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Immunosuppressive mechanisms of human bone marrow derived mesenchymal stromal cells in BALB/c host graft versus host disease murine models.

Robles JD, Liu YP, Cao J, Xiang Z, Cai Y, Manio M, Tang EH, Chan GC - Exp Hematol Oncol (2015)

Bottom Line: Documentation of suppression of RANTES, CCL3, CXCL9, CCR5 and CXCR3 with simultaneous decrease of donor T cell alloreactivity was demonstrated 6 days after transplantation, along with reduction of levels of inflammatory cytokines, suppression of STAT 5A/B phosphorylation, increased expression of CCR7 and increased production of nitrous oxide by hMSCs.Documentation of homing of hMSCs to lymphoid organs and target tissues was also performed.These mechanisms contribute to the current understanding of MSC mechanisms of immunosuppression and forms a comprehensive picture of how they exert immunosuppression in an in vivo model of immune dysregulation.

View Article: PubMed Central - PubMed

Affiliation: Department of Paediatrics and Adolescent Medicine, The University of Hong Kong Li Ka Shing Faculty of Medicine, Queen Mary Hospital, 102 Pokfulam Rd., HKSAR, PRC.

ABSTRACT

Background: Mesenchymal stromal cells (MSCs) are proven to have immunosuppressive functions via various mechanisms. These mechanisms were demonstrated by administering bone marrow derived human MSCs (hMSCs) to graft versus host disease (GVHD) murine models.

Methods: BALB/c host mice were irradiated prior to receiving C57BL/6 donor T cell depleted bone marrow (TCDBM) cells (negative control) and donor CD4+ T lymphocyte with (treatment group) or without hMSCs (positive control). The presence of hMSCs in target tissues and lymphoid organs was documented by using in vivo imaging and measuring the expression of EphB2 and ephrin-B2 by RTqPCR. Survival rate and GVHD score were also monitored. Tissue sections were obtained for histopathologic analysis. Flow cytometry was used to document donor T cell alloreactivity and expression of CCR5, CXCR3 and CCR7. ELISA was utilized to determine levels of proinflammatory cytokines, RANTES (CCL5) and phosphorylated STAT 5A/B. RTqPCR was performed to quantify expression of CCL3 and CXCL9. Western blotting was performed to qualitatively measure iNOS expression.

Results: Survival rate and GVHD score improved with hMSC treatment. Pathologic changes of GVHD were abrogated. Documentation of suppression of RANTES, CCL3, CXCL9, CCR5 and CXCR3 with simultaneous decrease of donor T cell alloreactivity was demonstrated 6 days after transplantation, along with reduction of levels of inflammatory cytokines, suppression of STAT 5A/B phosphorylation, increased expression of CCR7 and increased production of nitrous oxide by hMSCs. Documentation of homing of hMSCs to lymphoid organs and target tissues was also performed.

Conclusions: These mechanisms contribute to the current understanding of MSC mechanisms of immunosuppression and forms a comprehensive picture of how they exert immunosuppression in an in vivo model of immune dysregulation.

No MeSH data available.


Related in: MedlinePlus

Measurement of levels of proinflammatory cytokines. (A) TNF-α. (B) IFN-γ. Lethally irradiated BALB/c host mice were given intravenous injections of 2 × 10^6 TCDBM cells from C57BL/6 donors with or without 0.25 × 10 ^6 CD4+ T cells from donors and 1 × 10^6 hMSCs. Tissue cytokine levels [TNF-α(A), IFN- (B)] of BALB/c hosts 6d after transplantation *p<0.05. Data were combined from 2 independent experiments with 4 mice in each experiment.
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Fig3: Measurement of levels of proinflammatory cytokines. (A) TNF-α. (B) IFN-γ. Lethally irradiated BALB/c host mice were given intravenous injections of 2 × 10^6 TCDBM cells from C57BL/6 donors with or without 0.25 × 10 ^6 CD4+ T cells from donors and 1 × 10^6 hMSCs. Tissue cytokine levels [TNF-α(A), IFN- (B)] of BALB/c hosts 6d after transplantation *p<0.05. Data were combined from 2 independent experiments with 4 mice in each experiment.

Mentions: Documentation of the host mice’s long term survival and GVHD score after being given multiple doses of hMSCs were performed (days 0, 3 and 6 after transplantation). Multiple doses are given to overcome the transient nature of the immunosuppressive effects of hMSCs and keep the mice alive for a longer period. All the negative control group host mice that received donor TCDBM cells survived for 80 days. The survival of positive control group host mice given TCDBM and CD4+ was approximately 30%, 80 days after transplantation. Maximal death rates were observed around days 7–14 after transplantation. The group of mice given hMSCs survived better than the positive control, with around 80% of mice still alive 80 days after transplantation. The differences in survival between the 3 groups are significant (p = 0.0035) by log-rank (Mantel-Cox Test) analysis of survival (Figure 2A). The clinical score also shows the effect hMSCs have in systemic symptoms of GVHD (p = 0.006) (Figure 2B). An important early indicator for acute GVHD is donor T cell expansion in the host mice. Treatment of mice with hMSCs caused a marked reduction in donor T cell alloreactivity in the target organs and lymphoid tissues (Figure 2C). In line with these findings, a significant decrease in levels of proinflammatory cytokines in host tissue, notably TNF-α in the spleen (p = 0.0391), and IFN-γ in the spleen and colon was noted (p = 0.030) (Figure 3A-B). Lastly, host spleen, liver, colon, and lungs were harvested 14 days after transplantation and were noted to have less prominent GVHD features with hMSC treatment (Figure 4).Figure 2


Immunosuppressive mechanisms of human bone marrow derived mesenchymal stromal cells in BALB/c host graft versus host disease murine models.

Robles JD, Liu YP, Cao J, Xiang Z, Cai Y, Manio M, Tang EH, Chan GC - Exp Hematol Oncol (2015)

Measurement of levels of proinflammatory cytokines. (A) TNF-α. (B) IFN-γ. Lethally irradiated BALB/c host mice were given intravenous injections of 2 × 10^6 TCDBM cells from C57BL/6 donors with or without 0.25 × 10 ^6 CD4+ T cells from donors and 1 × 10^6 hMSCs. Tissue cytokine levels [TNF-α(A), IFN- (B)] of BALB/c hosts 6d after transplantation *p<0.05. Data were combined from 2 independent experiments with 4 mice in each experiment.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4440561&req=5

Fig3: Measurement of levels of proinflammatory cytokines. (A) TNF-α. (B) IFN-γ. Lethally irradiated BALB/c host mice were given intravenous injections of 2 × 10^6 TCDBM cells from C57BL/6 donors with or without 0.25 × 10 ^6 CD4+ T cells from donors and 1 × 10^6 hMSCs. Tissue cytokine levels [TNF-α(A), IFN- (B)] of BALB/c hosts 6d after transplantation *p<0.05. Data were combined from 2 independent experiments with 4 mice in each experiment.
Mentions: Documentation of the host mice’s long term survival and GVHD score after being given multiple doses of hMSCs were performed (days 0, 3 and 6 after transplantation). Multiple doses are given to overcome the transient nature of the immunosuppressive effects of hMSCs and keep the mice alive for a longer period. All the negative control group host mice that received donor TCDBM cells survived for 80 days. The survival of positive control group host mice given TCDBM and CD4+ was approximately 30%, 80 days after transplantation. Maximal death rates were observed around days 7–14 after transplantation. The group of mice given hMSCs survived better than the positive control, with around 80% of mice still alive 80 days after transplantation. The differences in survival between the 3 groups are significant (p = 0.0035) by log-rank (Mantel-Cox Test) analysis of survival (Figure 2A). The clinical score also shows the effect hMSCs have in systemic symptoms of GVHD (p = 0.006) (Figure 2B). An important early indicator for acute GVHD is donor T cell expansion in the host mice. Treatment of mice with hMSCs caused a marked reduction in donor T cell alloreactivity in the target organs and lymphoid tissues (Figure 2C). In line with these findings, a significant decrease in levels of proinflammatory cytokines in host tissue, notably TNF-α in the spleen (p = 0.0391), and IFN-γ in the spleen and colon was noted (p = 0.030) (Figure 3A-B). Lastly, host spleen, liver, colon, and lungs were harvested 14 days after transplantation and were noted to have less prominent GVHD features with hMSC treatment (Figure 4).Figure 2

Bottom Line: Documentation of suppression of RANTES, CCL3, CXCL9, CCR5 and CXCR3 with simultaneous decrease of donor T cell alloreactivity was demonstrated 6 days after transplantation, along with reduction of levels of inflammatory cytokines, suppression of STAT 5A/B phosphorylation, increased expression of CCR7 and increased production of nitrous oxide by hMSCs.Documentation of homing of hMSCs to lymphoid organs and target tissues was also performed.These mechanisms contribute to the current understanding of MSC mechanisms of immunosuppression and forms a comprehensive picture of how they exert immunosuppression in an in vivo model of immune dysregulation.

View Article: PubMed Central - PubMed

Affiliation: Department of Paediatrics and Adolescent Medicine, The University of Hong Kong Li Ka Shing Faculty of Medicine, Queen Mary Hospital, 102 Pokfulam Rd., HKSAR, PRC.

ABSTRACT

Background: Mesenchymal stromal cells (MSCs) are proven to have immunosuppressive functions via various mechanisms. These mechanisms were demonstrated by administering bone marrow derived human MSCs (hMSCs) to graft versus host disease (GVHD) murine models.

Methods: BALB/c host mice were irradiated prior to receiving C57BL/6 donor T cell depleted bone marrow (TCDBM) cells (negative control) and donor CD4+ T lymphocyte with (treatment group) or without hMSCs (positive control). The presence of hMSCs in target tissues and lymphoid organs was documented by using in vivo imaging and measuring the expression of EphB2 and ephrin-B2 by RTqPCR. Survival rate and GVHD score were also monitored. Tissue sections were obtained for histopathologic analysis. Flow cytometry was used to document donor T cell alloreactivity and expression of CCR5, CXCR3 and CCR7. ELISA was utilized to determine levels of proinflammatory cytokines, RANTES (CCL5) and phosphorylated STAT 5A/B. RTqPCR was performed to quantify expression of CCL3 and CXCL9. Western blotting was performed to qualitatively measure iNOS expression.

Results: Survival rate and GVHD score improved with hMSC treatment. Pathologic changes of GVHD were abrogated. Documentation of suppression of RANTES, CCL3, CXCL9, CCR5 and CXCR3 with simultaneous decrease of donor T cell alloreactivity was demonstrated 6 days after transplantation, along with reduction of levels of inflammatory cytokines, suppression of STAT 5A/B phosphorylation, increased expression of CCR7 and increased production of nitrous oxide by hMSCs. Documentation of homing of hMSCs to lymphoid organs and target tissues was also performed.

Conclusions: These mechanisms contribute to the current understanding of MSC mechanisms of immunosuppression and forms a comprehensive picture of how they exert immunosuppression in an in vivo model of immune dysregulation.

No MeSH data available.


Related in: MedlinePlus