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Molecular Mechanisms of Immunomodulation Properties of Mesenchymal Stromal Cells: A New Insight into the Role of ICAM-1 - PubMed

Molecular Mechanisms of Immunomodulation Properties of Mesenchymal Stromal Cells: A New Insight into the Role of ICAM-1

Yury Rubtsov et al. Stem Cells Int. 2017.

Abstract

Mesenchymal stromal cells (MSC) control excessive inflammation and create a microenvironment for tissue repair protecting from chronic inflammation and tissue fibrosis. We examined the molecular mechanisms of MSC immunomodulatory function in mixed cultures of human adipose-derived MSC with lymphocytes. Our data show that MSC promote unstimulated lymphocyte survival potentially by an increase in antigen presentation. Under inflammatory conditions, mimicked by stimulation of TCR in lymphocytes, MSC suppress activation and proliferation of stimulated T cells. Immunosuppression is accompanied by downregulation of IL-2Rα that negatively affects the survival of activated T cells. MSC upregulate transcription of indolamine-2,3-dioxygenase (IDO) and inducible NO synthase (iNOS), which generate products negatively affecting T cell function. Both MSC and lymphocytes dramatically increase the surface ICAM-1 level in mixed cultures. Antibody-mediated blockage of surface ICAM-1 partially releases MSC-mediated immune suppression in vitro. Our data suggest that MSC have cell-intrinsic molecular programs depending on the inflammatory microenvironment. We speculate that MSC sense soluble factors and respond by surface ICAM-1 upregulation. ICAM-1 is involved in the control of T cell activation leading to immunosuppression or modest stimulation depending on the T cell status. Immunomodulation by MSC ranging from support of naive T cell survival to immunosuppression of activated T cells may affect the tissue microenvironment protecting from aberrant regeneration.

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Figures

Figure 1
Figure 1

hASC suppress lymphocyte proliferation and downregulate IL-2Rα on activated T cells. Proliferation of PBMC activated with PHA and cultured separately and with hASC in contact or contactless conditions (25 : 1 ratio, 48 h) (a). CD25 expression in PBMC stimulated with plate-bound anti-CD3 and anti-CD28 and cultured with hASC (25 : 1, 48 h, mean + SEM, n = 5). Culture medium and medium conditioned by hASC served as negative controls. Proportion and absolute number of CD25+ CD4 T cells determined using immunofluorescent staining and FACS analysis are shown (mean + SEM, n = 6) (b, c). Similar analysis was performed using FACS sorted from PBMC pure (>95%) human CD4 T cells incubated with hASC (right plot); results obtained using total PBMC stained for CD25 are shown on the left plot; data of representative experiments are shown as histograms (d). Representative images of live PBMC-hASC cultures acquired using light microscope equipped with a digital camera. (48 h of incubation, magnification—10х) (e, f) (∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001).

Figure 2
Figure 2

Increase in IDO level in hASC in the presence of activated PBMC. Supernatants from mixed cultures of activated PBMC (anti-CD3, anti-CD28) cultured with hASC (48 h) affect the CD25 level in CD4 T cells. After 48 hours of incubation, PBMC were analyzed for CD25 (IL-2Rα) surface expression (n = 3, mean + SEM) (a). Changes in IDO transcription after treatment of hASC with supernantants (n = 3) (b). Transcriptional activity of IDO (c) and iNOS (d) genes in the course of hASC-mediated immune suppression (n = 4). IDO protein level in hASC cultured with anti-CD3- and anti-CD28-stimulated PBMC at 1 : 25 ratio for 48 hours (vinculin-normalization control) (e) (∗p < 0.05 and ∗∗p < 0.01; unstim—unstimulated PBMC, stim—PBMC stimulated with anti-CD3 and anti-CD28).

Figure 3
Figure 3

IDO inhibition blocks paracrine hASC-mediated immune suppression. Activated PBMC were cultured with or without hASC in contact and contactless conditions in the presence of 1-MT for 48 hours. Proportion of CD25+ CD4 T cells (a); IDO mRNA changes in hASC (b); IDO protein level in hASC cell lysates (c); kynurenine level in supernatants from hASC-PBMC cultures. Absorbance values (405 nm) were normalized to control samples (supernatant from hASC cultured alone). (d) All data represent mean + SEM of three independent experiments (1-MT—1-methyl-DL-tryptophan; ∗p < 0.05 and ∗∗p < 0.01).

Figure 4
Figure 4

hASC support survival of unstimulated T cells. CD25 expression in unstimulated PBMC cultured with hASC in contact and contactless conditions (48 h). Proportion (a) and absolute number (b) of CD25+ CD4 T cells (n = 6). Representative dot plot of CD25 expression on unstimulated T cells cultured with hASC (c). HLA-DR expression in nonstimulated PBMC cultured alone or with hASC under different conditions for 7 days. hASC (d) and PBMC (e), respectively (∗p < 0.05 and ∗∗p < 0.01).

Figure 5
Figure 5

ICAM-1 level changes in the course of hASC-mediated immunosuppression. ICAM-1 mRNA levels in hASC (a) and PBMC (b) from mixed cultures. ICAM-1 surface expression on hASC (c, d) and stimulated (e, f) or unstimulated (g, h) PBMC from mixed cultures (all data represent mean + SEM, n = 6, ∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001).

Figure 6
Figure 6

Supernatants from mixed cultures of hASC and activated PBMC stimulate ICAM-1 transcription in hASC and PBMC. PBMCs were activated with anti-CD3 and СD28 and cultured alone or with hASC in contact and contactless conditions (48 h). Supernatants were added to activated PBMC and hASC cultured alone. PBMC (a) and hASC (b) were analyzed for ICAM-1 mRNA level 48 hours later (mean + SEM, n = 3, ∗p < 0.05).

Figure 7
Figure 7

ICAM-1 blocking inhibits hASC-mediated immunosuppression independently of IDO. CD3+ CD4 T cells sorted from PBMC and activated with anti-CD3/anti-СD28 were cultured alone or with hASC in the presence of anti-ICAM-1 or control IgG1 in contact or contactless conditions for 48 hours. Proportion and absolute numbers of CD25+ CD4 T cells were determined (a, b, resp.). hASC IDO gene expression (c); IDO protein level (d). Kynurenine level in mixed cultures (e) (mean + SEM, n = 6, ∗p < 0.05 and ∗∗p < 0.01). T cell-hASC cultures after control IgG1 (e) or anti-ICAM (f) treatment. Magnification 10x.

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References

    1. Friedenstein A. J., Chailakhjan R. K., Lalykina K. S. The development of fibroblast colonies in monolayer cultures of guinea-pig bone marrow and spleen cells. Cell Tissue Kinetics. 1970;20(3):393–403. - PubMed
    1. Dominici M., Le Blanc K., Mueller I., et al. Minimal criteria for defining multipotent mesenchymal stromal cells. The International Society for Cellular Therapy position statement. Cytotherapy. 2006;8(4):315–317. doi: 10.1080/14653240600855905. - DOI - PubMed
    1. Prockop D. J. Marrow stromal cells as stem cells for nonhematopoietic tissues. Science. 1997;276(5309):71–74. doi: 10.1126/science.276.5309.71. - DOI - PubMed
    1. Salem H. K., Thiemermann C. Mesenchymal stromal cells: current understanding and clinical status. Stem Cells. 2010;28(3):585–596. doi: 10.1002/stem.269. - DOI - PMC - PubMed
    1. Javazon E. H., Beggs K. J., Flake A. W. Mesenchymal stem cells: paradoxes of passaging. Experimental Hematology. 2004;32(5):414–425. doi: 10.1016/j.exphem.2004.02.004. - DOI - PubMed