Characterization of Alport syndrome and DPP9 protease using Col4a3 knock-out mice and DPP9 protease dead mutant mice

1. Alport syndrome is a genetic disease of collagen IV (α3, 4, 5) resulting in defective assembly of glomerular basement membrane leads to renal failure. Col4a3KO mice are an established genetic model of autosomal recessive Alport syndrome. No sex differences in the evolution of body mass loss, renal pathology, biomarkers of tubular damage NGAL and KIM-1, or deterioration of kidney function were observed during the life span of Col4a3KO mice. These findings confirm that, similar to human autosomal recessive Alport syndrome, female and male Col4a3KO mice develop renal failure at the same age and with similar severity. The specific contribution of macrophage infiltration to Alport disease, one of the prominent features of the disease in human and Col4a3KO mice, remains unknown. This study shows that depletion of kidney macrophages in Col4a3KO male mice by administration of clodronate liposomes, prior to clinical onset of disease and throughout the study period, does not protect the mice from renal failure and interstitial fibrosis, nor delay disease progression. These results suggest that therapy targeting macrophage recruitment to kidney is unlikely to be effective as treatment of Alport syndrome.
2. Dipeptidyl peptidase 9 (DPP9), an intracellular N-terminal post-proline-cleaving enzyme, is implicated in cell behavior and immune responses but its physiological function remains largely unknown. Here we investigated in vivo role of DPP9 enzyme by characterizing DPP9 knock-in mice expressing a catalytically inactive S729A mutant of DPP9 (DPP9ki/ki mice). We show that impaired suckling response is a primary cause of perinatal lethality of DPP9ki/ki mice which die within 12-18 h after birth and can be rescued by manual feeding. Suckling defect is a result of microglossia characterized by defects in the formation of intrinsic distal tongue muscle known to derive from migratory muscle progenitors while intrinsic proximal and extrinsic tongue muscles deriving from head mesenchyme develop normally in DPP9ki/ki mice. Smaller tongue size of DPP9ki/ki mice correlates with reduced number and impaired survival of migratory tongue muscle progenitors. CXCR4 mediated survival and migration of muscle progenitors is not impaired in the absence of DPP9 enzymatic activity however, an effect of DPP9 enzyme on SDF-1, ligand of CXCR4, cannot be excluded. In addition, we show that DPP9ki/ki mice exhibit impaired fetal hematopoiesis however their hematopoietic stem cells are fully competent in reconstitution of myeloid and lymphoid lineages in lethally irradiated mice. Taken together, our studies revealed for the first time that DPP9 enzyme controls survival of migratory tongue muscle progenitors and its absence in mice leads to impaired tongue development, suckling defect and neonatal lethality but DPP9 enzymatic activity in hematopoietic cells is dispensable for normal hematopoiesis.