Significance Statement: Mesangial cells play a crucial role in maintaining glomerular homeostasis and injuries to these cells often result in progression to CKD like IgA and diabetic nephropathies. However, the transcription factors involved in mesangial cell development and function are largely unknown. The authors describe the role transcription factor GATA3 plays in mesangial cells in embryonic kidneys and healthy and injured adult glomeruli. Mice with haploinsufficiency of GATA3 have too few MC precursor cells and glomerular abnormalities. GATA3 expression increases in mesangial cells in mesangial proliferative GN in humans and rodent models suggesting GATA3 is important for glomerular homeostasis and response to injury. GATA3 also may be a useful a nuclear marker of human mesangial cells. Background: GATA3 is a dual-zinc finger transcription factor that regulates gene expression in many developing tissues. In the kidney, GATA3 is essential for ureteric bud branching, and mice without it fail to develop kidneys. In humans, autosomal dominant GATA3 mutations can cause renal aplasia as part of the hypoparathyroidism, renal dysplasia, deafness (HDR) syndrome that includes mesangioproliferative GN. This suggests that GATA3 may have a previously unrecognized role in glomerular development or injury. Methods: To determine GATA3’s role in glomerular development or injury, we assessed GATA3 expression in developing and mature kidneys from Gata3 heterozygous (⁺/−) knockout mice, as well as injured human and rodent kidneys. Results: We show that GATA3 is expressed by FOXD1 lineage stromal progenitor cells, and a subset of these cells mature into mesangial cells (MCs) that continue to express GATA3 in adult kidneys. In mice, we uncover that GATA3 is essential for normal glomerular development, and mice with haploinsufficiency of Gata3 have too few MC precursors and glomerular abnormalities. Expression of GATA3 is maintained in MCs of adult kidneys and is markedly increased in rodent models of mesangioproliferative GN and in IgA nephropathy, suggesting that GATA3 plays a critical role in the maintenance of glomerular homeostasis. Conclusions: These results provide new insights on the role GATA3 plays in MC development and response to injury. It also shows that GATA3 may be a novel and robust nuclear marker for identifying MCs in tissue sections.

Vascular calcification is a strong predictor of cardiovascular mortality and lacks effective treatment. The transformation of vascular smooth muscle cells (VSMCs) into osteoblast-like phenotypes is a key driver of calcification. This study identifies a regulatory role for Hyaluronan (HA) in VSMC osteogenic differentiation and arterial calcification. Human aortic VSMCs stimulated with high phosphate and/or pro-inflammatory cytokines (IL6 and TGF-β1) exhibited increased RUNX2, alkaline phosphatase and osteopontin expression, along with reduced contractility and elevated calcium deposition. This corresponded with reduced HA deposition and downregulation of HA synthase enzymes (HAS1, HAS2), Hyaluronidase enzymes (Hyal1), and HA binding proteins (CD44, TSG-6), whilst HAS3 and versican were upregulated. Comparable alterations in HA and protein expression were observed in an in vivo model of arterial calcification using vitamin K-deficient warfarin-fed mice. Pharmacological inhibition of HA synthesis, enzyme-mediated HA degradation and siRNA/plasmid modulation of HAS isoenzymes demonstrated a possible functional link between HA regulation and VSMC osteogenic differentiation. This study establishes HA and its associated binding proteins as key regulators of arterial calcification, highlighting a novel pathway for potential therapeutic intervention.

The stroma plays key roles in development and tissue regeneration after injury, but its expansion following injury can drive fibrosis and chronic disease. In the kidney, hyaluronan (HA) is largely absent from the healthy stroma but becomes abundant during disease. To investigate HA regulation in stromal responses, we compared HA and hyaluronan synthase (HAS) expression during kidney development, homeostasis, injury, and recovery. A rat model of ischaemia–reperfusion injury (IRI) with or without ischaemic pre-conditioning (IPC) was studied alongside embryonic and Has1/3^−/− mouse kidneys, with mechanistic studies performed in primary human fibroblasts. In healthy kidneys, stromal HA was absent, HAS1 showed limited epithelial expression, and HAS2 was undetectable. Following IRI, stromal HA accumulation and increased HAS2 expression were associated with chronic fibrosis. In contrast, during development, prominent stromal HA matrices were evident in areas of tubular generation, with strong HAS1 (not HAS2) expression. During recovery, stromal HA and HAS2 were attenuated, whereas HAS1+ stromal cells expanded but were distinct from α-SMA+ myofibroblasts. Functionally, HAS1+ fibroblasts displayed enhanced migration, fibroblast activation protein (FAP) expression and reduced profibrotic marker expression, whereas HAS2+ fibroblasts exhibited a contractile myofibroblast phenotype with high extra domain A fibronectin (EDA-FN) and type I collagen, alpha-1 subunit (COL1A1). Together, these findings identify HA as a regulator of stromal fibroblast phenotype and associate HAS1 and HAS2 in divergent fibrotic outcomes following kidney injury.