DEFINING TISSUE LEVEL ARCHITECTURE CHANGES IN EXTRACELLULAR MATRIX DURING MURINE KIDNEY AND FORELIMB MYOTENDINOUS JUNCTION DEVELOPMENT

Congenital diseases of the kidney are the leading cause of chronic kidney disease in pediatric patients. Tissue engineering models used to investigate these diseases are limited by an immature phenotype. Models cultured in an extracellular matrix (ECM), a network of proteins and glycosaminoglycans surrounding cells and providing structural support that mimic the matrix found in development will be likely more mature. However, developing kidney ECM composition and structural dynamics are unknown. To address this gap, we studied ECM composition using mass spectrometry and organization by visualizing the ECM in 3D.

In this work, we used mass spectrometry to resolve ECM basement membrane and interstitial matrix dynamics between embryonic, perinatal, and adult kidneys. Surprisingly, we observed a transient upregulation of interstitial matrix structures that corresponded to dynamic 3D structures in the cortex (vertical fibers) and at the corticomedullary junction (medullary ray sheath fibers). Notably, in a model of abnormal Foxd1+ stromal cells, the vertical fibers were disorganized, and medullary ray sheath fibers were no longer associated with blood vessels, suggesting the dynamic 3D structures depended on stromal cell modulation.

One of the effects of abnormal kidney development is decreased amniotic fluid, which limits embryonic movement and subsequent limb development. In additional studies, we looked at the implications of the lost motility in the muscular dysgenesis (mdg) mouse on the development of the myotendinous junction (MTJ). The MTJ links contractile muscle with tendon. We found the ECM protein COL22A1 was specific to the developing MTJ as early as embryonic day (E)13.5. The development of the MTJ from a linear structure to a cap-like structure with invaginations in adolescent mice depended on muscle contraction. Furthermore, we used a model to decouple the muscle-tendon-bone complex at an ectopic lateral triceps insertion (Prrx1Cretg/+; Tbx3fl/fl). We observed disorganized tendon and MTJ markers at the termination of the ectopic lateral triceps muscle but negligible cartilage markers. Together, this indicated MTJ maturation depended on motility but not on the enthesis.

The information gleaned from our studies on how stromal cells affect dynamic 3D interstitial ECM structures and composition change during kidney development can be used as a template for 3D kidney culture systems. Combined with forelimb MTJ development, our results indicate the importance of the interstitial matrix in tissue morphogenesis.