Limits...
The Macrophage Galactose-Type C-Type Lectin (MGL) Modulates Regulatory T Cell Functions.

Zizzari IG, Martufi P, Battisti F, Rahimi H, Caponnetto S, Bellati F, Nuti M, Rughetti A, Napoletano C - PLoS ONE (2015)

Bottom Line: In this study, we demonstrate that Treg immunosuppressive activity can be modulated by the cross-linking between the CD45RA expressed by Tregs and the C-type lectin MGL.This effect can be attributed to changes in CD45RA and TCR signalling through the inhibition of Lck and inactivation of Zap-70, an increase in the Foxp3 methylation status and, ultimately, the reduced production of suppressive cytokines.These results indicate a role of MGL as an immunomodulator within the tumour microenvironment interfering with Treg functions, suggesting its possible use in the design of anticancer vaccines.

View Article: PubMed Central - PubMed

Affiliation: Department of Experimental Medicine, "Sapienza" University, Rome, Italy.

ABSTRACT
Regulatory T cells (Tregs) are physiologically designed to prevent autoimmune disease and maintain self-tolerance. In tumour microenvironments, their presence is related to a poor prognosis, and they influence the therapeutic outcome due to their capacity to suppress the immune response by cell-cell contact and to release immunosuppressive cytokines. In this study, we demonstrate that Treg immunosuppressive activity can be modulated by the cross-linking between the CD45RA expressed by Tregs and the C-type lectin MGL. This specific interaction strongly decreases the immunosuppressive activity of Tregs, restoring the proliferative capacity of co-cultured T lymphocytes. This effect can be attributed to changes in CD45RA and TCR signalling through the inhibition of Lck and inactivation of Zap-70, an increase in the Foxp3 methylation status and, ultimately, the reduced production of suppressive cytokines. These results indicate a role of MGL as an immunomodulator within the tumour microenvironment interfering with Treg functions, suggesting its possible use in the design of anticancer vaccines.

No MeSH data available.


Related in: MedlinePlus

Binding of rhMGL-Fc to Tregs increases Foxp3 methylation and downregulates TCR signalling.(A) Percentage of Foxp3 methylation (TSDR region) of Treg cells alone or after treatment with rhMGL-Fc. The results correspond to the mean values obtained from three donors ± SD. (B) MFI values of FOXP3 expression of Tregs alone or after treatment with rhMGL-Fc stimulated with anti-CD3. The results correspond with the mean of three independent experiments ± SD. (C) Western blot analysis of Treg lysates with or without the treatment of rhMGL-Fc (left). Samples were analysed for pLck (Tyr 505), pLck (Tyr 394) and Zap-70. β-actin was used as a loading control. Proteins were resolved in 4–12% SDS-PAGE gel. Densitometric evaluation of the Lck-p505, Lck p-394, Zap-70 signals normalized to the levels of β-Actin (right). These results are representative of one donor out of three. ** Corresponds to p<0.01.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4493043&req=5

pone.0132617.g002: Binding of rhMGL-Fc to Tregs increases Foxp3 methylation and downregulates TCR signalling.(A) Percentage of Foxp3 methylation (TSDR region) of Treg cells alone or after treatment with rhMGL-Fc. The results correspond to the mean values obtained from three donors ± SD. (B) MFI values of FOXP3 expression of Tregs alone or after treatment with rhMGL-Fc stimulated with anti-CD3. The results correspond with the mean of three independent experiments ± SD. (C) Western blot analysis of Treg lysates with or without the treatment of rhMGL-Fc (left). Samples were analysed for pLck (Tyr 505), pLck (Tyr 394) and Zap-70. β-actin was used as a loading control. Proteins were resolved in 4–12% SDS-PAGE gel. Densitometric evaluation of the Lck-p505, Lck p-394, Zap-70 signals normalized to the levels of β-Actin (right). These results are representative of one donor out of three. ** Corresponds to p<0.01.

Mentions: The development and immunosuppressive function of Tregs are regulated by the transcriptional factor FOXP3 [13]. The mechanism maintaining stable Foxp3 expression in Tregs requires demethylation of the Treg specific demethylated region (TSDR) [14]. Therefore, increased methylation of this region reduces Foxp3 transcription, impairing Treg function. To evaluate whether changes in Foxp3 gene profiling could account for the decreased immunosuppressive activity of Tregs upon MGL-CD45RA interaction, we examined the methylation status of CpG sites in TSDR. Tregs alone and Tregs after rhMGL-Fc triggering were lysed for cDNA isolation and analysis. The results in Fig 2A show that the MGL binding on Tregs induced a significant increase of Foxp3 methylation (p<0.01). In fact, in unstimulated Tregs, Foxp3 methylation was 42%, while after MGL-CD45RA interaction, Foxp3 methylation became significantly higher (50%; p<0.01).


The Macrophage Galactose-Type C-Type Lectin (MGL) Modulates Regulatory T Cell Functions.

Zizzari IG, Martufi P, Battisti F, Rahimi H, Caponnetto S, Bellati F, Nuti M, Rughetti A, Napoletano C - PLoS ONE (2015)

Binding of rhMGL-Fc to Tregs increases Foxp3 methylation and downregulates TCR signalling.(A) Percentage of Foxp3 methylation (TSDR region) of Treg cells alone or after treatment with rhMGL-Fc. The results correspond to the mean values obtained from three donors ± SD. (B) MFI values of FOXP3 expression of Tregs alone or after treatment with rhMGL-Fc stimulated with anti-CD3. The results correspond with the mean of three independent experiments ± SD. (C) Western blot analysis of Treg lysates with or without the treatment of rhMGL-Fc (left). Samples were analysed for pLck (Tyr 505), pLck (Tyr 394) and Zap-70. β-actin was used as a loading control. Proteins were resolved in 4–12% SDS-PAGE gel. Densitometric evaluation of the Lck-p505, Lck p-394, Zap-70 signals normalized to the levels of β-Actin (right). These results are representative of one donor out of three. ** Corresponds to p<0.01.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4493043&req=5

pone.0132617.g002: Binding of rhMGL-Fc to Tregs increases Foxp3 methylation and downregulates TCR signalling.(A) Percentage of Foxp3 methylation (TSDR region) of Treg cells alone or after treatment with rhMGL-Fc. The results correspond to the mean values obtained from three donors ± SD. (B) MFI values of FOXP3 expression of Tregs alone or after treatment with rhMGL-Fc stimulated with anti-CD3. The results correspond with the mean of three independent experiments ± SD. (C) Western blot analysis of Treg lysates with or without the treatment of rhMGL-Fc (left). Samples were analysed for pLck (Tyr 505), pLck (Tyr 394) and Zap-70. β-actin was used as a loading control. Proteins were resolved in 4–12% SDS-PAGE gel. Densitometric evaluation of the Lck-p505, Lck p-394, Zap-70 signals normalized to the levels of β-Actin (right). These results are representative of one donor out of three. ** Corresponds to p<0.01.
Mentions: The development and immunosuppressive function of Tregs are regulated by the transcriptional factor FOXP3 [13]. The mechanism maintaining stable Foxp3 expression in Tregs requires demethylation of the Treg specific demethylated region (TSDR) [14]. Therefore, increased methylation of this region reduces Foxp3 transcription, impairing Treg function. To evaluate whether changes in Foxp3 gene profiling could account for the decreased immunosuppressive activity of Tregs upon MGL-CD45RA interaction, we examined the methylation status of CpG sites in TSDR. Tregs alone and Tregs after rhMGL-Fc triggering were lysed for cDNA isolation and analysis. The results in Fig 2A show that the MGL binding on Tregs induced a significant increase of Foxp3 methylation (p<0.01). In fact, in unstimulated Tregs, Foxp3 methylation was 42%, while after MGL-CD45RA interaction, Foxp3 methylation became significantly higher (50%; p<0.01).

Bottom Line: In this study, we demonstrate that Treg immunosuppressive activity can be modulated by the cross-linking between the CD45RA expressed by Tregs and the C-type lectin MGL.This effect can be attributed to changes in CD45RA and TCR signalling through the inhibition of Lck and inactivation of Zap-70, an increase in the Foxp3 methylation status and, ultimately, the reduced production of suppressive cytokines.These results indicate a role of MGL as an immunomodulator within the tumour microenvironment interfering with Treg functions, suggesting its possible use in the design of anticancer vaccines.

View Article: PubMed Central - PubMed

Affiliation: Department of Experimental Medicine, "Sapienza" University, Rome, Italy.

ABSTRACT
Regulatory T cells (Tregs) are physiologically designed to prevent autoimmune disease and maintain self-tolerance. In tumour microenvironments, their presence is related to a poor prognosis, and they influence the therapeutic outcome due to their capacity to suppress the immune response by cell-cell contact and to release immunosuppressive cytokines. In this study, we demonstrate that Treg immunosuppressive activity can be modulated by the cross-linking between the CD45RA expressed by Tregs and the C-type lectin MGL. This specific interaction strongly decreases the immunosuppressive activity of Tregs, restoring the proliferative capacity of co-cultured T lymphocytes. This effect can be attributed to changes in CD45RA and TCR signalling through the inhibition of Lck and inactivation of Zap-70, an increase in the Foxp3 methylation status and, ultimately, the reduced production of suppressive cytokines. These results indicate a role of MGL as an immunomodulator within the tumour microenvironment interfering with Treg functions, suggesting its possible use in the design of anticancer vaccines.

No MeSH data available.


Related in: MedlinePlus