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The role of mmu-miR-155-5p-NF-?B signaling in the education of bone marrow-derived mesenchymal stem cells by gastric cancer cells.

ABSTRACT: Bone marrow-derived mesenchymal stem cells (BM-MSCs) are important precursors of tumor stromal cells. Previously, we have demonstrated that miR-155-5p inhibition directly induced transition of BM-MSCs into gastric cancer-associated MSCs. Whether miR-155-5p is involved in the education of BM-MSCs by gastric cancer cells has not been established. Murine BM-MSCs (mMSCs) were isolated and grown in conditioned medium derived from gastric cancer cell line MFC (MFC-CM). The tumor-promoting phenotype and function of mMSCs were detected by immunofluorescence staining, quantitative reverse transcription-polymerase chain reaction (qRT-PCR), cell colony formation assay, transwell migration, and invasion assays. Luciferase reporter assays and western blot analyses were conducted to reveal the relationship between nuclear factor kappa-light-chain-enhancer of activated B cells (NF-?B) p65 and mmu-miR-155-5p. miRNA mimics, inhibitor, and the NF-?B inhibitor pyrrolidine dithiocarbamic acid (PDTC) were used to evaluate the role of miR-155-5p-NF-?B signaling in the education of mMSCs by MFC-CM. We successfully established the education model of mMSCs by MFC-CM and found that mmu-miR-155-5p expression levels were reduced in mMSCs. Mimicking this deregulation by transfecting miRNA inhibitor into mMSCs produced a similar effect as that of MFC-CM on mMSCs. NF-?B p65 was validated as a target of mmu-miR-155-5p, which also negatively regulated NF-?B activation. Inhibition of NF-?B activation by PDTC abolished the effect of the miRNA inhibitor on mMSCs. mmu-miR-155-5p overexpression partially blocked the effect of MFC-CM in educating mMSCs, while PDTC treatment completely eliminated MFC-CM activity. These results indicate that miR-155-5p is not the sole miRNA mediating the education of BM-MSCs by gastric cancer cells, but downstream NF-?B signaling is indispensable for this process.

SUBMITTER: Wang M

PROVIDER: S-EPMC5852371 | biostudies-literature | 2018 Mar

REPOSITORIES: biostudies-literature

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The role of mmu-miR-155-5p-NF-κB signaling in the education of bone marrow-derived mesenchymal stem cells by gastric cancer cells.

Wang Mei M Yang Fang F Qiu Rong R Zhu Mengchu M Zhang Huiling H Xu Wenrong W Shen Bo B Zhu Wei W

Cancer medicine 20180214 3

Bone marrow-derived mesenchymal stem cells (BM-MSCs) are important precursors of tumor stromal cells. Previously, we have demonstrated that miR-155-5p inhibition directly induced transition of BM-MSCs into gastric cancer-associated MSCs. Whether miR-155-5p is involved in the education of BM-MSCs by gastric cancer cells has not been established. Murine BM-MSCs (mMSCs) were isolated and grown in conditioned medium derived from gastric cancer cell line MFC (MFC-CM). The tumor-promoting phenotype an ...[more]

PMID: 29441735

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Project description:BackgroundAcquired aplastic anemia (AA) is characterized by deficiency or dysfunction of the bone marrow (BM) microenvironment. However, little is known about the impairment of BM-derived mesenchymal stem cells (MSCs) in AA patients.MethodsWe used Illumina HiSeqTM 2000 sequencing, quantitative real-time polymerase chain reaction (qRT-PCR), flow cytometry (FCM), and Western blotting to test the expression of CD106 gene (vascular cell adhesion molecule 1 (VCAM1)) and CD106 protein of BM-MSCs. Furthermore, we used hematoxylin and eosin (H&E) and histochemical staining analysis, immunofluorescence, and the formation of capillary-like structures to analyze capillary tube-like formation in vitro; we also used the Matrigel plug assay to test in vivo vasculogenesis, and an assay of colony forming units (CFUs) and colony-forming unit-megakaryocyte (CFU-MK) to detect the support function of MSCs in vitro. The in vivo engraftment of CD34+ cells and MSCs in NOD/SCID mice was tested by FACS and survival assay; the expression of NF-κB was tested by NanoPro analysis and immunofluorescence. NF-κB-regulated CD106 gene (VCAM1) was confirmed by tumor necrosis factor alpha (TNF-α)-stimulated and lipopolysaccharide (LPS)-stimulated MSCs, blockade assay, and immunofluorescence.ResultsHere, we report that BM-MSCs from AA patients exhibited downregulation of the CD06 gene (VCAM1) and low expression of CD106 in vitro. Further analysis revealed that CD106+ MSCs from both AA patients and healthy controls had increased potential for in vitro capillary tube-like formation and in vivo vasculogenesis compared with CD106- MSCs, and the results were similar when healthy MSCs were compared with AA MSCs. CD106+ MSCs from both AA patients and healthy controls more strongly supported in vitro growth and in vivo engraftment of CD34+ cells in NOD/SCID mice than CD106- MSCs, and similar results were obtained when healthy MSCs and AA MSCs were compared. The expression of NF-κB was decreased in AA MSCs, and NF-κB regulated the CD106 gene (VCAM1) which supported hematopoiesis.ConclusionsThese results revealed the effect of CD106 and NF-κB in BM failure of AA.

Dataset Information

The role of mmu-miR-155-5p-NF-?B signaling in the education of bone marrow-derived mesenchymal stem cells by gastric cancer cells.

Publications

The role of mmu-miR-155-5p-NF-κB signaling in the education of bone marrow-derived mesenchymal stem cells by gastric cancer cells.

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