Regulation of ureteric bud branching morphogenesis by sulfated proteoglycans in the developing kidney

Abstract

Glycosaminoglycans in the form of heparan sulfate proteoglycans (HSPG) and chondroitin sulfate proteoglycans (CSPG) are required for normal kidney organogenesis. The specific roles of HSPGs and CSPGs on ureteric bud (UB) branching morphogenesis are unclear, and past reports have obtained differing results. Here we employ in vitro systems, including isolated UB culture, to clarify the roles of HSPGs and CSPGs on this process. Microarray analysis revealed that many proteoglycan core proteins change during kidney development (syndecan-1,2,4, glypican-1,2,3, versican, decorin, biglycan). Moreover, syndecan-1, syndecan-4, glypican-3, and versican are differentially expressed during isolated UB culture, while decorin is dynamically regulated in cultured isolated metanephric mesenchyme (MM). Biochemical analysis indicated that while both heparan sulfate (HS) and chondroitin sulfate (CS) are present, CS accounts for approximately 75% of the glycosaminoglycans (GAG) in the embryonic kidney. Selective perturbation of HS in whole kidney rudiments and in the isolated UB resulted in a significant reduction in the number of UB branch tips, while CS perturbation has much less impressive effects on branching morphogenesis. Disruption of endogenous HS sulfation with chlorate resulted in diminished FGF2 binding and proliferation, which markedly altered kidney area but did not have a statistically significant effect on patterning of the ureteric tree. Furthermore, perturbation of GAGs did not have a detectable effect on FGFR2 expression or epithelial marker localization, suggesting the expression of these molecules is largely independent of HS function. Taken together, the data suggests that nonselective perturbation of HSPG function results in a general proliferation defect; selective perturbation of specific core proteins and/or GAG microstructure may result in branching pattern defects. Despite CS being the major GAG synthesized in the whole developing kidney, it appears to play a lesser role in UB branching; however, CS is likely to be integral to other developmental processes during nephrogenesis, possibly involving the MM. A model is presented of how, together with growth factors, heterogeneity of proteoglycan core proteins and glycosaminoglycan sulfation act as a switching mechanism to regulate different stages of the branching process. In this model, specific growth factor-HSPG combinations play key roles in the transitioning between stages and their maintenance.

Versican Mediates Mesenchymal-Epithelial Transition

“…In contrast, the formation of tight junctions was found in the V1-transfected NIH3T3 cells, suggesting that these transfected cells are polarized, and epithelialization of NIH3T3 cells induced by V1 can therefore be considered complete. The expression of versican and WT-1 were both detected during renal development (Pritchard-Jones et al, 1990;Hosono et al, 1999;Erickson and Couchman, 2001;Steer et al, 2004).…”

Transcriptional profiling of Wnt4 mutant mouse kidneys identifies genes expressed during nephron formation

“…These include the transmembrane heparan sulfate proteoglycan Sdc4. Interestingly sulfated proteoglycans modulate epithelial branching morphogenesis in organ culture (Steer et al, 2004). Also Tcf2, which has been previously associated with kidney disease (Coffinier et al, 1999;Edghill et al, 2006).…”

The GDNF Target Vsnl1 Marks the Ureteric Tip

“…30 Vsnl1 was highly expressed at the UB tips of isolated UBs growing on GDNF containing Matrigel 43 Canonical ␤-catenin activity has been shown to be antagonized by the calcium sensors. 44,45 To explore the possible relationship between Vsnl1 and stabilized ␤-catenin, we used the mIMCD3 cell line, transiently cotransfected with the TCF reporter TOPFLASH and full-length murine Vsnl1 cDNA (pCAb-Vsnl1-IRES-EGFPm) or with a control vector (pCAb-IRESEGFPm).…”

“…As described previously, 43 UBs were microdissected from E12 mouse embryonic kidneys. The UBs were then suspended within extracellular matrix gels (1:2 mixture of growth factor-reduced Matrigel and culture medium; BD Biosciences).…”