Biology subjects

Caliandro, R.

Publications and source records attributed to Caliandro, R..

5 recordsLinked to original sources

bioRxiv2026

Hierarchical and context-dependent GR-MR signalling governs endogenous corticosteroid decoding in the heart

Endogenous corticosteroids bind both the glucocorticoid receptor (GR) and mineralocorticoid receptor (MR), yet how this shared ligand input is decoded in the heart remains unclear. Previous work established that corticosterone suppresses postnatal cardiomyocyte proliferation through GR. Here we show that corticosteroid responses are governed by a functional GR-MR hierarchy and by cellular context. Genetic or pharmacological GR inhibition redirects corticosterone towards cardiomyocyte proliferation, and MR antagonism or silencing abolishes this effect. GR disruption also enhances aldosterone-induced proliferation in cardiomyocyte-enriched cultures, indicating that GR restrains MR output beyond ligand allocation. However, aldosterone fails to increase cardiomyocyte proliferation in mixed cultures and instead stimulates fibroblast proliferation and activation. By contrast, corticosterone combined with GR inhibition promotes cardiomyocyte proliferation without inducing stromal proliferation or profibrotic activation. Both treatments induce MR nuclear localisation in fibroblasts, showing that their divergent stromal effects arise despite comparable receptor nuclear engagement. Following myocardial infarction, circulating corticosterone increases and GR inhibition enhances cardiomyocyte MR nuclear localisation. In adult murine myocardium, corticosterone plus GR antagonism increases cardiomyocyte cell-cycle activity in an MR-dependent manner, with analogous responses observed in porcine and human myocardium. These findings identify hierarchical and context-dependent GR-MR signalling as a mechanism of corticosteroid decoding and a potential route to cardiac regeneration.

molecular biology↗

bioRxiv2026

TBX5 dosage governs ventricular cardiomyocyte maturation, specialization and dedifferentiation in vivo

Variation in transcription factor (TF) activity modulates traits and disease susceptibility, yet how such variation translates into cellular phenotype and organ function in vivo is not well established. We utilized AAV-mediated gene delivery to express the dosage-sensitive TF TBX5 across a physiologically plausible range in postnatal ventricular cardiomyocytes. Transcriptomic profiling revealed that TBX5 dosage-dependent cardiomyocyte states changed gradually and often non-monotonically across the TBX5 dosage spectrum. The lowest dosages induced cardiomyocyte hypertrophy and upregulated gene programs governing oxidative metabolism, calcium handling, and contractility. In contrast, mid-to-high dosages induced a ventricular conduction system-like transcriptional profile. Supraphysiological dosages triggered cardiomyocyte dedifferentiation, characterized by cardiomyocyte size reduction, cell cycle re-entry, metabolic reprogramming, and impaired ventricular ejection fraction. Furthermore, the ventricular state of an Nppa-Nppb deficiency model characterized by cardiac hypertrophy and reduced Tbx5 expression was partially normalized by TBX5 delivery. By defining the non-linear relationship of TBX5 activity level, cardiomyocyte state and cardiac function, our study reveals how TF dosage regulates the transitions from physiological maturation to specialized lineage acquisition and dedifferentiation at the cell and organ level in vivo.

developmental biology↗

bioRxiv2026

Advancing Nuclei Isolation from Frozen Human Heart for Single-Nucleus RNA Sequencing Applications

While single-cell RNA sequencing (scRNA-seq) has been the first widely adopted single-cell transcriptomic approach, its reliance on fresh tissue samples has substantially limited its applicability to clinically relevant specimen. Single-nucleus RNA sequencing (snRNA-seq) overcomes this constrain by enabling transcriptomic profiling from frozen material. However, isolating high-quality nuclei from frozen cardiac tissue remains technically challenging due to the dense extracellular matrix, complex tissue architecture, and heterogeneous cellular composition of the heart. To address these challenges, numerous nuclei isolation protocols have been adapted and optimized, resulting in substantial methodological heterogeneity across studies. Despite the widespread use of snRNA-seq in cardiac research, a robust and standardized nuclei isolation protocol that consistently yields high-quality nuclei from frozen human heart tissue is still lacking. Here, we present a comprehensive, end-to-end protocol for nuclei isolation from frozen human left ventricle, along with a detailed downstream pipeline for snRNA-seq data analysis. Our hybrid nuclei isolation strategy integrates multiple sequential clean-up steps designed to preserve nuclear integrity and RNA quality prior to sequencing. Compared with commonly used nuclei isolation protocols, this approach yields substantially higher number of nuclei while maintaining comparable numbers of detected genes and counts, even at lower sequencing depth. Adoption of this protocol may reduce technical variability across studies and facilitate more reproducible snRNA-seq analyses of human cardiac tissue.

molecular biology↗

bioRxiv2024

A novel and robust method for assessing mitochondrial (dys)function in healthy and diseased frozen cardiac tissue

Cardiovascular diseases are often associated with impairment in mitochondrial function detected by reduced mitochondrial oxygen consumption using high-resolution respirometry. However, existing respirometry protocols are limited by the necessity for fresh tissue samples. This study developed a method with tailored substrate-inhibitor titration (TSIT) of mitochondrial electron transport complexes (ETC) to measure mitochondrial function in frozen cardiac samples using high-resolution respirometry. Briefly, acetyl-CoA was added to fuel the tricarboxylic acid (TCA) cycle for NADH production, enabling complex I (CI)-linked respiratory assessment. NADH was then added to measure maximum CI-linked respiratory capacity, followed by rotenone and succinate to assess complex II (CII)-linked respiratory capacity. TSIT detected mitochondrial functional differences between frozen atrial and ventricular tissue, with comparable results as measured in fresh samples. It also detected cardiac mitochondrial dysfunction across various (patho)physiological mouse models (including aging, ischemia reperfusion, obesity, and CI deficiency) as well as in frozen human donor samples, highlighting its clinical potential. Furthermore, we showed the first evidence for supercomplexes (SCs) formation between ETC-SCs and the TCA cycle metabolon, underpinning TSIT feasibility. In conclusion, we established a novel, robust, sensitive and translational method (TSIT) for assessing mitochondrial (dys)function in frozen cardiac samples from various species, enabling flexible analysis of mitochondrial function in both laboratory and clinical settings.

physiology↗

bioRxiv2023

Vascular abnormalities in heart and brain are associated with cardiovascular and neurological symptoms in a novel mouse model for Williams syndrome

Williams syndrome is a developmental disorder caused by a microdeletion entailing the loss of a single copy of 25-27 genes on chromosome 7q11.23. Patients suffer from cardiovascular and neuropsychological symptoms. Structural abnormalities of the cardiovascular system in Williams syndrome have been attributed to the hemizygous loss of the elastin (ELN) gene. In contrast, the neuropsychological consequences of Williams syndrome, including sensorimotor deficits, hypersociability and cognitive impairments, have been mainly attributed to altered expression of transcription factors like LIMK1, GTF2I and GTF2IRD1, while the potential impact of altered cerebrovascular function has been largely overlooked. To study the relationship between Williams syndrome mutations and vascularization of both the heart and brain, we generated a mouse model carrying a relatively long microdeletion (LD) that includes the Ncf1 gene, thereby minimizing the confounding impact of hypertension. LD mice had elongated and tortuous aortas but, unlike Eln haploinsufficient mice, showed no signs of structural cardiac hypertrophy. Remarkably, LD mice also displayed structural abnormalities in coronary and brain vessels, including disorganized extracellular matrices. Importantly, LD mice faithfully replicated both cardiovascular and neuropsychological symptoms observed in patients. The phenotype was even more comprehensive than former models, with structure-function correlations evident in aberrant auditory and motor behaviors resembling those in patients with Williams syndrome. Together, our findings suggest that not only cardiovascular but also neuropsychological symptoms in Williams syndrome may be driven in part by vascular abnormalities affecting both heart and brain. Significance StatementWilliams syndrome is caused by microdeletion of 25-27 genes on chromosome 7q11.23, resulting in cardiovascular and neuropsychological symptoms. It remains unclear how the affected genes interact and whether cardiovascular deficits influence brain function. We developed and characterized a mouse model with the longest Williams syndrome microdeletion to date. This model reveals interactions between genes that can be compensatory or additive: haploinsufficiency of Ncf1 may counteract the cardiac hypertrophy caused by Eln deletion, while vascular defects that are potentially due to Eln haploinsufficiency extend to the brain and may worsen neuropsychological symptoms. Our findings support the hypothesis that structural vascular deficits putatively contribute to both cardiac and cognitive phenotypes in Williams syndrome, opening new avenues for understanding and treating this syndrome.

systems biology↗