Regulation of Renal Differentiation by Trophic Factors

Classically, trophic factors are considered as proteins which support neurons in their growth, survival, and differentiation. However, most neurotrophic factors also have important functions outside of the nervous system. Especially essential renal growth and differentiation regulators are glial cell line-derived neurotrophic factor (GDNF), bone morphogenetic proteins (BMPs), and fibroblast growth factors (FGFs). Here we discuss how trophic factor-induced signaling contributes to the control of ureteric bud (UB) branching morphogenesis and to maintenance and differentiation of nephrogenic mesenchyme in embryonic kidney. The review includes recent advances in trophic factor functions during the guidance of branching morphogenesis and self-renewal versus differentiation decisions, both of which dictate the control of kidney size and nephron number. Creative utilization of current information may help better recapitulate renal differentiation in vitro, but it is obvious that significantly more basic knowledge is needed for development of regeneration-based renal therapies

MAPK/ERK Signaling in Regulation of Renal Differentiation

Congenital anomalies of the kidney and urinary tract (CAKUT) are common birth defects derived from abnormalities in renal differentiation during embryogenesis. CAKUT is the major cause of end-stage renal disease and chronic kidney diseases in children, but its genetic causes remain largely unresolved. Here we discuss advances in the understanding of how mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK) activity contributes to the regulation of ureteric bud branching morphogenesis, which dictates the final size, shape, and nephron number of the kidney. Recent studies also demonstrate that the MAPK/ERK pathway is directly involved in nephrogenesis, regulating both the maintenance and differentiation of the nephrogenic mesenchyme. Interestingly, aberrant MAPK/ERK signaling is linked to many cancers, and recent studies suggest it also plays a role in the most common pediatric renal cancer, Wilms’ tumor

ShapeMetrics: a userfriendly pipeline for 3D cell segmentation and spatial tissue analysis

The demand for single-cell level data is constantly increasing within life sciences. In order to meet this demand, robust cell segmentation methods that can tackle challenging in vivo tissues with complex morphology are required. However, currently available cell segmentation and volumetric analysis methods perform poorly on 3D images. Here, we generated ShapeMetrics, a MATLAB-based script that segments cells in 3D and, by performing unbiased clustering using a heatmap, separates the cells into subgroups according to their volumetric and morphological differences. The cells can be accurately segregated according to different biologically meaningful features such as cell ellipticity, longest axis, cell elongation, or the ratio between cell volume and surface area. Our machine learning based script enables dissection of a large amount of novel data from microscope images in addition to the traditional information based on fluorescent biomarkers. Furthermore, the cells in different subgroups can be spatially mapped back to their original locations in the tissue image to help elucidate their roles in their respective morphological contexts. In order to facilitate the transition from bulk analysis to single-cell level accuracy, we emphasize the user-friendliness of our method by providing detailed step-by-step instructions through the pipeline hence aiming to reach users with less experience in computational biology.Peer reviewe

Development of the urogenital system is regulated via the 3 ' UTR of GDNF

Mechanisms controlling ureter lenght and the position of the kidney are poorly understood. Glial cellline derived neurotrophic factor (GDNF) induced RET signaling is critical for ureteric bud outgrowth, but the function of endogenous GDNF in further renal differentiation and urogenital system development remains discursive. Here we analyzed mice where 3' untranslated region (UTR) of GDNF is replaced with sequence less responsive to microRNA-mediated regulation, leading to increased GDNF expression specifically in cells naturally transcribing Gdnf. We demonstrate that increased Gdnf leads to short ureters in kidneys located in an abnormally caudal position thus resembling human pelvic kidneys. High GDNF levels expand collecting ductal progenitors at the expense of ureteric trunk elongation and result in expanded tip and short trunk phenotype due to changes in cell cycle length and progenitor motility. MEK-inhibition rescues these defects suggesting that MAPK-activity mediates GDNF's effects on progenitors. Moreover, Gdnf(hyper) mice are infertile likely due to effects of excess GDNF on distal ureter remodeling. Our findings suggest that dysregulation of GDNF levels, for example via alterations in 3' UTR, may account for a subset of congenital anomalies of the kidney and urinary tract (CAKUT) and/or congenital infertility cases in humans and pave way to future studies.Peer reviewe

ETS-related transcription factors Etv4 and Etv5 are involved in proliferation and induction of differentiationassociated genes in embryonic stem (ES) cells

The pluripotency and self-renewal capacity of embryonic stem (ES) cells is regulated by several transcription factors. Here, we show that the ETS-related transcription factors Etv4 and Etv5 (Etv4/5) are specifically expressed in undifferentiated ES cells, and suppression of Oct3/4 results in down-regulation of Etv4/5. Simultaneous deletion of Etv4 and Etv5 (Etv4/5 double knock-out(dKO)) in ES cells resulted in a flat, epithelial cell-like appearance, whereas the morphology changed into compact colonies in a 2i medium (containing two inhibitors for GSK3 and MEK/ERK). Expression levels of self-renewal marker genes, including Oct3/4 and Nanog, were similar between wild-type and dKO ES cells, whereas proliferation of Etv4/5 dKO ES cells was decreased with overexpression of cyclin-dependent kinase inhibitors (p16/p19, p15, and p57). A differentiation assay revealed that the embryoid bodies derived from Etv4/5 dKO ES cells were smaller than the control, and expression of ectoderm marker genes, including Fgf5, Sox1, and Pax3, was not induced in dKO-derived embryoid bodies. Microarray analysis demonstrated that stem cell-related genes, including Tcf15, Gbx2, Lrh1, Zic3, and Baf60c, were significantly repressed in Etv4/5dKOEScells.Theartificial expression of Etv4 and/or Etv5 in Etv4/5 dKO ES cells induced re-expression of Tcf15 and Gbx2. These results indicate that Etv4 and Etv5, potentially through regulation of Gbx2 and Tcf15, are involved in the ES cell proliferation and induction of differentiation-associated genes in ES cells. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc. Published in the U.S.A

Sticklebacks adapted to divergent osmotic environments show differences in plasticity for kidney morphology and candidate gene expression

Novel physiological challenges in different environments can promote the evolution of divergent phenotypes, either through plastic or genetic changes. Environmental salinity serves as a key barrier to the distribution of nearly all aquatic organisms, and species diversification is likely to be enabled by adaptation to alternative osmotic environments. The threespine stickleback (Gasterosteus aculeatus) is a euryhaline species with populations found both in marine and freshwater environments. It has evolved both highly plastic and locally adapted phenotypes due to salinity-derived selection, but the physiological and genetic basis of adaptation to salinity is not fully understood. We integrated comparative cellular morphology of the kidney, a key organ for osmoregulation, and candidate gene expression to explore the underpinnings of evolved variation in osmotic plasticity within two populations of sticklebacks from distinct salinity zones in the Baltic Sea: the high salinity Kattegat, representative of the ancestral marine habitat; and the low salinity Bay of Bothnia. A common-garden experiment revealed that kidney morphology in the ancestral high-salinity population had a highly plastic response to salinity conditions whereas this plastic response was reduced in the low-salinity population. Candidate gene expression in kidney tissue revealed a similar pattern of population specific differences, with a higher degree of plasticity in the native high-salinity population. Together these results suggest that renal cellular morphology has become canalized to low salinity, and that these structural differences may have functional implications for osmoregulation.Peer reviewe