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. 2022 Sep;10(18):962.
doi: 10.21037/atm-22-3073.

Quantitative iTRAQ proteomics reveal the proteome profiles of bone marrow mesenchymal stem cells after cocultures with Schwann cells in vitro

Han Ding #  1 , Ang Li #  1   2 , Chao Sun #  1 , Jianping Zhang  1 , Jun Shang  1 , Haoshuai Tang  1 , Junjin Li  1 , Min Wang  3 , Xiaohong Kong  4 , Zhijian Wei  1 , Shiqing Feng  1
Affiliations

Quantitative iTRAQ proteomics reveal the proteome profiles of bone marrow mesenchymal stem cells after cocultures with Schwann cells in vitro

Han Ding et al. Ann Transl Med. 2022 Sep.

Abstract

Background: Bone marrow mesenchymal stem cells (BMSCs) combined with Schwann cells (SCs) represent a better therapeutic cell transplantation strategy for treating spinal cord injury (SCI) than transplantation with BMSCs or SCs alone. In previous studies, we demonstrated that BMSCs are able to differentiate in neuron-like cells when cocultured with SCs. The detailed mechanism underlying SCI repair that occurs during the combined transplantation of BMSCs and SCs has not yet been studied. In this study, we adopted an isobaric tag for relative and absolute quantitation (iTRAQ)-based protein identification/quantification approach to examine the effects of the SC and BMSC coculture process on the BMSCs and then obtained and analyzed the differentially expressed proteins (DEPs) and their possible related pathways.

Methods: This study included three groups based on the number of coculture days (i.e., 0, 3, and 7 days). Changes in BMSC protein expression levels were measured using the iTRAQ technique. A bioinformatics analysis of all the data was performed.

Results: In total, 6,760 types of proteins were detected, corresponding to 5,181 data points with quantitative information. Of these, a total of 243 DEPs were identified, of which 169 proteins were upregulated and 74 proteins were downregulated. These DEPs were identified by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses. Intercellular adhesion molecule-1 (ICAM-1), integrin, and dioxygenase may play crucial roles in the repair of SCI. The data analysis indicates that the relevant biological processes may be regulated by lysosome function, cell adhesion molecules (CAMs), leukocyte transendothelial migration, and the phosphatidylinositol-3-kinase (PI3K) and peroxisome proliferator-activated receptor (PPAR) signaling pathways.

Conclusions: The data provided in this study indicate that several molecular mechanisms and signaling pathways are involved in the BMSC and SC coculture process. This information may be useful for the further identification of specific targets and related mechanisms and guide new directions for SCI treatment.

Keywords: Proteomics analysis; Schwann cells (SCs); bone marrow mesenchymal stem cells (BMSCs); isobaric tag for relative and absolute quantitation (iTRAQ); spinal cord injury (SCI).

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Conflict of interest statement

Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at https://atm.amegroups.com/article/view/10.21037/atm-22-3073/coif). The authors have no conflicts of interest to declare.

Figures

Figure 1
Figure 1
Cell identification. (A) Immunofluorescence staining of S100 for SC identification. Magnification: 20×. (B) Flow cytometry for BMSC identification. (C) BMSCs have the ability to differentiate into adipocytes (oil red O), chondrocytes (Alcian blue) and osteoblasts (Alizarin red). Magnification: 20×. CD, cluster differentiation; SC, Schwann cell; BMSC, bone marrow mesenchymal stem cell.
Figure 2
Figure 2
A comparison of the number of the DEPs (up-regulated and down-regulated) in the three groups. SC3d group: the BMSCs were cocultured with SCs for 3 days; SC0d group: the BMSCs were cultured alone; SC7d group: the BMSCs were cocultured with SCs for 7 days. DEPs, differentially expressed proteins; BMSCs, bone marrow mesenchymal stem cells; SCs, Schwann cells.
Figure 3
Figure 3
GO enrichment analysis. (A) GO enrichment analysis of the DEPs in the SC3d group vs. the SC0d group. (B) GO enrichment analysis of the DEPs in the SC7d group vs. the SC0d group. (C) GO enrichment analysis of the DEPs in the SC7d group vs. the SC3d group. SC3d group: the BMSCs were cocultured with SCs for 3 days; SC0d group: the BMSCs were cultured alone; SC7d group: the BMSCs were cocultured with SCs for 7 days. UV, ultraviolet; GO, Gene Ontology; DEPs, differentially expressed proteins; BMSCs, bone marrow mesenchymal stem cells; SCs, Schwann cells.
Figure 4
Figure 4
KEGG pathway analysis. (A) KEGG pathway analysis of the DEPs in the SC3d vs. SC0d group. (B) KEGG pathway analysis of the DEPs in the SC7d vs. SC0d group. (C) KEGG pathway analysis of the DEPs in the SC7d vs. SC3d group. SC3d group: the BMSCs were cocultured with SCs for 3 days; SC0d group: the BMSCs were cultured alone; SC7d group: the BMSCs were cocultured with SCs for 7 days. ECM, extracellular matrix; HCM, hypertrophic cardiomyopathy; ARVC, arrhythmogenic right ventricular cardiomyopathy; PI3K, phosphatidylinositol-3-kinase; CAMs, cell adhesion molecules; ALS, amyotrophic lateral sclerosis; PPAR, peroxisome proliferator-activated receptor; KEGG, Kyoto Encyclopedia of Genes and Genomes; DEPs, differentially expressed proteins; BMSCs, bone marrow mesenchymal stem cells; SCs, Schwann cells.
Figure 5
Figure 5
PPI network analysis. (A) PPI network analysis of the DEPs in the SC3d vs. the SC0d group. (B) PPI network analysis of the DEPs in the SC7d vs. the SC0d group. (C) PPI network analysis of the DEPs in the SC7d vs. the SC3d group. SC3d group: the BMSCs were cocultured with SCs for 3 days; SC0d group: the BMSCs were cultured alone; SC7d group: the BMSCs were cocultured with SCs for 7 days. PPI, protein-protein interaction; DEPs, differentially expressed proteins; BMSCs, bone marrow mesenchymal stem cells; SCs, Schwann cells.
Figure 6
Figure 6
Western blot verification. (A) The expression level of the DEPs. (B) Quantification of the DEPs. *P<0.05, **P<0.01, compared to the control group. DEPs, differentially expressed proteins.
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