MSC-derived MMP-processed CCL2/CCL7 block IgG secretion. (A) MSCs block IgG secretion via CCL2/7. An ELISPOT assay was performed in the presence of CCL2/8 and CCL7 neutralizing antibodies. IgG secretion is partially restored on CCL2 and CCL7 neutralization, but a complete rescue is achieved after simultaneous inhibition of both CCLs (n = 3/group; P < .001). (B) MSCs secrete antagonist CCL2. To prove that a truncated form of CCL2 with antagonistic activities is responsible for IgG blockade, in vitro MMP1-mpCCL2 was added on OVA-derived splenocytes and led to 90% IgG inhibition (n = 3/group; P < .001). (C) MSCs secrete MMP-1, MMP-3, and MMP-8. To generate the cleaved form of CCL2, an MMP digestion is required. MMPs were indeed detected in MSC CM by Western blot (WB). (D) MMP inhibition rescues IgG blockade. Because MMP enzymatic cleavage of CCL2 as well as CCL7 is responsible for the IgG secretion inhibition, we added actinonin (a generic MMP inhibitor) and demonstrate that PCs can secrete IgG after MSC CM treatment in a way comparable with the nontreated splenocytes (n = 3/group; P < .05). The negative control consists of splenocytes from naive mice. All experiments were repeated 3 times. Error bars represent plus or minus SD.