Biology subjects

Sequeira-Lopez, M. L. S.

Publications and source records attributed to Sequeira-Lopez, M. L. S..

8 recordsLinked to original sources

bioRxiv2026

NetTracer3D Enables User-Friendly Analysis of Diverse Microscopic and Medical 3D Datasets

Recent advances in high-resolution imaging and spatial transcriptomics have enabled reconstruction of complex 3D tissue maps, providing unprecedented insights into cellular connectivity, organization, and tissue architecture. However, standardization challenges hinder integration, sharing, and analysis of these datasets across research communities. We developed NetTracer3D to simplify three-dimensional image analysis across diverse datasets. NetTracer3D is an integrated tool for defining, processing, and sharing 3D tissue maps with standardized data formats and interactive exploration capabilities. It provides three broadly applicable network analysis modalities: Connectivity networks for analyzing functional tissue units or cells connected via secondary structures such as nerves or vasculature; Branch Adjacency and Branchpoint networks for converting branched anatomical structures into analyzable representations; and Proximity networks for grouping structures by spatial relationships to identify cellular organization patterns. We demonstrate several use cases applying NetTracer3D to analyze multidimensional data from CODEX and label free Raman spectroscopy, multiscalar data encompassing subcellular and anatomical scales and a range of modalities. NetTracer3D was able to characterize neural relationships between functional tissue units in human and mouse kidneys and mouse bronchi. Branchpoint networks were used to identify vascular defects in human brain angiogram and define the innervation structure of a lymph node. Finally, we demonstrate how proximity networks characterize the tumor microenvironment in 3D light sheet cancer images and auto-detect cellular neighborhoods in multiplexed 2D CODEX datasets. Beyond network creation, NetTracer3D enables analysis, spatial statistics, and visual analytics tailored for volumetric tissue data. By establishing interoperable formats and analysis workflows, this work provides accessible and reproducible analytical tools for 3D spatial biology, enabling new discoveries of relationships between structure and physiology.

genomics↗

bioRxiv2025

Hypoxia-Induced Metabolic Reprogramming and Markings of Cell Fate in Concentric Arterial Hypertrophy

Chronic inhibition of the renin-angiotensin system (RAS), while widely used to treat hypertension, can lead to an underrecognized form of vascular disease marked by concentric arteriolar and arterial hypertrophy (CAAH). Here, using two lineage-traced mouse models of genetic renin deletion and sustained RAS blockade, we uncover a pathogenic cascade initiated by renin-lineage cell fate reprogramming. Loss of endocrine identity and transformation of smooth muscle cells drives a shift toward a fibrotic, inflammatory, and secretory phenotype that remodels the extracellular matrix and promotes vascular thickening and luminal narrowing. Integrated transcriptomic, proteomic, and metabolomic profiling revealed a hypoxia-linked metabolic switch--characterized by succinate accumulation and NAD+ depletion--coupled to Hif activation and disease progression. We identify Cdh13 and collagens (including Col1a1 and Col12a1) as early urinary biomarkers and define a 10-gene molecular signature of CAAH with potential clinical application. These findings establish renin-lineage cell plasticity and metabolic dysfunction as central drivers of CAAH and nominate candidate biomarkers for early detection and therapeutic targeting in RAS-inhibited patients.

physiology↗

bioRxiv2025

Renin Cells Drive Kidney Neurovascular Development and Arterial Remodeling when Renin Activity is Deficient

Renin cells synthesize and release the hormone-enzyme renin to regulate blood pressure and fluid-electrolyte homeostasis. Their function and identity depend on communication with surrounding cells and nerve fibers within complex kidney structure. Because renin cells are rare -0.01 % of kidney cells-conventional histological approaches cannot capture their interaction with nerve fibers and surrounding cells within the nephron and its vasculature. Using a novel ultrabright renin cell-specific tdTomato reporter mouse, high-resolution 3D imaging, and single-cell RNA-Seq, we mapped the interactions of renin cells with growing axons during normal kidney vascular development, in response to threats to homeostasis, and a severe arterial disease caused by a defective renin enzyme. During embryonic kidney development, stromal and renin cell progenitors assemble the arterioles, express axon attractants and neurotrophins that establish the precise innervation of renin cells and arterioles in a centrifugal pattern. Hypotension and sodium depletion led to an increase in the volume and number of renin cells along the arterioles. Renin enzymatic deficiency led to hypertrophy and endocrine transformation of renal arterioles, aberrant axon sprouting and sympathetic hyperinnervation suggesting a feed-forward mechanism whereby renin cells and axons co-induce each other, orchestrate neurovascular development and arteriolar remodeling when renin cells are over stimulated.

developmental biology↗

bioRxiv2024

Angiotensin II elicits robust calcium oscillations coordinated within juxtaglomerular cell clusters to suppress renin secretion.

BACKGROUNDJuxtaglomerular (JG) cells are sensors that control blood pressure (BP) and fluid-electrolyte homeostasis. They are arranged as clusters at the tip of each afferent arteriole. In response to a decrease in BP or extracellular fluid volume, JG cells secrete renin, initiating an enzymatic cascade that culminates in the production of angiotensin II (AngII), a potent vasoconstrictor that restores BP and fluid-electrolyte homeostasis. In turn, AngII exerts negative feedback on renin release concomitantly with increased intracellular Ca2+, preventing excessive circulating renin and hypertension. However, within their native structural organization, the intricacies of intracellular Ca2+ signaling dynamics and their sources remain uncharacterized. METHODSWe generated mice expressing the JG cell-specific genetically encoded Ca2+ indicator (GCaMP6f) to investigate Ca2+ dynamics within JG cell clusters ex vivo and in vivo. For ex vivo Ca2+ imaging, acutely prepared kidney slices were perfused continuously with a buffer containing variable Ca2+ and AngII concentrations {+/-} Ca2+ channel inhibitors. For in vivo Ca2+ image capture, native mouse kidneys were imaged in situ using multi-photon microscopy with and without AngII administration. ELISA measurements of renin concentrations determined acute renin secretion ex vivo and in vivo, respectively. RESULTSEx vivo Ca2+ imaging revealed that JG cells exhibit robust and coordinated intracellular oscillatory signals with cell-cell propagation following AngII stimulation. AngII dose-dependently induced stereotypical burst patterns characterized by consecutive Ca2+ spikes, which inversely correlated with renin secretion. Pharmacological channel inhibition identified key sources of these oscillations: endoplasmic reticulum Ca2+ storage and release, extracellular Ca2+ uptake via ORAI channels, and intercellular communication through gap junctions. Blocking ORAI channels and gap junctions reduced AngII inhibitory effect on renin secretion. In vivo Ca2+ imaging demonstrated robust intracellular and intercellular Ca2+ oscillations within JG cell clusters under physiological conditions, exhibiting spike patterns consistent with those measured in ex vivo preparations. Administration of AngII enhanced the Ca2+ oscillatory signals and suppressed acute renin secretion in vivo. CONCLUSIONAngII elicits coordinated intracellular and intercellular Ca2+ oscillations within JG cell clusters, ex vivo and in vivo. The effect is driven by endoplasmic reticulum-derived Ca2+ release, ORAI channels, and gap junctions, leading to suppressed renin secretion.

cell biology↗

bioRxiv2024

The transcription factor Tcf21 is required for specifying Foxd1 cells to the juxtaglomerular cell lineage

Renin is crucial for blood pressure regulation and electrolyte balance, and its expressing cells arise from Foxd1+ stromal progenitors. However, factors guiding these progenitors toward renin-secreting cell fate remain unclear. Tcf21, a basic helix-loop-helix (bHLH) transcription factor, is essential in kidney development. Utilizing Foxd1Cre/+;Tcf21f/f and Ren1dCre/+;Tcf21f/f mouse models, we investigated the role of Tcf21 in the differentiation of Foxd1+ progenitor cells into juxtaglomerular (JG) cells. Immunostaining and in-situ hybridization demonstrated fewer renin-positive areas and altered renal arterial morphology, including the afferent arteriole, in Foxd1Cre/+;Tcf21f/f kidneys compared to controls, indicating Tcf21s critical role in the emergence of renin-expressing cells. However, Tcf21 inactivation in renin-expressing cells (Ren1dCre/+;Tcf21f/f) did not recapitulate this phenotype, suggesting Tcf21 is dispensable once renin cell identity is established. Using an integrated analysis of single-cell RNA sequencing (scRNA-seq) and single-cell assay for transposase-accessible chromatin sequencing (scATAC-seq) on GFP+ cells (stromal lineage) from E12, E18, P5, and P30 Foxd1Cre/+;Rosa26mTmG control kidneys, we analyzed the temporal dynamics of Tcf21 expression in cells comprising the JG lineage (n=2,054). A pseudotime trajectory analysis revealed that Tcf21 expression is highest in metanephric mesenchyme and stromal cells at early developmental stages (E12), with a decline in expression as cells mature into renin-expressing JG cells. Motif enrichment analyses supported Tcf21s significant involvement in early kidney development. These findings underscore the critical role of Tcf21 in Foxd1+ cell differentiation into JG cells during early stages of kidney development, offering insights into the molecular mechanisms governing JG cell differentiation and highlight Tcf21s pivotal role in kidney development. NEW & NOTEWORTHYThis manuscript provides novel insights into the role of Tcf21 in the differentiation of Foxd1+ cells into JG cells. Utilizing integrated scRNA-seq and scATAC-seq, the study reveals that Tcf21 expression is crucial during early embryonic stages, with its peak at embryonic day 12. The findings demonstrate that inactivation of Tcf21 leads to fewer renin-positive areas and altered renal arterial morphology, underscoring the importance of Tcf21 in the specification of renin-expressing JG cells and kidney development.

developmental biology↗

bioRxiv2024

An efficient inducible model for the control of gene expression in renin cells

BackgroundFate mapping and genetic manipulation of renin cells have relied on either non-inducible Cre lines that can introduce developmental effects of gene deletion or BAC transgene-based inducible models that may be prone to spurious and/or ectopic gene expression. MethodsWe generated an inducible mouse model in which CreERT2 is under the control of the endogenous Akr1b7 gene, an independent marker of renin cells. ResultsWe evaluated the pattern of Cre expression in Akr1b7CreERT2/+;R26RmTmG/+ mice in which Akr1b7+/renin+ cells become GFP+ upon tamoxifen administration. At E18.5 and P5, GFP was found in Juxtaglomerular cells, along the arterioles, and in the glomerular mesangium. In adult kidneys, GFP was present mainly in Juxtaglomerular cells. In mice treated with captopril and a low sodium diet to induce recruitment of renin cells, GFP extended along the afferent arterioles and in the mesangium. In addition, we deleted renin in adult mice and found a marked reduction in kidney renin expression and mean arterial pressure in mutant animals. When subjected to a homeostatic threat, mutant mice were unable to recruit renin+ cells. Most importantly, mice with renin deletion induced in the adult developed concentric vascular hypertrophy ruling out potential developmental effects on the vasculature due to the lack of renin. ConclusionsRenin can be efficiently and specifically deleted in renin cells in adult life using our conditional model. Akr1b7CreERT2 mice constitute an excellent model for the fate mapping of renin cells and for the spatial and temporal control of gene expression in renin cells.

physiology↗

bioRxiv2023

Protein Kinase A Inhibition Epigenetically Silences Ren1

RationaleRenin-expressing cells are myoendocrine cells crucial for survival which detect changes in blood pressure and release renin to maintain homeostasis. One of the pathways responsible for renin expression includes cAMP as a crucial factor. cAMP binds to subunits of protein kinase A (PKA), ultimately recruiting both CBP and p300. Binding to the cAMP-responsive element in the renin enhancer region thus amplifies renin transcription. ObjectiveTo evaluate transcriptomic and epigenomic changes occurring at the renin locus via cAMP pathway inhibition. Methods and ResultsWe treated As4.1 cells (a tumoral cell line that constitutively expresses renin) with the PKA inhibitor H89 (treated) or DMSO (control). We then performed independent ATAC-seq, scRNA-seq, and ChIP-seq for H3K27Ac and P300 binding on biological replicates of treated and control As4.1 cells. Ren1 expression is significantly reduced following PKA inhibition with a corresponding loss in H3K27Ac and P300 binding at the locus. A restricted set of nine genes with overlapping dynamically accessible regions, differential gene expression, and H3K27Ac and P300 binding were identified with roles among three primary renin regulatory paradigms. ConclusionsThe data suggests that cAMP pathway inhibition controls renin expression through a reduction not in accessibility alone, but via a switch from an active to poised state of epigenetic control, a shift towards a less differentiated cellular identity, and the disruption of not only cAMP, but baroreceptor and Notch mediated renin regulatory pathways.

genomics↗

bioRxiv2023

Determinants of renin cell differentiation: a single cell epi-transcriptomics approach

RationaleRenin cells are essential for survival. They control the morphogenesis of the kidney arterioles, and the composition and volume of our extracellular fluid, arterial blood pressure, tissue perfusion, and oxygen delivery. It is known that renin cells and associated arteriolar cells descend from FoxD1+ progenitor cells, yet renin cells remain challenging to study due in no small part to their rarity within the kidney. As such, the molecular mechanisms underlying the differentiation and maintenance of these cells remain insufficiently understood. ObjectiveWe sought to comprehensively evaluate the chromatin states and transcription factors (TFs) that drive the differentiation of FoxD1+ progenitor cells into those that compose the kidney vasculature with a focus on renin cells. Methods and ResultsWe isolated single nuclei of FoxD1+ progenitor cells and their descendants from FoxD1cre/+;R26R-mTmG mice at embryonic day 12 (E12) (ncells=1234), embryonic day 18 (E18) (ncells=3696), postnatal day 5 (P5) (ncells=1986), and postnatal day 30 (P30) (ncells=1196). Using integrated scRNA-seq and scATAC-seq we established the developmental trajectory that leads to the mosaic of cells that compose the kidney arterioles, and specifically identified the factors that determine the elusive, myo-endocrine adult renin-secreting juxtaglomerular (JG) cell. We confirm the role of Nfix in JG cell development and renin expression, and identified the myocyte enhancer factor-2 (MEF2) family of TFs as putative drivers of JG cell differentiation. ConclusionsWe provide the first developmental trajectory of renin cell differentiation as they become JG cells in a single-cell atlas of kidney vascular open chromatin and highlighted novel factors important for their stage-specific differentiation. This improved understanding of the regulatory landscape of renin expressing JG cells is necessary to better learn the control and function of this rare cell population as overactivation or aberrant activity of the RAS is a key factor in cardiovascular and kidney pathologies.

developmental biology↗