Biology subjects

Rocha, S. F.

Publications and source records attributed to Rocha, S. F..

5 recordsLinked to original sources

bioRxiv2026

Endothelial β3-Adrenergic Receptor activation prevents pulmonary hypertension

BackgroundPulmonary hypertension (PH) is a progressive vascular disease characterized by endothelial dysfunction, vascular remodeling and increased pulmonary vascular resistance. The {beta}3-adrenergic receptor ({beta}3-AR) has been implicated in cardiovascular regulation and cardioprotective mechanisms; however, its role in pulmonary vascular disease remains poorly understood. We investigated whether activation of {beta}3-AR protects pulmonary endothelial function and prevents the development of pre-capillary PH. Methods{beta}3-AR expression was evaluated in pulmonary endothelium from patients with Chronic Obstructive Pulmonary Disease (COPD) and in murine models of hypoxia-induced PH. Genetic mouse models including {beta}3-AR knockout (KO) and conditional {beta}3-AR overexpression in endothelial cells (EC) or in smooth muscle cells (SMC), were used to determine cell-specific roles. Pharmacological activation of {beta}3-AR was achieved using the selective {beta}3-agonist mirabegron in hypoxia-induced PH mice and monocrotaline-induced PH rats. Pulmonary vascular reactivity and vasodilatory responses to {beta}3-AR stimulation were evaluated by wire myography in isolated pulmonary arteries. Mechanistic studies were performed in human pulmonary artery endothelial cells (HPAEC) under hypoxic conditions, in human pulmonary arterial smooth muscle cells (HPASMC) and in endothelial nitric oxide synthase (NOS3) KO mice. Results{beta}3-AR was upregulated in pulmonary endothelium of COPD patients and mice exposed to chronic hypoxia. Genetic deletion of {beta}3-AR aggravated PH, whereas endothelial-specific overexpression attenuated the disease phenotype, reducing right ventricular systolic pressure (RVSP), vascular remodeling and right ventricular (RV) hypertrophy. Activation of {beta}3-AR with mirabegron improved pulmonary hemodynamics, reduced vascular remodeling and preserved RV function. {beta}3-AR activation promoted endothelial nitric oxide synthase (eNOS)-dependent NO production, indirectly inhibiting SMC proliferation. Additionally, {beta}3-AR activation improved mitochondrial fitness in endothelial cells by increasing uncoupling protein 2 (UCP2) expression, reducing reactive oxygen species (ROS) generation and preventing mitochondrial fragmentation. ConclusionsThese findings identify endothelial {beta}3-AR as a previously unrecognized regulator of pulmonary vascular homeostasis and provide a strong translational rationale for targeting the {beta}3-adrenergic pathway in PH. Given that mirabegron is already approved for clinical use, our results support its repurposing as a therapeutic strategy for pre-capillary forms of PH.

pharmacology and toxicology↗

bioRxiv2025

Disrupted astrocyte-endothelial crosstalk drives hemangioblastoma lesions in VHL disease

Hemangioblastomas (HBs) are highly vascularized central nervous system (CNS) tumours that can become life-threatening, especially in the context of Von Hippel-Lindau (VHL) disease, caused by the loss of VHL function. The limited pharmacological options targeting VHL-HBs stem from an incomplete understanding of their cellular origin, development, and molecular pathogenesis. Here we use advanced mouse genetics to show that mosaic deletion of Vhl in Apln+ cells leads to the formation of precursor tumour-like lesions, composed by clusters of Vhl-knockout (VhlKO) astrocytes and surrounding Vhl-wild-type (VhlWT) vessels that become malformed, resembling early-stage HBs linked to VHL disease. VhlKO astrocytes morphologically and transcriptomically resembled the reactive astrocytes characteristic of ischemic CNS injury. They exhibited metabolic rewirement towards glycolysis and upregulation of cell growth pathways. They also expressed several secreted proangiogenic molecules that activate and prevent the normal maturation of neighbouring vessels, leading to VHL-HBs. Temporal conditional genetic analysis revealed that Vhl loss need to happen during postnatal development for HBs to form, and that lesions become quiescent in early adulthood. HIF-2 deletion, or MTORC1 inhibition with rapamycin, efficiently inhibited VHL-HBs growth and the associated vascular malformations. Our work shows that the loss of Vhl in single astrocytes induces their growth and pathogenic crosstalk with neighbouring endothelial cells, driving hemangioblastoma development in VHL disease. Our new somatic mosaic mouse models will also enable testing of novel drugs against this disease.

cancer biology↗

bioRxiv2025

Genetic mosaics reveal mechanisms of resistance to VEGF signaling loss during angiogenesis

The VEGF ligand and its main receptor VEGFR2 are considered to be essential for endothelial differentiation, proliferation, sprouting and survival. Blood vessels cannot form in growing embryos or tissues when VEGF/VEGFR2 signalling is compromised in all cells. The Anti-VEGF blocking antibody is one of the most widely used antibodies in the clinics, blocking angiogenesis in cancer, wound healing or in ischemic diseases. However, vascular resistance to Anti-VEGF has been reported. Here we used iFlpMosaics and iSuRe-HadCre to induce and track genetic mosaics of endothelial cells (ECs) lacking VEGFR2 during the entire embryonic and postnatal development. Surprisingly, Vegfr2KOECs adapt, proliferate normally and compete with wild-type cells over time, ultimately forming a substantial portion of the capillary network in most organs. We found that Vegfr2KO ECs are not able to sprout during the first wave of angiogenesis in most tissues, because it is highly VEGF-dependent and favours the growth and mobilization of wild-type ECs. However, due to their inability to respond to VEGF, over time Vegfr2KOECs become dominant in veins, which provide for a long term and continuous source of ECs for subsequent waves of VEGF-independent angiogenesis. Comparative scRNAseq analysis of ECs with acute and long-term loss of VEGFR2, revealed both common and organ-specific molecular mechanisms of adaptation and resistance to VEGF signalling loss. This included significant endothelial venousization and the upregulation of ligands for VEGFR1 and VEGFR3. VEGFR1 only partially compensated for VEGFR2 loss in capillary ECs, whereas VEGFR3 only compensated in arterial ECs. Loss of the three VEGF receptors did not compromise venous growth. This work changes our understanding of the role of VEGF signaling and its receptors in angiogenesis and reveals mechanisms of adaptation and resistance to their loss.

developmental biology↗

bioRxiv2024

Vascular HIF2 signaling prevents cardiomegaly, alveolar congestion and capillary remodeling during chronic hypoxia

Hypoxia is associated with the onset of cardiovascular diseases including cardiac hypertrophy and pulmonary arterial hypertension (PAH). Endothelial HIF2 signaling mediates pulmonary arterial remodeling and subsequent right ventricular systolic pressure (RVSP) elevation during chronic hypoxia, encouraging novel therapeutic opportunities for PAH based on specific HIF2 inhibitors. Nevertheless, HIF2 relevance beyond the pulmonary endothelium or in the cardiac adaptation to hypoxia remains elusive. Wilms tumor 1 lineage contributes to heart and lung vascular compartments including pericytes, endothelial and smooth muscle cells. Here we describe the response to chronic hypoxia of a novel HIF2 mutant mouse model in the Wt1 lineage (Hif2/Wt1 cKO). Hif2/Wt1 cKO is protected against pulmonary remodeling and increased RVSP induced by hypoxia, but displays alveolar congestion, inflammation and hemorrhages associated with microvascular instability. Furthermore, lack of HIF2 in the Wt1 lineage leads to cardiomegaly, capillary remodeling, right and left ventricular hypertrophy, systolic dysfunction and left ventricular dilation, suggesting pulmonary-independent cardiac direct roles of HIF2 in hypoxia. These structural defects are partially restored upon reoxygenation, while functional parameters remain altered. Our results suggest that cardiopulmonary HIF2 signaling prevents excessive vascular proliferation during chronic hypoxia and define novel protective roles of HIF2 to warrant stable microvasculature and organ function.

pathology↗

bioRxiv2023

iFlpMosaics: A method for the ratiometric induction and high-throughput comparative analysis of mutant and wildtype cells

To understand gene function, it is necessary to compare cells carrying the mutated target gene with normal cells. In most biomedical studies, the cells being compared are in different mutant and control animals and therefore do not experience the same epigenetic changes and tissue microenvironment. The experimental induction of genetic mosaics is essential to determine a gene cell-autonomous function and to model the etiology of diseases caused by somatic mutations. Current technologies used to induce genetic mosaics in mice lack either accuracy, throughput or barcoding diversity. Here, we present a large set of new genetic tools and mouse lines that enable Flp recombinase-dependent ratiometric induction and single-cell clonal tracking of multiple fluorescently labeled wildtype and Cre-mutant cells within the same time window and tissue microenvironment. The labeled cells can be profiled by multispectral imaging or by FACS and scRNA-seq. This technology facilitates the induction and analysis of genetic mosaics in any cell type and for any given single or combination of floxed genes. iFlpMosaics enables a more accurate understanding of how induced genetic mutations affect the biology of single cells during tissue development, homeostasis, and disease.

cell biology↗