These observations are in agreement with other studies demonstrating that Dex treated DCs exhibit a re duced T cell stimulatory capacity either in allogeneic or antigen precise co cultures. Considering the MPLA tDC phenotype, characterized by a higher IL 10 and low IL 12 production, along with a lowered costimulatory and activation machinery, all features associated with induction of regulatory T populations, it nonetheless remains to be elucidated no matter whether the CD4 T cells that proliferate in co cultures with tDCs and MPLA tDCs correspond to induced Tregs or Tr1 cells. As we stated above, TolDCs will have to preserve their lymph node homing capacity primarily via CCR7 expression in an effort to be able to migrate to DC T cell zones and exert their regulatory impact.
Study on semi mature DCs with tolerogenic properties and activated with LPS soon after toler ance induction, demonstrated that this feature is crucial for TolDC migratory and antigen presentation capacities. Considering that our target order Neratinib is usually to generate TolDCs for clinical pur poses, we employed MPLA for tDCs activation, replacing the LPS stimulus described for cell activation, and further evaluated MPLA tDCs chemokine receptors expression and also the migratory response to their cognate ligands. Ana lysis showed that in addition to CCR7, activated TolDCs also upregulated CXCR4, a further chemokine receptor de scribed to be expressed on mature DCs and involved in DC migration to secondary lymphoid organs.
Ex pression of both chemokine receptors in MPLA tDCs at the same time as in mDCs suggests that these DC types are capable of migrating to DC T cell speak to websites for antigen presen tation, data supported by the results obtained in migration assays, demonstrating that mDCs and MPLA tDCs display migratory skills in response to CCL19 and CXCL12, ligands of CYC116 CCR7 and CXCR4, respectively. On the con trary, each mDCs and MPLA tDCs exhibited a decreased migratory response to CCL5, ligand of CCR5 and CCR1, a chemokine related to migration of leukocytes towards inflamed tissues, whilst each chemokine receptors were shown to be upregulated on iDCs. The distinction ob served in MPLA tDC migratory response towards CCL19 in comparison with mDCs, could be related to CCR7 expression levels determined for every single DC form, with mDCs exhibiting high response towards CCL19, and displaying a tendency to a larger expression of CCR7 in comparison to MPLA tDCs. These benefits are concordant with those reported by Anderson et al.
applying LPS, and with other previous research on DCs, showing that MPLA stimulation is sufficient to induce tDCs activation and migration, by triggering a switch in their chemokine receptor expression profile. Conclusions In synthesis, the present study describes a 5 day proto col for TolDCs generation employing Dex as immunomod ulatory agent, and MPLA, a LPS substitute, as tDCs activator for obtaining clinical grade TolDCs.