Biology subjects

Chieca, M.

Publications and source records attributed to Chieca, M..

8 recordsLinked to original sources

bioRxiv2026

Schwann cell p75NTR sustains persistent pain downstream to NGF through ROS-dependent TRPA1 signaling

Nerve growth factor (NGF) is a key mediator of pain through activation of the high-affinity tropomyosin receptor kinase A (TrkA) and the low-affinity neurotrophin receptor (p75NTR). Although neuronal TrkA signaling is well established, the contribution of non-neuronal cells to NGF- dependent pain remains unclear. Here, we show that NGF and its precursor proNGF engage distinct cellular mechanisms. Intraplantar NGF induced acute nociception, heat hyperalgesia, mechanical allodynia, and cold hypersensitivity, whereas cleavage-resistant proNGF selectively evoked mechanical allodynia and cold hypersensitivity. Pharmacological and cell-specific genetic approaches demonstrated that acute nociception and heat hyperalgesia require neuronal TrkA, whereas mechanical and cold hypersensitivity depend on p75NTR activation in Schwann cells. In Schwann cells, NGF and proNGF induced p75NTR-dependent calcium release, followed by TRPA1 activation, mitochondrial ROS production, and NOX1-dependent oxidative amplification. Inhibition of ROS or TRPA1, or Schwann cell-specific Trpa1 deletion, markedly reduced mechanical allodynia and cold hypersensitivity without affecting acute nociception or heat hyperalgesia. These findings identify a Schwann cell p75NTR-ROS-TRPA1 pathway sustaining persistent pain and highlight non-neuronal p75NTR signaling as a potential therapeutic target.

neuroscience↗

bioRxiv2026

Targeting Synaptic Vesicle Endocytosis in Nociceptors Provides Sustained Pain Relief

Endocytosis replenishes synaptic vesicle (SV) pools that are required for persistent transmission of chronic pain signals within nociceptive spinal circuits. The nociceptor-specific contribution of SV endocytosis to pain and the therapeutic potential of endocytosis inhibitors are unclear. We identified SV endocytosis in nociceptors as a critical driver of ongoing pain and developed a gene-based strategy to target this mechanism. Nociceptor-specific adeno-associated virus-mediated knockdown of adaptor-associated kinase 1 (AAK1) or dynamin 1 (Dnm1) in dorsal root ganglia Nav1.8-positive neurons inhibited postoperative and neuropathic hypersensitivity without affecting baseline mechanical or thermal sensitivity, locomotion or spontaneous behavior. Electrophysiological recordings from spinal neurons combined with optogenetic activation of nociceptor afferents showed that AAK1 or Dnm1 downregulation blocked the sustained synaptic transmission between nociceptors and dorsal horn neurons by disrupting SV recycling and reducing neurotransmitter release probability. Lipid nanoparticle (LNP)-encapsulated CRISPR/dCas9-repressor mRNA constructs (dCas9-R) were engineered to achieve sustained and reversible transcriptional and epigenetic repression of Aak1 or Dnm1 following intrathecal delivery. LNP-mediated gene modulation produced sustained downregulation of Aak1 or Dnm1 mRNA in sensory neurons and resulted in robust and long-lasting analgesia in preclinical models of postoperative, inflammatory, neuropathic and osteoarthritis pain without impairing acute nociception or locomotor activity. Mechanistically, targeting endocytic machinery disrupted SV recycling at nociceptor terminals, thereby reducing excitatory neurotransmission within spinal pain circuits. Together, these findings establish presynaptic endocytic regulation as a convergent mechanism underlying chronic pain and demonstrate the translational potential of LNP-delivered CRISPR/dCas9-R as a durable, non-opioid pain therapy that surmounts inherent redundancy of pain signaling mechanisms. One Sentence SummarySynaptic vesicle endocytosis in nociceptors is a critical mechanism driving ongoing pain and targeting this process with intrathecal LNP-delivered CRISPR/dCas9-mediated gene repression produces durable, non-opioid analgesia across multiple chronic pain models.

neuroscience↗

bioRxiv2026

Programming cell behavior with synthetic protease-activated receptors

Extracellular proteases are important signaling molecules in coagulation, inflammation, cell migration, and pain. Dysregulation of extracellular protease activity is common in diseases that perturb these critical functions. Engineering cells to sense and respond programmatically to protease activity has applications in biosensing, cell-based screening for protease activity, and therapeutics. Here we report synthetic protease-activated receptors (SynPARs) based on engineered, auto-inhibited G protein-coupled receptors (GPCRs). Relief of autoinhibition by proteolysis enables receptor activation by an exogenous or tethered agonist to generate transgene expression, real-time fluorescence, or endogenous G-protein signaling. We demonstrate SynPAR modularity with diverse secreted proteases, establish a cell-based SynPAR library selection to optimize protease recognition sequences, and control neuronal activity in response to protease activity. Finally, we use SynPARs in the dorsal root ganglion of mice to counteract hyperalgesia produced by trypsin activity, rewiring neurons to produce an analgesic response to a pain-inducing stimulus. Our study establishes SynPAR as a versatile and modular platform for recording, sensing, and responding to pericellular proteolysis. This fills a critical gap in protease-sensing tools and lays the groundwork for protease-activated genetic and cell-based medicines.

bioengineering↗

bioRxiv2026

Triptans reprogram Schwann cells to drive medication-overuse headache via β-glycan/TGF-β3 signaling

Medication-overuse headache (MOH) is one of the leading causes of chronic daily headache worldwide and arises from the repeated use of acute anti-migraine medications, including triptans. However, the cellular substrates and intracellular pathways driving this paradoxical chronification remain unknown. Here, using Schwann cell-selective silencing of the 5-HT1B/D receptor, we observed that acute triptan administration counteracts CGRP-induced, endosome-confined cAMP accumulation, preventing the development of periorbital mechanical allodynia in mice. In contrast, prolonged 5-HT1B/D activation in Schwann cells induces epigenetic and transcriptomic dysregulation associated with MOH in mice. Specifically, intronic hypermethylation-driven overexpression of BETAGLYCAN promotes activation of a non-canonical TGF-{beta}-dependent signaling cascade. The resulting TGF-{beta}3 upregulation establishes a feed-forward loop that sustains proalgesic paracrine communication between Schwann cells and primary sensory neurons. Analysis of plasma levels from patients with MOH confirmed elevated TGF-{beta}3 levels specifically associated with triptan-dependent MOH, supporting the translational relevance of these findings. Together, our data identify Schwann cell 5-HT1B/D signaling as a dual mediator of both the acute anti-migraine efficacy and the maladaptive mechanisms underlying MOH. These results provide a conceptual framework for strategies aimed at preserving the therapeutic benefits of triptans while limiting the adverse consequences of chronic administration. One Sentence SummarySchwann cell 5-HT1B/D receptor signaling mediates the dual effects of triptans by acutely inhibiting CGRP-driven nociceptive pathways while chronically promoting epigenetically driven TGF-{beta}3-dependent proalgesic signaling that causes medication-overuse headache.

pharmacology and toxicology↗

bioRxiv2026

Disentangling Schwann Cell and Neuronal TRPA1 Function in Mouse Models of Familial Episodic Pain Syndrome

Familial Episodic Pain Syndrome (FEPS) is a rare inherited disorder characterized by episodes of severe upper-body pain triggered by different stimuli including cold, stress, or fasting. A gain-of-function point mutation (N855S) in the Transient Receptor Potential Ankyrin 1 (TRPA1) ion channel has been identified in affected individuals, altering its biophysical properties, and leading to sustained nociceptive signaling. While TRPA1 is predominantly studied in sensory neurons, recent findings highlight its key modulatory role for Schwann cells in chronic pain. Here, we investigated the cell-specific contributions of mutant TRPA1 (TRPA1*) in FEPS by developing mouse models with TRPA1* selectively expressed in either Schwann cells or sensory neurons, using CRISPR-based and Cre-loxP strategies. Patch-clamp analyses confirmed that TRPA1* exhibits enhanced current responses to agonists compared to wild-type. Through behavioral assays we revealed that TRPA1* expressed in sensory neurons mediates acute nociception, while TRPA1* in Schwann cells drives mechanical allodynia in response to subthreshold doses of TRPA1 agonists and to physiological pain triggers commonly observed in FEPS patients, including fasting, cold exposure, and restraint stress. Pain responses were associated with the increase in reactive oxygen species (ROS) and accumulation of 4-hydroxynonenal (4-HNE) in TRPA1* sciatic nerves and these effects were reduced by a treatment with an antioxidant. We reveal distinct roles of neuronal and non-neuronal TRPA1 in pain and provide novel in vivo models to investigate the mechanisms of chronic pain in FEPS and related channelopathies. Overall, this study offers new insights into the development of targeted therapies for Schwann cell-TRPA1 to relieve pain in affected individuals.

genetics↗

bioRxiv2024

PoreMeth2: decoding the evolution of methylome alterations with Nanopore sequencing

In epigenetic analysis, identifying differentially methylated regions (DMRs) typically involves detecting groups of consecutive CpGs that show significant changes in their average methylation levels. However, the methylation state of a genomic region can also be characterized by a mixture of patterns (epialleles) with variable frequencies, and the relative proportions of such patterns can provide insights into its mechanisms of formation. Traditional methods based on bisulfite conversion and NGS, due to the read size (150 bp), allow epiallele frequency analysis only in high-CpG-density regions, limiting differential methylation studies to just 50% of the human methylome. Nanopore sequencing, with its long reads, enables the analysis of epiallele frequency across both high- and low-CpG-density regions. We introduce a novel computational approach, PoreMeth2, an R library that integrates epiallelic diversity and methylation frequency changes from Nanopore data to identify DMRs, assess their formation mechanisms, and annotate them to genic and regulatory elements. We applied PoreMeth2 to cancer and glial cell datasets, demonstrating its ability to distinguish epigenomic changes with a strong effect on gene expression from those with a weaker impact on transcriptional activity. PoreMeth2 is publicly available at https://github.com/Lab-CoMBINE/PoreMeth2.

bioinformatics↗

bioRxiv2024

Targeting the Schwann Cell EP2/cAMP Nanodomain to Block Pain but not Inflammation

Analgesia by non-steroidal anti-inflammatory drugs (NSAIDs) is ascribed to inhibition of prostaglandin (PG) biosynthesis and ensuing inflammation. However, NSAIDs have life-threatening side effects, and inhibition of inflammation delays pain resolution. Decoupling the mechanisms underlying PG-evoked pain vs. protective inflammation would facilitate pain treatment. Herein, we reveal that selective silencing of the PGE2 EP2 receptor in Schwann cells via an adeno-associated viral vector abrogates the indomethacin-sensitive component of pain-like responses in mice elicited by inflammatory stimuli without affecting inflammation. In human Schwann cells and in mice, EP2 activation and optogenetic stimulation of adenylyl cyclase evokes a plasma membrane-compartmentalized cyclic adenosine monophosphate (cAMP) signal that, via A-kinase anchor protein-associated protein kinase A, sustains inflammatory pain-like responses, but does not delay their resolution. Thus, an unforeseen and druggable EP2 receptor in Schwann cells, via specific cAMP nanodomains, encodes PG-mediated persistent inflammatory pain but not protective inflammation.

pharmacology and toxicology↗

bioRxiv2018

Dimerisation of APOBEC1 is dispensable for its RNA editing activity

The AID/APOBECs are DNA/RNA deaminases whose mutagenic activity has been linked to cancer. Among them, APOBEC1 physiologically partakes into a complex that edits a CAA codon into UAA Stop codon in the transcript of Apolipoprotein B (APOB), a protein crucial in the transport of lipids in the blood. Catalytically inactive mutants of APOBEC1 have a dominant negative effect on its activity, as they compete for the targeting of the APOB mRNA. Here we titrate APOBEC1-mediated editing in presence of catalytically inactive chimeras and mutants of APOBEC1, and we show that APOBEC1 inability to dimerise is the main determinant for its activity. This property is especially evident in an APOBEC1 mutant (L173A G227A) with increased activity on RNA despite decreased self-interaction. Moreover, dimerisation protects APOBEC1 from degradation and regulates its availability. Considering APOBEC1 capability to target DNA, we demonstrate that increased availability of the protein due to dimerisation leads to increase in the DNA damage induced by APOBEC1. These findings demonstrate that dimerisation, a property common to other APOBECs targeting DNA, might represent another layer in the regulation of this editing enzyme. BULLET POINTSO_LIAPOBEC1 inability to dimerise is the main determinant for its activity. C_LIO_LIDimerisation protects APOBEC1 from degradation and regulates its availability. C_LIO_LIAlterations in the balance between monomeric and dimeric APOBEC1 increase DNA damage. C_LI

molecular biology↗