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Little is known about regulation of the intestinal stem cell (ISC) compartment in gut GVHD. We have found that Interleukin-22 (IL-22) is important for ISC recovery after BMT. However, the mechanism and specific epithelial targets of IL-22 are poorly understood.
Using clinically modeled LP into C57BL/6 (B6) minor antigen-mismatched BMT (H-2b intoH-2b), we found that daily treatment with recombinant murine IL-22 (4ug IP starting D7 post-BMT) led to reduced GVHD pathology in small intestine (SI) and large intestine (LI) three weeks after transplant (p<0.001) without altering GVL or alloreactivity. Mice treated with IL-22 (no pharmacologic immunosuppression) maintained more Lgr5+ ISCs and demonstrated significantly greater ISC proliferation (p<0.01). Although IL-22 is thought to regulate Paneth cells (PCs), and PCs make up the SI stem cell niche, IL-22-mediated protection of ISCs was not due to niche augmentation, as PC numbers, PC-derived growth factors (EGF, Wnt3), and stroma-derived growth factors (R-spondin 3) were all unchanged after IL-22 administration. However, antimicrobial proteins Reg3β and Reg3γ were both upregulated in IL-22-treated mice (p<0.01), suggesting enhanced gut barrier function.
To evaluate direct effects on epithelial regeneration, we performed intestinal organoid cultures in the presence of IL-22. Organoids generated from wild-type B6 crypts demonstrated substantially increased size after seven days of culture with IL-22 (SI p<0.001, Fig. 1; LI p<0.05). Co-culturing crypts with innate lymphoid cells, potent producers of IL-22 in vivo, also led to increased organoid size. IL-22 significantly increased organoid EdU incorporation and new crypt budding, and serial passaging with IL-22 greatly increased organoid expansion, suggesting that IL-22 could augment ISC regeneration. IL-22 receptor staining further indicated a direct effect of IL-22 on ISCs rather than the stem cell niche, as PCs were found to express very little IL-22 receptor (Fig. 2). Indeed, IL-22 led to expansion of the Lgr5+ ISC pool within SI organoids (Fig. 2), activated STAT3 phosphorylation in Lgr5+ cells, and increased size and budding of organoids cultured from purified SI ISCs (p<0.05).
To investigate potential for clinical translation, we next tested a recombinant human IL-22 dimer/Fc fusion molecule (F-652, Generon Corp., Shanghai). F-652 significantly increased the size of SI and LI organoids (p<.05). Using the LP into B6 BMT model described above, we also found that treatment with F-652 (100 ug/kg subQ every other day starting D7 post-BMT) significantly improved both systemic GVHD scoring and survival (Fig. 3).
In summary, we found that IL-22 can bridge immune function and tissue regeneration by acting directly on epithelial stem cells. IL-22 therapy may represent a novel approach to promote intestinal recovery in GVHD patients without increasing post-transplant immunodeficiency.
