Biology subjects

Podini, P.

Publications and source records attributed to Podini, P..

8 recordsLinked to original sources

bioRxiv2025

Early endothelial activation at the blood-nerve barrier defines a hallmark of ALS

Vascular defects are common in Amyotrophic Lateral Sclerosis (ALS). The prevailing view is that breakdown of the blood-brain and blood-spinal cord barriers contributes to neurodegeneration. Here, we reveal that selective and early vulnerability of peripheral nerve endothelium --manifested as endothelial cell activation and interlinked blood-nerve barrier dysfunction-- constitutes a core feature of ALS pathogenesis, arising before vascular alterations in the central nervous system (CNS) and motor neuron pathology. Vascular changes in ALS patients have been largely studied in postmortem samples, limiting insight into their onset, causes, and pathogenic role. Surveying diagnostic motor nerve samples from ALS patients across clinical stages, we observed endothelial damage that preceded axonal loss and demyelination, marking vascular dysfunction as an early disease event. Similar ultrastructural abnormalities were detected in pre-symptomatic ALS mouse models (SOD1 and TARDBP mutants). Notably, endothelial cells became dysfunctional even when not carrying mutant TDP-43, indicating they respond to non-cell autonomous disease signals. Transcriptional, histological, and functional analyses revealed that these alterations were largely confined to peripheral nerves, while spinal cord vessels exhibited delayed and more focal changes. Single-cell sequencing identified ALS-susceptible endothelial cell subsets prone to a pro-inflammatory phenotype and impaired blood-nerve barrier function, increasing permeability via the transcellular route. These changes coincided with reactivity of nerve-resident macrophages and neutrophil infiltration. Neutrophil depletion attenuated endothelial activation and barrier leakiness, mitigating axonopathy in ALS mice. Our work unmasks the greater susceptibility of the peripheral nerve vasculature in ALS relative to the CNS. The early activation of peripheral nerve endothelium, combined with its potential reversibility, identifies a therapeutic window and suggests strategies for targeting the vascular-immune axis to protect the motor system.

neuroscience↗

bioRxiv2024

Glia-enriched stem-cell 3D model of the human brain mimics the glial-immune neurodegenerative phenotypes of multiple sclerosis

The role of central nervous system (CNS) glia in sustaining self-autonomous inflammation and driving clinical progression in multiple sclerosis (MS) is gaining scientific interest. We applied a single transcription factor (SOX10)-based protocol to accelerate oligodendrocyte differentiation from hiPSC-derived neural precursor cells, generating self-organizing forebrain organoids. These organoids include neurons, astrocytes, oligodendroglia, and hiPSC-derived microglia to achieve immunocompetence. Over 8 weeks, organoids reproducibly generated mature CNS cell types, exhibiting single-cell transcriptional profiles similar to the adult human brain. Exposed to inflamed cerebrospinal fluid (CSF) from MS patients, organoids properly mimic macroglia-microglia neuro-degenerative phenotypes and intercellular communication seen in chronic active MS. Oligodendrocyte vulnerability emerged by day 6 post-MS-CSF exposure, with nearly 50% reduction. Temporally-resolved organoid data support and expand on the role of soluble CSF mediators in sustaining downstream events leading to oligodendrocyte death and inflammatory neurodegeneration. Such findings support implementing this organoid model for drug screening to halt inflammatory neurodegeneration.

neuroscience↗

bioRxiv2024

Serum mitochondrial bioenergetics as fingerprint of the hepatic one: how to monitor genetic MASLD

Background & AimsMetabolic Dysfunction-Associated Steatotic Liver Disease (MASLD) pathogenesis is shaped by genetics and mitochondrial dysfunction. Recently, we demonstrated that the co-presence of patatin- like phospholipase domain-containing 3 (PNPLA3), transmembrane 6 superfamily member 2 (TM6SF2) and membrane bound o-acyltransferase domain-containing 7 (MBOAT7) polymorphisms predisposes to disease progression in MASLD patients and that their deletion contributes to mitochondrial (mt) maladaptation in an in- vitro model. In this work we deepened the impact of genes silencing on mitochondrial dynamism. Then we restored TM6SF2 and/or MBOAT7 wild-type (WT) proteins in the in-vitro model to evaluate the rescue of organelles morphology/function. Finally, we compared hepatic and peripheral mt-bioenrgetics in MASLD patients carrying PNPLA3, MBOAT7 and/or TM6SF2 loss-of-function variations. MethodsWT proteins were overexpressed through lentiviral transfection, mt-respiration was assessed by Seahorse. ResultsThe restore of MBOAT7 and/or TM6SF2 wild-type proteins resulted in the assembly of spaghetti- shaped mitochondria with improved OXPHOS capacity. Mitochondrial activity was assessed in liver biopsies and peripheral blood mononuclear cells of biopsy-proven (n=44;Discovery cohort) and noninvasively assessed (n=45;Fibroscan-MASLD cohort) MASLD patients stratified according to the presence of the 3 at-risk variants alongside in unrelated liver disease patients (n=45;Unrelated liver disease cohort). In the Discovery cohort, the hepatic bioenergetic profile fully reflecting the circulating one, was impaired in carriers of the risk variants, more so when in combination. We confirmed the lowered serum respirometry in the Fibroscan-MASLD cohort. Finally, the circulating respiration did not change in unrelated liver disease patients, thus demonstrating that it was specifically impaired in MASLD. ConclusionsThese results boosted the relevance of mitochondrial circulating respirometry to outline genetically-based MASLD.

genetics↗

bioRxiv2024

Prostaglandin D2 synthase controls Schwann cells metabolism

We previously reported that in the absence of Prostaglandin D2 synthase (L-PGDS) peripheral nerves are hypomyelinated in development and that with aging they present aberrant myelin sheaths. We now demonstrate that L-PGDS expressed in Schwann cells is part of a coordinated program aiming at preserving myelin integrity. In vivo and in vitro lipidomic, metabolomic and transcriptomic analyses confirmed that myelin lipids composition, Schwann cells energetic metabolism and key enzymes controlling these processes are altered in the absence of L-PGDS. Moreover, Schwann cells undergo a metabolic rewiring and turn to acetate as the main energetic source. Further, they produce ketone bodies to ensure glial cell and neuronal survival. Importantly, we demonstrate that all these changes correlate with morphological myelin alterations and describe the first physiological pathway implicated in preserving PNS myelin. Collectively, we posit that myelin lipids serve as a reservoir to provide ketone bodies, which together with acetate represent the adaptive substrates Schwann cells can rely on to sustain the axo-glial unit and preserve the integrity of the PNS.

neuroscience↗

bioRxiv2024

NEK1 haploinsufficiency impairs ciliogenesis in human iPSC-derived motoneurons and brain organoids

Primary cilia are microtubule-based organelles acting as specialized signalling antennae that respond to specific stimuli to maintain cellular integrity and homeostasis. Recent studies indicate defective primary cilia in post-mortem human brains and animal models of neurodegenerative conditions, including Amyotrophic Lateral Sclerosis (ALS). Heterozygous loss-of-function mutations (LOF) in NEK1 gene are present in about 1% of familial and sporadic ALS cases. The protein kinase NEK1 regulates various cellular processes, including ciliogenesis, but a clear link between NEK1 LOF mutation in ALS and primary cilia is unknown. In this study we generated a human iPSC line carrying a NEK1 LOF mutation by gene editing, leading to NEK1 protein haploinsufficiency. In differentiated iPSC-motoneurons (MNs) we observed that primary cilia were significantly shorter in NEK1-LOF iPSC-MNs compared to wild-type (WT) iPSC-MNs and that also the percentage of ciliated iPSC-MNs was significantly decreased in NEK1-LOF cells. We also investigated ciliogenesis in NEK1-LOF iPSC-brain organoids confirming that primary cilia were thinner with no apparent alteration in the ultrastructure by transmission electron microscopy. Our data suggest that NEK1 protein plays a role in regulating ciliogenesis in both 2D and 3D human iPSC-derived neuronal models and that NEK1 LOF mutations associated to ALS, leading to NEK1 haploinsufficiency and likely to reduced kinase activity, impair primary cilium formation. The involvement of ciliogenesis dysfunction in ALS deserves further investigation providing novel therapeutic targets and strategies to be addressed for this incurable disease.

neuroscience↗

bioRxiv2023

Two neuronal models of TDP-43 proteinopathy display reduced axonal translation, increased oxidative stress, and defective exocytosis.

Amyotrophic lateral sclerosis (ALS) is a progressive, lethal neurodegenerative disease mostly affecting people around 50-60 years of age. TDP-43, a ubiquitously expressed RNA-binding protein involved in pre-mRNA splicing and controlling mRNA stability and translation, forms neuronal cytoplasmic inclusions in an overwhelming majority of ALS patients, of both sporadic and familial origin, a phenomenon referred to as TDP-43 proteinopathy. These cytoplasmic aggregates disrupt the subcellular transport and localization of mRNA. The axon, like dendrites, is a site of mRNA translation, permitting the local synthesis of selected proteins, both constitutively and in response to stimuli reaching the axon and presynaptic terminal. This is especially relevant in upper and lower motor neurons, whose axon spans long distances, likely accentuating their susceptibility to ALS-related noxae. In this work we have generated and characterized two models of TDP-43 proteinopathy, consisting of virtually pure populations of mouse cortical neurons expressing a human TDP-43 fusion protein, wt or mutant, which accumulates as cytoplasmic aggregates. Neurons expressing human TDP-43 exhibit a global impairment in axonal protein synthesis, an increase in oxidative stress, and defects in presynaptic function and electrical activity. These changes correlate with deregulation in the axonal levels of polysome-engaged mRNAs playing relevant roles in those processes. Our data support the emerging notion that deregulation of mRNA metabolism and of axonal mRNA transport may trigger the dying-back neuropathy that initiates motor neuron degeneration in ALS.

neuroscience↗

bioRxiv2021

Heparin protects human neural progenitor cells from Zika Virus-induced cell death and preserves their differentiation into mature neural-glia cells

Zika virus (ZIKV) is an arbovirus member of the Flaviviridae family that causes severe congenital brain anomalies in infected fetuses. Human neural progenitor cells (hNPCs) are highly permissive to ZIKV infection, causing inhibition of cell proliferation concomitant with an induction of cell death. We previously demonstrated that pharmaceutical-grade heparin inhibited virus-induced cell death with minor effects on in vitro virus replication in ZIKV-infected hNPCs. Here we show that heparin prevented ZIKV-induced intracellular vacuoles, a signature characteristic of paraptosis, but also inhibited necrosis and apoptosis of hNPCs when grown as neurospheres (NS). Furthermore, heparin preserved the differentiation of both ZIKV-infected human-induced pluripotent stem cells (hiPSC) derived-NPCs and fetus-derived NPCs into neural-glial cells. Collectively, these results highlight the potential neuroprotective effect of heparin that could be re-purposed and exploited to drive the development of novel agents for preventing ZIKV damage.

microbiology↗

bioRxiv2020

ZFP423 regulates early patterning and multiciliogenesis in the hindbrain choroid plexus

The choroid plexus (ChP) is a secretory tissue that produces cerebrospinal fluid (CSF) and secretes it into the ventricular system. CSF flows from the lateral to the third ventricle, and then to the fourth ventricle through the cerebral aqueduct. Recent studies have uncovered new, active roles for this structure in the regulation of neural stem cell maintenance and differentiation into neurons. Zfp423, encoding a Kruppel-type zinc finger transcription factor essential for cerebellar development and mutated in rare cases of cerebellar vermis hypoplasia / Joubert syndrome and other ciliopathies, is expressed in the hindbrain roof plate (RP), from which the IV ventricle ChP arises, and in mesenchymal cells giving rise to the stroma and leptomeninges. Zfp423 mutants display a marked reduction of the hindbrain ChP (hChP), which fails to express key markers of its secretory function and genes implicated in its development and maintenance (Lmx1a, Otx2). The mutant hChP displays a complete lack of multiciliated ependymal cells. A transcriptome analysis conducted at the earliest stages of hChP development and subsequent validations demonstrate that the mutant hChp displays a strong deregulation of pathways involved in early hindbrain patterning and multiciliated cell fate specification. Our results propose Zfp423 as a master gene and one of the earliest known determinants of hChP development.

developmental biology↗