The effect of hyaluronic acid size and concentration on branching morphogenesis and tubule differentiation in developing kidney culture systems: Potential applications to engineering of renal tissues

Abstract

Hyaluronic acid (HA) is a glycosaminoglycan of tissue engineering importance that plays a vital role in mammalian development. In vitro kidney culture methods were utilized to investigate the importance of HA during renal organogenesis. We found that HA has the ability to simultaneously modulate ureteric bud (UB) branching, promote mesenchymal-to-epithelial transformation, and promote differentiation of both metanephric mesenchyme (MM) and the UB depending on the concentration and molecular weight (MW) of HA. Hyaluronidase inhibited branching morphogenesis in both isolated UB and whole kidney cultures, suggesting endogenous HA is required for branching morphogenesis. HA exhibited morphogen-like properties, stimulating branching morphogenesis at low concentrations (0.1%) and low MW (6.55 kDa) but inhibiting at high concentrations (3.75%) and high MW (234.4 kDa). Furthermore, HA of every MW tested promoted collecting duct differentiation as measured by AQP-2 expression. E-cadherin immunostaining and qPCR of nephron differentiation markers (OAT-1, NaP i -2, AQP-1, and THP), demonstrated that HA of a variety of MWs strongly promotes mesenchymal epithelialization and nephron differentiation in a concentration-dependent manner. Since the HA synthesis and degradation genes, has-2 and hyal-2, are highly expressed during kidney development, this data suggests that specific sizes and concentrations of HA may act to independently regulate UB branching and promote tubular maturation, representing a potential switch for ending branching morphogenesis, as well as initiating nephron differentiation. In addition, the ability of HA to promote in vitro embryonic kidney growth and maturation, together with the biocompatibility and crosslinking capability of HA, suggests a potential use of HA for both creating an instructive, 3D scaffold for in vitro kidney engineering from developmental tissues, as well as promoting tubule regeneration in injured or cryopreserved kidneys.

Constructing Kidney-like Tissues from Cells Based on Programs for Organ Development: Toward a Method ofIn VitroTissue Engineering of the Kidney

“…6,11,17 The localization of Flk-1, Pod, vW factor, E-cadherin, cytokeratin, and/or PAX-2 (1:500 in blocking solution) was determined in the samples. For PNA staining, after the blocking step, the tissues were washed twice with Neuraminidase buffer (150 mM NaCl and 50 mM sodium acetate, pH 5.5), and incubated with Neuraminidase (1 unit/mL) for 4 h at 378C, and then with rhodamine-conjugated PNA (50 mg/mL) and FITC-DB (1:500) for 24 h at 48C.…”

Immuno-Localization of CD44 and Osteopontin in Developing Human Kidney

“…By this mechanism, local HA molecular weight and concentration may promote ureteric bud branching early in development, and bring about the cessation of branching at the appropriate time. Local differences in HA molecular weight and concentration may be regulated via developmental and topographic expression of enzymes responsible for HA synthesis and degradation (27). These observations suggest a critical role for HA during early renal development.…”

Correlation between hyaluronic acid, hyaluronic acid synthase and human renal clear cell carcinoma

“…HA has broad participation in the embryogenesis of mammalian kidney [7] , has important role in the process of urinary concentration in normal kidney [8] . Furthermore, HA has taken part in the process of occurrence, development and fibrosis in different kinds of nephritis and diabetic nephropathy; HA also makes a difference to the pathologic process of renal ischemia-reperfusion injury and kidney tubular crystallization [9][10][11][12] .…”