Microglial Correction After Fetal Therapy Without Conditioning In Mice With Mucopolysaccharidosis Type Vii

Abstract Introduction Mucopolysaccharidosis type 7 (MPS7) is a lysosomal storage disorder typically fatal in utero. Postnatal enzyme replacement therapy (ERT) to replace missing glucuronidase (GUS) does not penetrate the blood-brain barrier (BBB). We investigated whether in utero ERT (IUERT) specifically targets brain microglia (natural GUS storehouses and key brain inflammation mediators) and whether in utero hematopoietic stem cell transplantation (IUHSCT) results in microglial engraftment as a strategy for permanent correction. Methods We performed IUERT by injecting GUS into MPS7 fetuses mid-gestation, and analyzed tissue homogenates (via colorimetric substrate) and brain microglia (via flow cytometry) for enzyme activity after 4-7 days. We performed IUHSCT by transplanting HSCs mid-gestation from CX3CR1-GFP donors. We examined blood, bone marrow, and brain for engraftment. We assessed brain inflammation by staining for CD68. We performed RNA sequencing to characterize engrafted microglia. Results IUERT resulted in detectable brain GUS activity. Flow cytometry showed that GUS activity after IUERT was near wild-type levels, and brains harvested in adulthood had decreased inflammation via CD68 immunohistochemistry. IUHSCT resulted in multilineage engraftment of hematopoietic cells in blood and bone. Confocal microscopy revealed multifocal engraftment of donor-derived microglia. RNA sequencing indicated that engrafted microglia were nearly identical to endogenous microglia. MPS7 chimeras had evidence of reduced brain inflammation near donor microglia. Conclusion Both IUERT and IUHSCT are complementary treatment modalities that can penetrate the BBB and ameliorate neurologic manifestations of diseases such as MPS7. These results lay the foundation for future studies using in utero molecular therapies for MPS7 as well as other storage disorders.