Cells of the renin lineage promote kidney regeneration post-release of ureteral obstruction in neonatal mice.

AIM: Ureteral obstruction leads to significant changes in kidney renin expression. It is unclear whether those changes are responsible for the progression of kidney damage, repair, or regeneration. In the current study, we aimed to elucidate the contribution of renin-producing cells (RPCs) and the cells of the renin lineage (CoRL) towards kidney damage and regeneration using a model of partial and reversible unilateral ureteral obstruction (pUUO) in neonatal mice. METHODS: Renin cells are progenitors for other renal cell types collectively called CoRL. We labeled the CoRL with green fluorescent protein (GFP) using genetic approaches. We performed lineage tracing to analyze the changes in the distribution of CoRL during and after the release of obstruction. We also ablated the RPCs and CoRL by cell-specific expression of Diphtheria Toxin Sub-unit A (DTA). Finally, we evaluated the kidney damage and regeneration during and after the release of obstruction in the absence of CoRL. RESULTS: In the obstructed kidneys, there was a 163% increase in the renin-positive area and a remarkable increase in the distribution of GFP+ CoRL. Relief of obstruction abrogated these changes. In addition, DTA-expressing animals did not respond to pUUO with increased RPCs and CoRL. Moreover, reduction in CoRL significantly compromised the kidney's ability to recover from the damage after the release of obstruction. CONCLUSIONS: CoRL play a role in the regeneration of the kidneys post-relief of obstruction.

Overexpression of notch signaling in renin cells leads to a polycystic kidney phenotype.

Polycystic kidney disease (PKD) is an inherited disorder that results in large kidneys, numerous fluid-filled cysts, and ultimately end-stage kidney disease. PKD is either autosomal dominant caused by mutations in PKD1 or PKD2 genes or autosomal recessive caused by mutations in the PKHD1 or DZIP1L genes. While the genetic basis of PKD is known, the downstream molecular mechanisms and signaling pathways that lead to deregulation of proliferation, apoptosis, and differentiation are not completely understood. The Notch pathway plays critical roles during kidney development including directing differentiation of various progenitor cells, and aberrant Notch signaling results in gross alternations in cell fate. In the present study, we generated and studied transgenic mice that have overexpression of an intracellular fragment of mouse Notch1 ('NotchIC') in renin-expressing cells. Mice with overexpression of NotchIC in renin-expressing cells developed numerous fluid-filled cysts, enlarged kidneys, anemia, renal insufficiency, and early death. Cysts developed in both glomeruli and proximal tubules, had increased proliferation marks, and had increased levels of Myc. The present work implicates the Notch signaling pathway as a central player in PKD pathogenesis and suggests that the Notch-Myc axis may be an important target for therapeutic intervention.

Renin Cell Baroreceptor, a Nuclear Mechanotransducer Central for Homeostasis.

[Figure: see text].

A primitive type of renin-expressing lymphocyte protects the organism against infections.

The hormone renin plays a crucial role in the regulation of blood pressure and fluid-electrolyte homeostasis. Normally, renin is synthesized by juxtaglomerular (JG) cells, a specialized group of myoepithelial cells located near the entrance to the kidney glomeruli. In response to low blood pressure and/or a decrease in extracellular fluid volume (as it occurs during dehydration, hypotension, or septic shock) JG cells respond by releasing renin to the circulation to reestablish homeostasis. Interestingly, renin-expressing cells also exist outside of the kidney, where their function has remained a mystery. We discovered a unique type of renin-expressing B-1 lymphocyte that may have unrecognized roles in defending the organism against infections. These cells synthesize renin, entrap and phagocyte bacteria and control bacterial growth. The ability of renin-bearing lymphocytes to control infections-which is enhanced by the presence of renin-adds a novel, previously unsuspected dimension to the defense role of renin-expressing cells, linking the endocrine control of circulatory homeostasis with the immune control of infections to ensure survival.

Renin-Expressing Cells Require ß1-Integrin for Survival and for Development and Maintenance of the Renal Vasculature.

Juxtaglomerular cells are crucial for blood pressure and fluid-electrolyte homeostasis. The factors that maintain the life of renin cells are unknown. In vivo, renin cells receive constant cell-to-cell, mechanical, and neurohumoral stimulation that maintain their identity and function. Whether the presence of this niche is crucial for the vitality of the juxtaglomerular cells is unknown. Integrins are the largest family of cell adhesion molecules that mediate cell-to-cell and cell-to-matrix interactions. Of those, ß1-integrin is the most abundant in juxtaglomerular cells. However, its role in renin cell identity and function has not been ascertained. To test the hypothesis that cell-matrix interactions are fundamental not only to maintain the identity and function of juxtaglomerular cells but also to keep them alive, we deleted ß1-integrin in vivo in cells of the renin lineage. In mutant mice, renin cells died by apoptosis, resulting in decreased circulating renin, hypotension, severe renal-vascular abnormalities, and renal failure. Results indicate that cell-to-cell and cell-to-matrix interactions via ß1-integrin is essential for juxtaglomerular cells survival, suggesting that the juxtaglomerular niche is crucial not only for the tight regulation of renin release but also for juxtaglomerular cell survival-a sine qua non condition to maintain homeostasis.

Super-enhancers maintain renin-expressing cell identity and memory to preserve multi-system homeostasis.

Renin cells are crucial for survival - they control fluid-electrolyte and blood pressure homeostasis, vascular development, regeneration, and oxygen delivery to tissues. During embryonic development, renin cells are progenitors for multiple cell types that retain the memory of the renin phenotype. When there is a threat to survival, those descendants are transformed and reenact the renin phenotype to restore homeostasis. We tested the hypothesis that the molecular memory of the renin phenotype resides in unique regions and states of these cells' chromatin. Using renin cells at various stages of stimulation, we identified regions in the genome where the chromatin is open for transcription, mapped histone modifications characteristic of active enhancers such as H3K27ac, and tracked deposition of transcriptional activators such as Med1, whose deletion results in ablation of renin expression and low blood pressure. Using the rank ordering of super-enhancers, epigenetic rewriting, and enhancer deletion analysis, we found that renin cells harbor a unique set of super-enhancers that determine their identity. The most prominent renin super-enhancer may act as a chromatin sensor of signals that convey the physiologic status of the organism, and is responsible for the transformation of renin cell descendants to the renin phenotype, a fundamental process to ensure homeostasis.

Identification of cardiac hemo-vascular precursors and their requirement of sphingosine-1-phosphate receptor 1 for heart development.

The cardiac endothelium plays a crucial role in the development of a functional heart. However, the precise identification of the endocardial precursors and the mechanisms they require for their role in heart morphogenesis are not well understood. Using in vivo and in vitro cell fate tracing concomitant with specific cell ablation and embryonic heart transplantation studies, we identified a unique set of precursors which possess hemogenic functions and express the stem cell leukemia (SCL) gene driven by its 5' enhancer. These hemo-vascular precursors give rise to the endocardium, atrioventricular cushions and coronary vascular endothelium. Furthermore, deletion of the sphingosine-1-phosphate receptor 1 (S1P1) in these precursors leads to ventricular non-compaction cardiomyopathy, a poorly understood condition leading to heart failure and early mortality. Thus, we identified a distinctive population of hemo-vascular precursors which require S1P1 to exert their functions and are essential for cardiac morphogenesis.

Lemierre's Syndrome as a Trigger for Secondary Hemophagocytic Lymphohistiocytosis.

Secondary hemophagocytic lymphohistiocytosis (sHLH) is a rare hyperinflammatory disorder caused by an aberrant immune response to a number of infectious or inflammatory conditions. Successful treatment of this potentially fatal condition requires early recognition and prompt therapy directed at the underlying trigger. In this report, we describe the clinical presentation, diagnostic findings, management, and outcome of a child with Lemierre's syndrome-associated sHLH. This is the first reported association of these 2 rare conditions and expands the number of known triggers for sHLH.

Loss of Jagged1 in renin progenitors leads to focal kidney fibrosis.

The Notch signaling pathway is required to maintain renin expression within juxtaglomerular (JG) cells. However, the specific ligand which activates Notch signaling in renin-expressing cells remains undefined. In this study, we found that among all Notch ligands, Jagged1 is differentially expressed in renin cells with higher expression during neonatal life. We therefore hypothesized that Jagged1 was involved in renin expression and/or vascular integrity. We used a conditional knockout approach to delete Jagged1 in cells of the renin lineage. Deletion of Jagged1 specifically within renin cells did not result in decreased renin production within the kidney. However, animals with conditional deletion of Jagged1 did develop focal kidney fibrosis and elevated blood urea nitrogen. Our data demonstrate that Jagged1-mediated Notch signaling is dispensable in renin cells of the kidney in regard to renin expression. However, deletion of Jagged1 in renin cells descendants affects perivascular-interstitial integrity leading to focal fibrosis and diminished renal function.

Identification of renin progenitors in the mouse bone marrow that give rise to B-cell leukaemia.

The cell of origin and triggering events for leukaemia are mostly unknown. Here we show that the bone marrow contains a progenitor that expresses renin throughout development and possesses a B-lymphocyte pedigree. This cell requires RBP-J to differentiate. Deletion of RBP-J in these renin-expressing progenitors enriches the precursor B-cell gene programme and constrains lymphocyte differentiation, facilitated by H3K4me3 activating marks in genes that control the pre-B stage. Mutant cells undergo neoplastic transformation, and mice develop a highly penetrant B-cell leukaemia with multi-organ infiltration and early death. These renin-expressing cells appear uniquely vulnerable as other conditional models of RBP-J deletion do not result in leukaemia. The discovery of these unique renin progenitors in the bone marrow and the model of leukaemia described herein may enhance our understanding of normal and neoplastic haematopoiesis.

Human rhabdomyosarcoma cell lines for rhabdomyosarcoma research: utility and pitfalls.

Rhabdomyosarcoma (RMS) is the most common soft tissue sarcoma of childhood and adolescence. Despite intergroup clinical trials conducted in Europe and North America, outcomes for high risk patients with this disease have not significantly improved in the last several decades, and survival of metastatic or relapsed disease remains extremely poor. Accrual into new clinical trials is slow and difficult, so in vitro cell-line research and in vivo xenograft models present an attractive alternative for preclinical research for this cancer type. Currently, 30 commonly used human RMS cell lines exist, with differing origins, karyotypes, histologies, and methods of validation. Selecting an appropriate cell line for RMS research has important implications for outcomes. There are also potential pitfalls in using certain cell lines including contamination with murine stromal cells, cross-contamination between cell lines, discordance between the cell line and its associated original tumor, imposter cell lines, and nomenclature errors that result in the circulation of two or more presumed unique cell lines that are actually from the same origin. These pitfalls can be avoided by testing for species-specific isoenzymes, microarray analysis, assays for subtype-specific fusion products, and short tandem repeat analysis.

Inhibition of the Notch-Hey1 axis blocks embryonal rhabdomyosarcoma tumorigenesis.

PURPOSE: Rhabdomyosarcoma (RMS) is the most common soft tissue sarcoma of childhood and remains refractory to combined-modality therapy in patients with high risk disease. In skeletal myogenesis, Notch signaling prevents muscle differentiation and promotes proliferation of satellite cell progeny. Given its physiologic role in myogenesis and oncogenic role in other human cancers, we hypothesized that aberrant Notch signaling may contribute to RMS tumorigenesis and present novel therapeutic opportunities. EXPERIMENTAL DESIGN: Human RMS cell lines and tumors were evaluated by immunoblot, IHC, and RT-PCR to measure Notch ligand, receptor, and target gene expression. Manipulation of Notch signaling was accomplished using genetic and pharmacologic approaches. In vitro cell growth, proliferation, and differentiation were assessed using colorimetric MTT and BrdU assays, and biochemical/morphologic changes after incubation in differentiation-promoting media, respectively. In vivo tumorigenesis was assessed using xenograft formation in SCID/beige mice. RESULTS: Notch signaling is upregulated in human RMS cell lines and tumors compared with primary skeletal muscle, especially in the embryonal (eRMS) subtype. Inhibition of Notch signaling using Notch1 RNAi or γ-secretase inhibitors reduced eRMS cell proliferation in vitro. Hey1 RNAi phenocopied Notch1 loss and permitted modest myogenic differentiation, while overexpression of an activated Notch moiety, ICN1, promoted eRMS cell proliferation and rescued pharmacologic inhibition. Finally, Notch inhibition using RNAi or γ-secretase inhibitors blocked tumorigenesis in vivo. CONCLUSIONS: Aberrant Notch-Hey1 signaling contributes to eRMS by impeding differentiation and promoting proliferation. The efficacy of Notch pathway inhibition in vivo supports the development of Notch-Hey1 axis inhibitors in the treatment of eRMS.

Epidermal growth factor potentiates renal cell death in hydronephrotic neonatal mice, but cell survival in rats.

BACKGROUND: Epidermal growth factor (EGF) markedly attenuates tubular apoptosis induced by unilateral ureteral obstruction (UUO) in the neonatal rat, and reduces apoptosis induced by mechanical stretch of cultured rat tubular cells. METHODS: To investigate the role of EGF in modulating apoptosis resulting from UUO, neonatal wild type and mutant mice lacking EGF (knockout), or with diminished EGF receptor activity (waved-2 mutant) were compared to control mice for tubular apoptosis and atrophy. Rat and mouse kidneys were compared for localization of the EGF receptor. Apoptosis was also measured in cultured mouse tubular cells subjected to stretch and exposed to EGF. RESULTS: UUO reduced endogenous renal EGF expression in wild-type mice. Unlike the rat, exogenous EGF did not decrease tubular apoptosis or atrophy in the obstructed kidney, and significantly increased stretch-induced apoptosis of cultured mouse tubular cells. Tubular apoptosis was 50% lower in the obstructed kidney of EGF knockout and waved-2 mice relative to wild type and heterozygous animals. Exogenous EGF increased tubular apoptosis and doubled atrophy in the obstructed kidney of waved-2 mice. Species differences in EGF receptor localization were detected in 3-day-old kidneys. CONCLUSION: EGF acts as a survival factor in the neonatal rat, but potentiates tubular cell death in the neonatal mouse. Species differences are maintained in cultured cells, suggesting that differences in EGF receptor signaling underlie these opposing effects.