Limits...
Mechanisms of the immunosuppressive effects of mouse adipose tissue-derived mesenchymal stromal cells on mouse alloreactively stimulated spleen cells.

Nagaya R, Mizuno-Kamiya M, Takayama E, Kawaki H, Onoe I, Tanabe T, Nagahara K, Kondoh N - Exp Ther Med (2013)

Bottom Line: The suppressive effect of ASCs mediated by direct cell contact was partially reversed following knockdown of β2 microglobulin, a component of the major histocompatibility complex (MHC) class I molecule, with siRNA.The results of the study demonstrated that ASCs have significant immune modulatory effects on alloreactively stimulated spleen cells.The effects of ASCs on spleen cells are dependent on soluble factor(s) and cell contact, which is mediated by the MHC class I complex on ASCs.

View Article: PubMed Central - PubMed

Affiliation: Department of Oral Implantology, Asahi University School of Dentistry, Mizuho-shi, Gifu 501-0296, Japan.

ABSTRACT
The mechanisms of immunomodulation by mesenchymal stromal cells remain poorly understood. In this study, the effects of mouse adipose tissue-derived mesenchymal stromal cells (ASCs) on mouse spleen cells alloreactively stimulated by anti-CD3 and anti-CD28 antibody-coated (anti-CD3/CD28) beads were observed. Production of interferon-γ by the anti-CD3/CD28 bead-stimulated spleen cells was significantly suppressed in co-culture with ASCs. However, an augmented intensity of CD69 on the stimulated spleen cells was not suppressed in the presence of ASCs. The immunosuppressive effects of ASCs were partially mediated by one or more soluble factors (26% suppression). However, the ASCs require cell-cell contact in order to maximally exert suppression (88%). The suppressive effect of ASCs mediated by direct cell contact was partially reversed following knockdown of β2 microglobulin, a component of the major histocompatibility complex (MHC) class I molecule, with siRNA. The results of the study demonstrated that ASCs have significant immune modulatory effects on alloreactively stimulated spleen cells. The effects of ASCs on spleen cells are dependent on soluble factor(s) and cell contact, which is mediated by the MHC class I complex on ASCs.

No MeSH data available.


Related in: MedlinePlus

(A) Suppression of endogenous β2M transcripts in ASCs transfected with β2M-specific siRNA. The expression of endogenous β2M mRNA in siRNA-transfected ASCs was confirmed by semi-quantitative PCR. The relative expression level of β2M mRNA was normalized as a ratio to that of RPS5-mRNA. Results are described as a mean of duplicate experiments. (B) IFN-γ production by stimulated spleen cells in the presence or absence of siRNA-transfected ASCs. Anti-CD3/CD28-stimulated spleen cells were directly (a), or indirectly (b) co-cultured with β2M-specific or control (scrambled) siRNA-transfected ASCs. Relative IFN-γ levels in the supernatant are indicated by the ordinate. Experiments were repeated twice with good reproducibility. Results from a representative experiment are summarized. Values in more than triplicates were described as the mean ± standard deviation. The Student’s t-test was used to test the probability of significant differences between samples. #P<0.05 β2M siRNA vs. scrambled siRNA; **P<0.01 β2M siRNA or scrambled siRNA vs. control (without ASCs). β2M, β2 microglobulin; ASCs, adipose tissue-derived mesenchymal stromal cells; RPS5, ribosomal protein S5; IFN-γ, interferon-γ.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC3860983&req=5

f4-etm-07-01-0017: (A) Suppression of endogenous β2M transcripts in ASCs transfected with β2M-specific siRNA. The expression of endogenous β2M mRNA in siRNA-transfected ASCs was confirmed by semi-quantitative PCR. The relative expression level of β2M mRNA was normalized as a ratio to that of RPS5-mRNA. Results are described as a mean of duplicate experiments. (B) IFN-γ production by stimulated spleen cells in the presence or absence of siRNA-transfected ASCs. Anti-CD3/CD28-stimulated spleen cells were directly (a), or indirectly (b) co-cultured with β2M-specific or control (scrambled) siRNA-transfected ASCs. Relative IFN-γ levels in the supernatant are indicated by the ordinate. Experiments were repeated twice with good reproducibility. Results from a representative experiment are summarized. Values in more than triplicates were described as the mean ± standard deviation. The Student’s t-test was used to test the probability of significant differences between samples. #P<0.05 β2M siRNA vs. scrambled siRNA; **P<0.01 β2M siRNA or scrambled siRNA vs. control (without ASCs). β2M, β2 microglobulin; ASCs, adipose tissue-derived mesenchymal stromal cells; RPS5, ribosomal protein S5; IFN-γ, interferon-γ.

Mentions: It has been identified that the immunosuppressive effects of MSCs involve a nonclassical MHC, HLA-G (28,29). HLA-G is expressed in membrane-bound and soluble isoforms (29). In order to examine the association of MHC molecules and the suppressive effects of mouse ASCs, knockdown of endogenous β2M in ASCs was performed, as the molecule is a component of MHC class I molecules, and also necessary for cell surface expression of MHC class I molecules and stability (30). As shown in Fig. 4A, β2M siRNA-transfected ASCs expressed a lower level of endogenous β2M mRNA (<20%) compared with that of the non-specific scrambled RNA-transfected cells.


Mechanisms of the immunosuppressive effects of mouse adipose tissue-derived mesenchymal stromal cells on mouse alloreactively stimulated spleen cells.

Nagaya R, Mizuno-Kamiya M, Takayama E, Kawaki H, Onoe I, Tanabe T, Nagahara K, Kondoh N - Exp Ther Med (2013)

(A) Suppression of endogenous β2M transcripts in ASCs transfected with β2M-specific siRNA. The expression of endogenous β2M mRNA in siRNA-transfected ASCs was confirmed by semi-quantitative PCR. The relative expression level of β2M mRNA was normalized as a ratio to that of RPS5-mRNA. Results are described as a mean of duplicate experiments. (B) IFN-γ production by stimulated spleen cells in the presence or absence of siRNA-transfected ASCs. Anti-CD3/CD28-stimulated spleen cells were directly (a), or indirectly (b) co-cultured with β2M-specific or control (scrambled) siRNA-transfected ASCs. Relative IFN-γ levels in the supernatant are indicated by the ordinate. Experiments were repeated twice with good reproducibility. Results from a representative experiment are summarized. Values in more than triplicates were described as the mean ± standard deviation. The Student’s t-test was used to test the probability of significant differences between samples. #P<0.05 β2M siRNA vs. scrambled siRNA; **P<0.01 β2M siRNA or scrambled siRNA vs. control (without ASCs). β2M, β2 microglobulin; ASCs, adipose tissue-derived mesenchymal stromal cells; RPS5, ribosomal protein S5; IFN-γ, interferon-γ.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3860983&req=5

f4-etm-07-01-0017: (A) Suppression of endogenous β2M transcripts in ASCs transfected with β2M-specific siRNA. The expression of endogenous β2M mRNA in siRNA-transfected ASCs was confirmed by semi-quantitative PCR. The relative expression level of β2M mRNA was normalized as a ratio to that of RPS5-mRNA. Results are described as a mean of duplicate experiments. (B) IFN-γ production by stimulated spleen cells in the presence or absence of siRNA-transfected ASCs. Anti-CD3/CD28-stimulated spleen cells were directly (a), or indirectly (b) co-cultured with β2M-specific or control (scrambled) siRNA-transfected ASCs. Relative IFN-γ levels in the supernatant are indicated by the ordinate. Experiments were repeated twice with good reproducibility. Results from a representative experiment are summarized. Values in more than triplicates were described as the mean ± standard deviation. The Student’s t-test was used to test the probability of significant differences between samples. #P<0.05 β2M siRNA vs. scrambled siRNA; **P<0.01 β2M siRNA or scrambled siRNA vs. control (without ASCs). β2M, β2 microglobulin; ASCs, adipose tissue-derived mesenchymal stromal cells; RPS5, ribosomal protein S5; IFN-γ, interferon-γ.
Mentions: It has been identified that the immunosuppressive effects of MSCs involve a nonclassical MHC, HLA-G (28,29). HLA-G is expressed in membrane-bound and soluble isoforms (29). In order to examine the association of MHC molecules and the suppressive effects of mouse ASCs, knockdown of endogenous β2M in ASCs was performed, as the molecule is a component of MHC class I molecules, and also necessary for cell surface expression of MHC class I molecules and stability (30). As shown in Fig. 4A, β2M siRNA-transfected ASCs expressed a lower level of endogenous β2M mRNA (<20%) compared with that of the non-specific scrambled RNA-transfected cells.

Bottom Line: The suppressive effect of ASCs mediated by direct cell contact was partially reversed following knockdown of β2 microglobulin, a component of the major histocompatibility complex (MHC) class I molecule, with siRNA.The results of the study demonstrated that ASCs have significant immune modulatory effects on alloreactively stimulated spleen cells.The effects of ASCs on spleen cells are dependent on soluble factor(s) and cell contact, which is mediated by the MHC class I complex on ASCs.

View Article: PubMed Central - PubMed

Affiliation: Department of Oral Implantology, Asahi University School of Dentistry, Mizuho-shi, Gifu 501-0296, Japan.

ABSTRACT
The mechanisms of immunomodulation by mesenchymal stromal cells remain poorly understood. In this study, the effects of mouse adipose tissue-derived mesenchymal stromal cells (ASCs) on mouse spleen cells alloreactively stimulated by anti-CD3 and anti-CD28 antibody-coated (anti-CD3/CD28) beads were observed. Production of interferon-γ by the anti-CD3/CD28 bead-stimulated spleen cells was significantly suppressed in co-culture with ASCs. However, an augmented intensity of CD69 on the stimulated spleen cells was not suppressed in the presence of ASCs. The immunosuppressive effects of ASCs were partially mediated by one or more soluble factors (26% suppression). However, the ASCs require cell-cell contact in order to maximally exert suppression (88%). The suppressive effect of ASCs mediated by direct cell contact was partially reversed following knockdown of β2 microglobulin, a component of the major histocompatibility complex (MHC) class I molecule, with siRNA. The results of the study demonstrated that ASCs have significant immune modulatory effects on alloreactively stimulated spleen cells. The effects of ASCs on spleen cells are dependent on soluble factor(s) and cell contact, which is mediated by the MHC class I complex on ASCs.

No MeSH data available.


Related in: MedlinePlus