, mitochondrial dysfunction, and aging[J]. Rejuvenation Res, 2005, 8(1): 3-5.
[10] Bhatia-Kissová I, Camougrand N. Mitophagy: a process that adapts to the cell physiology[J]. Int J Biochem Cell Biol, 2013, 45(1): 30-33.
[11] Huang C, Hong L, Xu J, et al. Protective roles of autophagy in retinal pigment epithelium under high glucose condition via regulating PINK1/Parkin pathway and BNIP3L[J]. Biol Res, 2018, 51(1):22. doi: 10.1186/s40659-018-0169-4.
[12] Arciuch AVG, Elguero ME, Poderoso JJ, et al. Mitochondrial regulation of cell cycle and proliferation[J]. Antioxid Redox Signal, 2012, 16(10): 1150-1180.
[13] Zhu W, Yuan Y, Liao G, et al. Mesenchymal stem cells ameliorate hyperglycemia-induced endothelial injury through modulation of mitophagy[J]. Cell Death Dis, 2018, 9(8): 837. doi: 10.1038/s41419-018-0861-x.
[14] Graeme CM, Edward LH, Wilhelm H, et al. Increasing the multiplexing capacity of TMTs using reporter ion isotopologues with isobaric masses[J]. Anal Chem, 2012, 84(17): 7469-7478.
[15] Mesa KR, Kawaguchi K, Cockburn K, et al. Homeostatic epidermal stem cell self-renewal is driven by local differentiation[J]. Cell Stem Cell, 2018, 23(5): 677-686.
[16] Toivola DM, Boor P, Alam C, et al. Keratins in health and disease[J]. Curr Opin Cell Biol, 2015, 32: 73-81. doi: 10.1016/j.ceb.2014.12.008.
[17] Ichiro K, Hitoshi M, Kenji K, et al. Cytokeratin, filaggrin, and p63 expression in reepithelialization during human cutaneous wound healing[J]. Wound Repair Regen, 2006, 14(1): 38-45.
[18] Jacek RW, Alexandre Z, Nagarjuna N, et al. Universal sample preparation method for proteome analysis[J]. Nat Methods, 2009, 6(5): 359-362.
[19] Novak I, Kirkin V, McEwan DG, et al. Nix is a selective autophagy receptor for mitochondrial clearance[J]. EMBO Rep, 2010, 11(1): 45-51.
[20] Weidberg H, Shvets E, Shpilka T, et al. LC3 and GATE-16/GABARAP subfamilies are both essential yet act differently in autophagosome biogenesis[J]. EMBO J, 2010, 29(11): 1792-1802.
[21] Rõtig A. Human diseases with impaired mitochondrial protein synthesis[J]. Biochim Biophys Acta, 2011, 1807(2011): 1198-1205.
[22] Nottia M, Marchese M, Verrigni D, et al. A homozygous MRPL24 mutation causes a complex movement disorder and affects the mitoribosome assembly[J]. Neurobiol Dis, 2020, 141:104880. doi: 10.1016/j.nbd.2020.104880.
[23] Azakir BA, Desrochers G, Angers A. The ubiquitin ligase Itch mediates the antiapoptotic activity of epidermal growth factor by promoting the ubiquitylation and degradation of the truncated C-terminal portion of Bid[J]. FEBS J, 2010, 277(5): 1319-1330.
[24] Ferreira AF, Cunha PS, Carregal VM, et al. Extracellular vesicles from adipose-derived mesenchymal stem/stromal cells accelerate migration and activate Akt pathway in human keratinocytes and fibroblasts independently of mir-205 activity[J]. Stem Cells Int, 2017, 2017: 9841035. doi: 10.1155/2017/9841035.
[25] Cheong A, Lingutla R, Mager J. Expression analysis of mammalian mitochondrial ribosomal protein genes[J]. Gene Expr Patterns, 2020, 38: 119147. doi: 10.1016/j.gep.2020.119147.
[26] Pennisi E. The race to the ribosome structure[J]. Science, 1999, 285(5436): 2048-2051.
[27] Schramm JC, Dinh T, Veves A. Microvascular changes in the diabetic foot[J]. Int J Low Extrem Wounds, 2006, 5(3): 149-159.
[1] XIONG Chao, LIU Li, FENG Jianguo, WEI Jicheng. Effects of sevoflurane preconditioning on silent information regulator of transcription 3 expression and acetylation in H9C2 cardiomyocytes after hypoxia/reoxygenation [J]. Journal of Shandong University (Health Sciences), 2019, 57(3): 25-30.
[2] ZHAO Xuelian, YU Jun, XIE Zhaohong, CAO Yanjun, LIU Zhen, WANG Xiao, XU Linlin, YANG Hui, ZHENG Xiaolei, SHEN Yang, BI Jianzhong. Mechanism of mitophagy in a cell model of Alzheimer's disease [J]. JOURNAL OF SHANDONG UNIVERSITY (HEALTH SCIENCES), 2015, 53(10): 1-5.
[3] HANG Zhong-xia1, ZHANG Hong2, CUI Dong-sheng1, WANG Tao3, NIU Jing-ya1, ZHANG Rui1, ZHOU Jie3, PEI Yunhai3, LI Jiang-jing3, XU Shun-jiang1. Neuroprotective effect of L-carnitine on ischemic brain injury in rats [J]. JOURNAL OF SHANDONG UNIVERSITY (HEALTH SCIENCES), 2012, 50(8): 14-19.
[4] ZHANG Hua1，3, LIU Xue-xia2, SONG Xi-cheng3， LEI Da-peng1, PAN Xin-liang1， ZHANG Qing-quan3. Preliminary proteomic analysis of human serum from patients with
 T1 glottic laryngeal carcinoma [J]. JOURNAL OF SHANDONG UNIVERSITY (HEALTH SCIENCES), 2012, 50(2): 110-115.
[5] CHEN Meng1, REN Ning2, LIU Wen-jun1, QIN Xiao-min1, LI Jin-song1. Screening of molecular markers for epidermal stem cells in mice by the double immunofluorescence labeling technique [J]. JOURNAL OF SHANDONG UNIVERSITY (HEALTH SCIENCES), 2011, 49(4): 65-69.
[6] ZHANG Wen-hua, XU Shu-jun, JIANG Yu-quan, LI Gang, LI Xin-gang, ZHOU Mao-de, WU Cheng-yuan. Protective effect of tricyclic antidepressant nortryptyline of cortical  neurons on rats with OGD-induced injury and its mechanism [J]. JOURNAL OF SHANDONG UNIVERSITY (HEALTH SCIENCES), 2011, 49(4): 29-32.
[7] QIN Wen,ZHANG Xiao-xia,LIU Ping,WANG Zi-ying. Protective effects of N-acetylcysteine on oxygen-glucose deprivation injured cortical astrocytes in rats [J]. JOURNAL OF SHANDONG UNIVERSITY (HEALTH SCIENCES), 2008, 46(9): 859-862.
Viewed
Full text


Abstract

Cited
  Shared   
  Discussed   
[1] ZHANG Jie,LI Zhen-hua,SUN Jin-hao,BAO Li-hua,LIU Yue-peng. Protective effects of the invariable magnetic field on oxidative injured Schwann cells[J]. JOURNAL OF SHANDONG UNIVERSITY (HEALTH SCIENCES), 2007, 45(3): 229 -232 .
[2] ZHENG Min,HAO Yue-wei,LIU Xue-ping,ZHAO Ting-ting. Relationship between the HPA-2 and Kozak sequence polymorphism of platelet membrane glycoprotein and cerebral infarction[J]. JOURNAL OF SHANDONG UNIVERSITY (HEALTH SCIENCES), 2008, 46(3): 292 -295 .
[3] . [J]. JOURNAL OF SHANDONG UNIVERSITY (HEALTH SCIENCES), 2007, 45(10): 995 .
[4] JIANG Hong-ju,LI Run-zhi, WANG Ying,XU Dong-mei,ZHANG Mei,ZHANG Yun,LI Ji-fu. Relationship between the plasma level of matrix metalloproteinase-9 and structural characteristics of plaques and percutaneous coronary intervention [J]. JOURNAL OF SHANDONG UNIVERSITY (HEALTH SCIENCES), 2008, 46(10): 966 -970 .
[5] . [J]. JOURNAL OF SHANDONG UNIVERSITY (HEALTH SCIENCES), 2008, 46(8): 828 -830 .
[6] . [J]. JOURNAL OF SHANDONG UNIVERSITY (HEALTH SCIENCES), 2009, 47(9): 5 -6 .
[7] NIU Rui,LIU Bo,SHAO Ming-ju,WANG Wei. mRNA expression of lunx in regional lymph nodes of non-small cell lung cancer and its correlation with prognosis[J]. JOURNAL OF SHANDONG UNIVERSITY (HEALTH SCIENCES), 2007, 45(9): 884 -885 .
[8] JIAO Fang-fang,LIU Shi-qing,LI Fei,LI Chang-sheng,WANG Qin,SUN Qing,LU Wei. Therapeutic effects of Huayulifei on expression of SMAD7 and TGF-β in rats with bleomycin-induced pulmonary fibrosis[J]. JOURNAL OF SHANDONG UNIVERSITY (HEALTH SCIENCES), 2007, 45(10): 1054 -1058 .
[9] ZHAO Ying， YAN Lei， ZHANG Hui， YU Peng， LI Ming-jiang， ZHAO Xing-bo. Expression of SPAG9 in patients with serous epithelial ovarian tumors[J]. JOURNAL OF SHANDONG UNIVERSITY (HEALTH SCIENCES), 2012, 50(2): 98 .
[10] WANG Shu-qin,QI Feng,WU Jian-bo,SUN Bao-zhu. Effect of calcium in ropivacaine-induced smooth muscle contraction of the aorta in rats in vitro[J]. JOURNAL OF SHANDONG UNIVERSITY (HEALTH SCIENCES), 2008, 46(8): 773 -776 .
Copyright 2018 © Editorial office of Journal of Shandong University (Health Sciences)
Supported by Beijing Magtech Co.Ltd