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DNA methyltransferase inhibition accelerates the immunomodulation and migration of human mesenchymal stem cells.

Lee S, Kim HS, Roh KH, Lee BC, Shin TH, Yoo JM, Kim YL, Yu KR, Kang KS, Seo KW - Sci Rep (2015)

Bottom Line: However, the effect of DNMT inhibitors on the immunomodulatory capacity of human mesenchymal stem cells (hMSCs) has not been investigated.We also determined that the migration of hMSCs toward ligands for CXCR2/CXCR4 was increased after 5-aza treatment.Moreover, using an experimental colitis model, we showed that 5-aza pre-treatment could enhance the therapeutic effect of MSCs against immune-related diseases.

View Article: PubMed Central - PubMed

Affiliation: 1] Research Institute for Veterinary Medicine, College of Veterinary Medicine, Seoul National University, Seoul 151-742, South Korea [2] Institute for Stem Cell and Regenerative Medicine in KangstemBiotech, Biotechnology Incubating Center, Seoul National University, Seoul 151-742, South Korea.

ABSTRACT
DNA methyltransferase (DNMT) inhibitors regulate target gene expression through epigenetic modifications, and these compounds have primarily been studied for cancer therapy or reprogramming. However, the effect of DNMT inhibitors on the immunomodulatory capacity of human mesenchymal stem cells (hMSCs) has not been investigated. In the present study, we treated hMSCs with 5-azacytidine (5-aza), a DNMT inhibitor, and confirmed that the inhibitory effects on mononuclear cell proliferation and cell migration toward activated T cells were increased. To identify the immunomodulatory factors stimulated through 5-aza treatment, we investigated the changes in promoter methylation patterns using methylation arrays and observed that the promoters of immunomodulatory factors, COX2 and PTGES, and migration-related factors, CXCR2 and CXCR4, were hypomethylated after 5-aza treatment. In addition, we observed that the COX2-PGE2 pathway is one of the main pathways for the enhanced immunosuppressive activity of hMSCs through 5-aza treatment. We also determined that the migration of hMSCs toward ligands for CXCR2/CXCR4 was increased after 5-aza treatment. Moreover, using an experimental colitis model, we showed that 5-aza pre-treatment could enhance the therapeutic effect of MSCs against immune-related diseases.

No MeSH data available.


Related in: MedlinePlus

5-aza stimulates the migration ability of hMSCs through the increase of CXCR2 and CXCR4.(A-B) After treating hMSCs with 5-aza, CXCR2 and CXCR4 expression was confirmed through (A) real-time qPCR and (B) western blot analysis. (C) After 5-aza treatment, the migration of hMSCs was determined after treatment with the CXCR2 or CXCR4 ligands. (D) Schematic diagrams indicating the locations of the promoter primers. (E) After treating hMSCs with 5-aza, bisulfide conversion and methyl-specific PCR were performed. *, p < 0.05; **, p < 0.01; ***, p < 0.001. Results show one representative experiment out of at least three. Results are shown as mean ± SD.
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f5: 5-aza stimulates the migration ability of hMSCs through the increase of CXCR2 and CXCR4.(A-B) After treating hMSCs with 5-aza, CXCR2 and CXCR4 expression was confirmed through (A) real-time qPCR and (B) western blot analysis. (C) After 5-aza treatment, the migration of hMSCs was determined after treatment with the CXCR2 or CXCR4 ligands. (D) Schematic diagrams indicating the locations of the promoter primers. (E) After treating hMSCs with 5-aza, bisulfide conversion and methyl-specific PCR were performed. *, p < 0.05; **, p < 0.01; ***, p < 0.001. Results show one representative experiment out of at least three. Results are shown as mean ± SD.

Mentions: MSCs express CXCR2 and CXCR4 and show chemotactic activity towards their ligands, SDF-1/CXCL12 and IL-8/CXCL832. Moreover, the SDF-1/CXCR4 axis is important for the migration of MSCs towards target tissues with inflammatory reactivity1433. We investigated whether the increased CXCR2 and CXCR4 expression after 5-aza treatment affects the hMSC migration. After the treatment of 5-aza on hMSCs for 24 hr, the expression of CXCR2 and CXCR4 was increased on mRNA and protein levels (Fig. 5A, B). Next, we confirmed whether the migration of hMSCs toward CXCR2 and CXCR4 ligands is altered through 5-aza treatment. The number of migrated cells toward IL-8, CXCL1 and SDF-1 was significantly increased after 5-aza treatment (P = 0.0028, 0.0001, 0.0023, respectively, Fig. 5C). To determine whether the increase in CXCR2 and CXCR4 expression through 5-aza is associated with demethylation of the gene promoter, changes in the methylation pattern following 5-aza treatment were analyzed using methyl-specific PCR (Fig. 5D). The promoters of both CXCR2 and CXCR4 were hypomethylated after 5-aza treatment (Fig. 5E).


DNA methyltransferase inhibition accelerates the immunomodulation and migration of human mesenchymal stem cells.

Lee S, Kim HS, Roh KH, Lee BC, Shin TH, Yoo JM, Kim YL, Yu KR, Kang KS, Seo KW - Sci Rep (2015)

5-aza stimulates the migration ability of hMSCs through the increase of CXCR2 and CXCR4.(A-B) After treating hMSCs with 5-aza, CXCR2 and CXCR4 expression was confirmed through (A) real-time qPCR and (B) western blot analysis. (C) After 5-aza treatment, the migration of hMSCs was determined after treatment with the CXCR2 or CXCR4 ligands. (D) Schematic diagrams indicating the locations of the promoter primers. (E) After treating hMSCs with 5-aza, bisulfide conversion and methyl-specific PCR were performed. *, p < 0.05; **, p < 0.01; ***, p < 0.001. Results show one representative experiment out of at least three. Results are shown as mean ± SD.
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4306122&req=5

f5: 5-aza stimulates the migration ability of hMSCs through the increase of CXCR2 and CXCR4.(A-B) After treating hMSCs with 5-aza, CXCR2 and CXCR4 expression was confirmed through (A) real-time qPCR and (B) western blot analysis. (C) After 5-aza treatment, the migration of hMSCs was determined after treatment with the CXCR2 or CXCR4 ligands. (D) Schematic diagrams indicating the locations of the promoter primers. (E) After treating hMSCs with 5-aza, bisulfide conversion and methyl-specific PCR were performed. *, p < 0.05; **, p < 0.01; ***, p < 0.001. Results show one representative experiment out of at least three. Results are shown as mean ± SD.
Mentions: MSCs express CXCR2 and CXCR4 and show chemotactic activity towards their ligands, SDF-1/CXCL12 and IL-8/CXCL832. Moreover, the SDF-1/CXCR4 axis is important for the migration of MSCs towards target tissues with inflammatory reactivity1433. We investigated whether the increased CXCR2 and CXCR4 expression after 5-aza treatment affects the hMSC migration. After the treatment of 5-aza on hMSCs for 24 hr, the expression of CXCR2 and CXCR4 was increased on mRNA and protein levels (Fig. 5A, B). Next, we confirmed whether the migration of hMSCs toward CXCR2 and CXCR4 ligands is altered through 5-aza treatment. The number of migrated cells toward IL-8, CXCL1 and SDF-1 was significantly increased after 5-aza treatment (P = 0.0028, 0.0001, 0.0023, respectively, Fig. 5C). To determine whether the increase in CXCR2 and CXCR4 expression through 5-aza is associated with demethylation of the gene promoter, changes in the methylation pattern following 5-aza treatment were analyzed using methyl-specific PCR (Fig. 5D). The promoters of both CXCR2 and CXCR4 were hypomethylated after 5-aza treatment (Fig. 5E).

Bottom Line: However, the effect of DNMT inhibitors on the immunomodulatory capacity of human mesenchymal stem cells (hMSCs) has not been investigated.We also determined that the migration of hMSCs toward ligands for CXCR2/CXCR4 was increased after 5-aza treatment.Moreover, using an experimental colitis model, we showed that 5-aza pre-treatment could enhance the therapeutic effect of MSCs against immune-related diseases.

View Article: PubMed Central - PubMed

Affiliation: 1] Research Institute for Veterinary Medicine, College of Veterinary Medicine, Seoul National University, Seoul 151-742, South Korea [2] Institute for Stem Cell and Regenerative Medicine in KangstemBiotech, Biotechnology Incubating Center, Seoul National University, Seoul 151-742, South Korea.

ABSTRACT
DNA methyltransferase (DNMT) inhibitors regulate target gene expression through epigenetic modifications, and these compounds have primarily been studied for cancer therapy or reprogramming. However, the effect of DNMT inhibitors on the immunomodulatory capacity of human mesenchymal stem cells (hMSCs) has not been investigated. In the present study, we treated hMSCs with 5-azacytidine (5-aza), a DNMT inhibitor, and confirmed that the inhibitory effects on mononuclear cell proliferation and cell migration toward activated T cells were increased. To identify the immunomodulatory factors stimulated through 5-aza treatment, we investigated the changes in promoter methylation patterns using methylation arrays and observed that the promoters of immunomodulatory factors, COX2 and PTGES, and migration-related factors, CXCR2 and CXCR4, were hypomethylated after 5-aza treatment. In addition, we observed that the COX2-PGE2 pathway is one of the main pathways for the enhanced immunosuppressive activity of hMSCs through 5-aza treatment. We also determined that the migration of hMSCs toward ligands for CXCR2/CXCR4 was increased after 5-aza treatment. Moreover, using an experimental colitis model, we showed that 5-aza pre-treatment could enhance the therapeutic effect of MSCs against immune-related diseases.

No MeSH data available.


Related in: MedlinePlus