Role of endoscopic ultrasound-guided fine needle aspiration biopsies in diagnosing pancreatic neoplasms in the paediatric population: experience from a tertiary center and review of the literature.

BACKGROUND: Endoscopic ultrasound-guided fine needle aspiration biopsy (EUS FNAB) is a well established diagnostic method in adult patients, but is rarely used in the paediatric population. The Clinical Department of Gastroenterology at the University Clinical Centre Ljubljana and the Department of Cytopathology at the Institute of Pathology, Faculty of Medicine, University of Ljubljana, Slovenia, have been closely collaborating on EUS FNAB since the introduction in 2010. The aim of the study was to review the cases of EUS FNAB of pancreatic neoplasms in children. PATIENTS AND METHODS: In the digital archive of the Institute of Pathology (IP), Faculty of Medicine (FM), University of Ljubljana (UL), we found 6 cases of EUS FNAB in children, 3 had EUS FNAB of the pancreas, 2 of whom had a cytopathologic diagnosis of a tumour. In the first case, the lesion was ultrasonographically solid, and the cell sample contained branching papillary structures surrounded by aggregates of small cells with nuclear grooves. In the second case, the lesion was ultrasonographically cystic, and predominantly necrosis was seen, with only single preserved cells. Positive nuclear reaction for ß-catenin was found in both cases by immunohistochemical staining. RESULTS: In both cases, the cytopathological diagnosis of solid pseudopapillary neoplasm of the pancreas was made, the cases represent the totality of paediatric cases of pancreatic neoplasms from the Children's Hospital Ljubljana since 2010. There were no adverse events during and after EUS FNAB. A histopathological examination of the tumour resection specimens confirmed the cytopathological diagnosis. CONCLUSIONS: Our experience indicates that EUS FNAB is a safe and effective method for diagnosing pancreatic neoplasms in the pediatric population, as supported by the findings in the literature.

TECPR1 is activated by damage-induced sphingomyelin exposure to mediate noncanonical autophagy.

Cells use noncanonical autophagy, also called conjugation of ATG8 to single membranes (CASM), to label damaged intracellular compartments with ubiquitin-like ATG8 family proteins in order to signal danger caused by pathogens or toxic compounds. CASM relies on E3 complexes to sense membrane damage, but so far, only the mechanism to activate ATG16L1-containing E3 complexes, associated with proton gradient loss, has been described. Here, we show that TECPR1-containing E3 complexes are key mediators of CASM in cells treated with a variety of pharmacological drugs, including clinically relevant nanoparticles, transfection reagents, antihistamines, lysosomotropic compounds, and detergents. Interestingly, TECPR1 retains E3 activity when ATG16L1 CASM activity is obstructed by the Salmonella Typhimurium pathogenicity factor SopF. Mechanistically, TECPR1 is recruited by damage-induced sphingomyelin (SM) exposure using two DysF domains, resulting in its activation and ATG8 lipidation. In vitro assays using purified human TECPR1-ATG5-ATG12 complex show direct activation of its E3 activity by SM, whereas SM has no effect on ATG16L1-ATG5-ATG12. We conclude that TECPR1 is a key activator of CASM downstream of SM exposure.

Small regions as key sources of traditional knowledge: a quantitative ethnobotanical survey in the central Balkans.

BACKGROUND: Starting from the idea that unexplored areas may yield new and different ethnobotanical information, we performed a survey of traditional uses of plants in two neighboring districts situated in east Serbia (Bor and Aleksinac), both lacking in previous ethnobotanical reports, but characterized by an interesting history and culture, together with some specific features. In this study, we hypothesized that such small and specific areas could be of high ethnobotanical importance. METHODS: Semi-structured interviews were used with 155 informants. Relative cultural importance (RCI) indices, such as the frequency of citation (FC), relative frequency of citation (RFC), relative importance index (RI), informant consensus factor (ICF-FIC), use value (UV), fidelity level (FL) and Jaccard index (JI), were calculated, and principal coordinate analysis (PCoA) was performed. RESULTS: In this study, 2333 use-reports and 114 plants were recorded. Of the 101 medical herbs, 33 are included in the European Pharmacopoeia Edition 8.0. The most frequently used mode of preparation was as an infusion (50.0%), while leaf (44.7%) was the most used plant part. The highest FC and RFC values were recorded for Hypericum perforatum L. (13.1 and 0.2, respectively), while the highest RI was documented for Urtica dioica L. (1.0). ICF and FL indices showed important differences among selected groups of informants. The PCoA showed three homogeneous plant groups. Plants were mostly used for the treatment of digestive (49.1%), circulatory (41.2%) and respiratory system disorders (35.1%). Thirty-seven (32.5%) herbs were used for human nutrition, 14 (12.3%) in veterinary medicine, 17 (14.9%) in rituals and ethnoculture, while 24 (21.0%) for miscellaneous purposes. The highest degree of similarity was determined with studies conducted in close proximity. Four species are new to Balkan ethnobotany. New uses for some well-known plants are highlighted. CONCLUSION: The study indicated that small and specific areas in the Balkans may be an important reservoir of ethnobotanical knowledge.

Cholesterol transfer via endoplasmic reticulum contacts mediates lysosome damage repair.

Lysosome integrity is essential for cell viability, and lesions in lysosome membranes are repaired by the ESCRT machinery. Here, we describe an additional mechanism for lysosome repair that is activated independently of ESCRT recruitment. Lipidomic analyses showed increases in lysosomal phosphatidylserine and cholesterol after damage. Electron microscopy demonstrated that lysosomal membrane damage is rapidly followed by the formation of contacts with the endoplasmic reticulum (ER), which depends on the ER proteins VAPA/B. The cholesterol-binding protein ORP1L was recruited to damaged lysosomes, accompanied by cholesterol accumulation by a mechanism that required VAP-ORP1L interactions. The PtdIns 4-kinase PI4K2A rapidly produced PtdIns4P on lysosomes upon damage, and knockout of PI4K2A inhibited damage-induced accumulation of ORP1L and cholesterol and led to the failure of lysosomal membrane repair. The cholesterol-PtdIns4P transporter OSBP was also recruited upon damage, and its depletion caused lysosomal accumulation of PtdIns4P and resulted in cell death. We conclude that ER contacts are activated on damaged lysosomes in parallel to ESCRTs to provide lipids for membrane repair, and that PtdIns4P generation and removal are central in this response.

Pheochromocytoma presenting as a pancreatic mass: Avoiding a dangerous pitfall in endoscopic ultrasound-guided fine needle aspiration biopsy using GATA3 immunostain.

Pheochromocytomas and sympathetic paragangliomas are rare tumours arising from chromaffin cells, producing catecholamines in various amounts. Fatal hypertensive episodes may occur perioperatively, which are preventable by alpha adrenergic receptor blockers. The perioperative mortality rate of diagnosed versus undiagnosed catecholamine-producing tumours is significant, considering that only a minority of tumours develop metastasis. Herein we describe a case of a primary adrenal pheochromocytoma referred to as a pancreatic tumour, successfully diagnosed by endoscopic ultrasound-guided fine needle aspiration biopsy, with a distinct morphology (prominent nuclear anisonucleosis, intranuclear pseudoinclusions, and multinucleation) and immunohistochemical signature.

Inhibiting autophagy increases the efficacy of low-dose photodynamic therapy.

Rupture and permeabilization of endocytic vesicles can be triggered by various causes, such as pathogenic invasions, amyloid proteins, and silica crystals leading to cell death and degeneration. A cellular quality control process, called lysophagy was recently described to target damaged lysosomes for autophagic sequestration within isolation membranes in order to protect the cell from the consequences of lysosomal leakage. This protective process, however, might interfere with treatment conditions, such as photodynamic therapy (PDT) and the intracellular drug delivery method photochemical internalization (PCI). PCI-induced permeabilization of endosomes and lysosomes is purposely triggered to release drugs that are sequestered in these organelles into the cytosol in order to synergistically kill cancer cells. Here, we show that photochemical treatment with the PCI-photosensitizer TPCS2a/fimaporfin results in both induction of autophagy and inhibition of the autophagic flux. The autophagic response is accompanied by recruitment of ubiquitin (Ubq), p62, and microtubule-associated protein 1A/1B-light chain 3 (LC3) to damaged vesicles, marked by Galectin 3 (Gal3). Furthermore, ultrastructural analysis revealed a homogenously thick p62-positive layer surrounding these permeabilized vesicles. Although p62 seems to be important during the selective autophagic sequestration, we show that its presence is not essential for the effective removal of damaged vesicles or the recovery of the lysosomal content. An active autophagic response and the presence of p62, however, is important for cancer cells to survive low-dose TPCS2a-PDT. Thus, targeting both p62 and autophagy together and independently, in a light-controlled/PCI based delivery of cancer therapeutics could increase the effectiveness of the treatment regime.

The thrombin receptor modulates astroglia-neuron trophic coupling and neural repair after spinal cord injury.

Excessive activation of the thrombin receptor, protease activated receptor 1 (PAR1) is implicated in diverse neuropathologies from neurodegenerative conditions to neurotrauma. PAR1 knockout mice show improved outcomes after experimental spinal cord injury (SCI), however information regarding the underpinning cellular and molecular mechanisms is lacking. Here we demonstrate that genetic blockade of PAR1 in female mice results in improvements in sensorimotor co-ordination after thoracic spinal cord lateral compression injury. We document improved neuron preservation with increases in Synapsin-1 presynaptic proteins and GAP43, a growth cone marker, after a 30 days recovery period. These improvements were coupled to signs of enhanced myelin resiliency and repair, including increases in the number of mature oligodendrocytes, their progenitors and the abundance of myelin basic protein. These significant increases in substrates for neural recovery were accompanied by reduced astrocyte (Serp1) and microglial/monocyte (CD68 and iNOS) pro-inflammatory markers, with coordinate increases in astrocyte (S100A10 and Emp1) and microglial (Arg1) markers reflective of pro-repair activities. Complementary astrocyte-neuron co-culture bioassays suggest astrocytes with PAR1 loss-of-function promote both neuron survival and neurite outgrowth. Additionally, the pro-neurite outgrowth effects of switching off astrocyte PAR1 were blocked by inhibiting TrkB, the high affinity receptor for brain derived neurotrophic factor. Altogether, these studies demonstrate unique modulatory roles for PAR1 in regulating glial-neuron interactions, including the capacity for neurotrophic factor signaling, and underscore its position at neurobiological intersections critical for the response of the CNS to injury and the capacity for regenerative repair and restoration of function.

Sealing holes in cellular membranes.

The compartmentalization of eukaryotic cells, which is essential for their viability and functions, is ensured by single or double bilayer membranes that separate the cell from the exterior and form boundaries between the cell's organelles and the cytosol. Nascent nuclear envelopes and autophagosomes, which both are enveloped by double membranes, need to be sealed during the late stage of their biogenesis. On the other hand, the integrity of cellular membranes such as the plasma membrane, lysosomes and the nuclear envelope can be compromised by pathogens, chemicals, radiation, inflammatory responses and mechanical stress. There are cellular programmes that restore membrane integrity after injury. Here, we review cellular mechanisms that have evolved to maintain membrane integrity during organelle biogenesis and after injury, including membrane scission mediated by the endosomal sorting complex required for transport (ESCRT), vesicle patching and endocytosis.

The thrombin receptor links brain derived neurotrophic factor to neuron cholesterol production, resiliency and repair after spinal cord injury.

Despite concerted efforts to identify CNS regeneration strategies, an incomplete understanding of how the needed molecular machinery is regulated limits progress. Here we use models of lateral compression and FEJOTA clip contusion-compression spinal cord injury (SCI) to identify the thrombin receptor (Protease Activated Receptor 1 (PAR1)) as an integral facet of this machine with roles in regulating neurite growth through a growth factor- and cholesterol-dependent mechanism. Functional recovery and signs of neural repair, including expression of cholesterol biosynthesis machinery and markers of axonal and synaptic integrity, were all increased after SCI in PAR1 knockout female mice, while PTEN was decreased. Notably, PAR1 differentially regulated HMGCS1, a gene encoding a rate-limiting enzyme in cholesterol production, across the neuronal and astroglial compartments of the intact versus injured spinal cord. Pharmacologic inhibition of cortical neuron PAR1 using vorapaxar in vitro also decreased PTEN and promoted neurite outgrowth in a cholesterol dependent manner, including that driven by suboptimal brain derived neurotrophic factor (BDNF). Pharmacologic inhibition of PAR1 also augmented BDNF-driven HMGCS1 and cholesterol production by murine cortical neurons and by human SH-SY5Y and iPSC-derived neurons. The link between PAR1, cholesterol and BDNF was further highlighted by demonstrating that the deleterious effects of PAR1 over-activation are overcome by supplementing cultures with BDNF, cholesterol or by blocking an inhibitor of adenylate cyclase, Gαi. These findings document PAR1-linked neurotrophic coupling mechanisms that regulate neuronal cholesterol metabolism as an important component of the machinery regulating CNS repair and point to new strategies to enhance neural resiliency after injury.

A novel combined animal tissue model for freehand and ultrasound-guided fine needle aspiration biopsy and smear preparation techniques training.

OBJECTIVE: A variety of models are used for fine needle aspiration biopsy (FNAB) and smear preparation techniques training: human, animal and silicon models or combined models. We present fresh animal tissues as models for freehand and ultrasound (US)-guided FNAB technique training, enabling an integrated approach from tumour detection to smear evaluation. METHODS: We introduced a novel combined animal tissue model using dietary animal meat with covering skin as a substrate. Animal liver tissue of various sizes, representing tumour, was inserted into the various layers of the substrate (subcutaneous fat, muscle tissue, proximity of bone). Freehand and US-guided FNAB smear preparation, including fixation, was then performed and assessed. RESULTS: The use of a combined animal tissue model for 6 freehand and 3 US-guided FNAB sessions showed a statistically significant improvement in the US-guided FNAB retrieval of liver tissue (Fisher's exact test, p =  .0216), in smear preparation technique reflected in a decrease in the number of too thick smears after freehand FNAB (Fisher's exact test, p  =  .0070), in the overall number of smears satisfactory for evaluation by US-guided FNAB (Fisher's exact test, p =  .0206) and in the number of flawless smears obtained in the freehand FNAB training sessions (Fisher's exact test, p =  .0020). CONCLUSIONS: A unique advantage of the presented model encompassing various layers of animal tissues with covering skin, offers an integrated approach for FNAB training from "tumour" detection, puncture precision, to smear preparation and cytological evaluation for a wider audience and does not compromise patient safety.

LRRK2 to the rescue of damaged endomembranes.

Mutations in several genes encoding for lysosomal proteins are involved in Parkinson's disease (PD). In this issue, Herbst et al (2020) show that PD-related leucine-rich repeat kinase 2 (LRRK2) is activated in response to pathogen or membranolytic drug-induced damage of phagolysosomes and lysosomes in macrophages, and regulates endolysosomal homeostasis by controlling the balance between membrane repair and degradation.

Erythroblastic sarcoma presenting as a unilateral pleural effusion in a patient with myelodysplastic syndrome/myeloproliferative neoplasm-Case report with cytologic and histologic findings.

Erythroblastic sarcoma (ES) is an extremely rare extramedullary solid tumor of immature erythroid cells, with around a dozen human cases to our knowledge presented so far in the english literature, none of the cases presenting as pleural effusion. Here we describe a case of ES in a thoracic wall diagnosed by cytological examination of pleural effusion with histologic correlation in a 53-year-old patient with a 10-month history of myelodysplastic syndrome/myeloproliferative neoplasm with grade III reticulin myelofibrosis.

ESCRT-mediated phagophore sealing during mitophagy.

Inactivation of the endosomal sorting complex required for transport (ESCRT) machinery has been reported to cause autophagic defects, but the exact functions of ESCRT proteins in macroautophagy/autophagy remain incompletely understood. Using live-cell fluorescence microscopy we found that the filament-forming ESCRT-III subunit CHMP4B was recruited transiently to nascent autophagosomes during starvation-induced autophagy and mitophagy, with residence times of about 1 and 2 min, respectively. Correlative light microscopy and electron tomography revealed CHMP4B recruitment at a late step in mitophagosome formation. The autophagosomal dwell time of CHMP4B was strongly increased by depletion of the regulatory ESCRT-III subunit CHMP2A. Using a novel optogenetic closure assay we observed that depletion of CHMP2A inhibited phagophore sealing during mitophagy. Consistent with this, depletion of CHMP2A and other ESCRT-III subunits inhibited both PRKN/PARKIN-dependent and -independent mitophagy. We conclude that the ESCRT machinery mediates phagophore closure, and that this is essential for mitophagic flux.Abbreviations: BSA: bovine serum albumin; CHMP: chromatin-modifying protein; CLEM: correlative light and electron microscopy; EGFP: enhanced green fluorescent protein; ESCRT: endosomal sorting complex required for transport; HEPES: 2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid; HRP: horseradish peroxidase; ILV: intralumenal vesicle; MAP1LC3/LC3: microtubule-associated protein 1 light chain 3; LOV2: light oxygen voltage 2; MLS: mitochondrial localization sequence; MT-CO2: mitochondrially encoded cytochrome c oxidase II; O+A: oligomycin and antimycin A; PBS: phosphate-buffered saline; PIPES: piperazine-N,N-bis(2-ethanesulfonic acid); PRKN/PARKIN: parkin RBR E3 ubiquitin protein ligase; RAB: RAS-related in brain; SD: standard deviation; SEM: standard error of the mean; TOMM20: TOMM20: translocase of outer mitochondrial membrane 20; VCL: vinculin; VPS4: vacuolar protein sorting protein 4; Zdk1: Zdark 1; TUBG: Tubulin gamma chain.

The many functions of ESCRTs.

Cellular membranes can form two principally different involutions, which either exclude or contain cytosol. The 'classical' budding reactions, such as those occurring during endocytosis or formation of exocytic vesicles, involve proteins that assemble on the cytosol-excluding face of the bud neck. Inverse membrane involution occurs in a wide range of cellular processes, supporting cytokinesis, endosome maturation, autophagy, membrane repair and many other processes. Such inverse membrane remodelling is mediated by a heteromultimeric protein machinery known as endosomal sorting complex required for transport (ESCRT). ESCRT proteins assemble on the cytosolic (or nucleoplasmic) face of the neck of the forming involution and cooperate with the ATPase VPS4 to drive membrane scission or sealing. Here, we review similarities and differences of various ESCRT-dependent processes, with special emphasis on mechanisms of ESCRT recruitment.

ESCRT-mediated lysosome repair precedes lysophagy and promotes cell survival.

Although lysosomes perform a number of essential cellular functions, damaged lysosomes represent a potential hazard to the cell. Such lysosomes are therefore engulfed by autophagic membranes in the process known as lysophagy, which is initiated by recognition of luminal glycoprotein domains by cytosolic lectins such as Galectin-3. Here, we show that, under various conditions that cause injury to the lysosome membrane, components of the endosomal sorting complex required for transport (ESCRT)-I, ESCRT-II, and ESCRT-III are recruited. This recruitment occurs before that of Galectin-3 and the lysophagy machinery. Subunits of the ESCRT-III complex show a particularly prominent recruitment, which depends on the ESCRT-I component TSG101 and the TSG101- and ESCRT-III-binding protein ALIX Interference with ESCRT recruitment abolishes lysosome repair and causes otherwise reversible lysosome damage to become cell lethal. Vacuoles containing the intracellular pathogen Coxiella burnetii show reversible ESCRT recruitment, and interference with this recruitment reduces intravacuolar bacterial replication. We conclude that the cell is equipped with an endogenous mechanism for lysosome repair which protects against lysosomal damage-induced cell death but which also provides a potential advantage for intracellular pathogens.

ESCRTs in membrane sealing.

The multisubunit endosomal sorting complex required for transport (ESCRT) machinery is a key regulator of cellular membrane dynamics. Initially characterized in the budding yeast Saccharomyces cerevisiae for its involvement in cargo sorting to the vacuole, the yeast lysosome, this protein complex has emerged over the past decade as a driver for diverse membrane remodeling processes. Its pleiotropic functional connection is mirrored in numerous cellular processes, such as cytokinetic abscission during the final step of cell division, nuclear pore quality control, nuclear envelope sealing and repair, plasma membrane repair, vesicle shedding from the plasma membrane, viral budding, and axonal pruning. Common to all the processes regulated by the ESCRT machinery is their assembly on the cytosolic side of the respective membrane to stabilize concave membranes, budding, and scission of narrow membrane necks away from the cytosol. Thus, this machinery has evolved to perform many functions in membrane dynamics, and given its importance, it is not surprising that the dysfunctional ESCRT machinery has been implicated in several diseases. In this mini-review, we summarize the role of ESCRT proteins in membrane deformation specifically during membrane sealing and repair.

Kallikrein-related peptidase 6 orchestrates astrocyte form and function through proteinase activated receptor-dependent mechanisms.

Kallikrein-related peptidase 6 (Klk6) is the most abundant serine proteinase in the adult central nervous system (CNS), yet we know little regarding its physiological roles or mechanisms of action. Levels of Klk6 in the extracellular environment are dynamically regulated in CNS injury and disease positioning this secreted enzyme to affect cell behavior by potential receptor dependent and independent mechanisms. Here we show that recombinant Klk6 evokes increases in intracellular Ca2+ in primary astrocyte monolayer cultures through activation of proteinase activated receptor 1 (PAR1). In addition, Klk6 promoted a condensation of astrocyte cortical actin leading to an elongated stellate shape and multicellular aggregation in a manner that was dependent on the presence of either PAR1 or PAR2. Klk6-evoked changes in astrocyte shape were accompanied by translocation of ß-catenin from the plasma membrane to the cytoplasm. These data are exciting because they demonstrate that Klk6 can influence astrocyte plasticity through receptor-dependent mechanisms. Furthermore, this study expands our understanding of the mechanisms by which kallikreins can contribute to neural homeostasis and remodeling and point to both PAR1 and PAR2 as new therapeutic targets to modulate astrocyte form and function.

Homocysteine regulates fatty acid and lipid metabolism in yeast.

S-Adenosyl-l-homocysteine hydrolase (AdoHcy hydrolase; Sah1 in yeast/AHCY in mammals) degrades AdoHcy, a by-product and strong product inhibitor of S-adenosyl-l-methionine (AdoMet)-dependent methylation reactions, to adenosine and homocysteine (Hcy). This reaction is reversible, so any elevation of Hcy levels, such as in hyperhomocysteinemia (HHcy), drives the formation of AdoHcy, with detrimental consequences for cellular methylation reactions. HHcy, a pathological condition linked to cardiovascular and neurological disorders, as well as fatty liver among others, is associated with a deregulation of lipid metabolism. Here, we developed a yeast model of HHcy to identify mechanisms that dysregulate lipid metabolism. Hcy supplementation to wildtype cells up-regulated cellular fatty acid and triacylglycerol content and induced a shift in fatty acid composition, similar to changes observed in mutants lacking Sah1. Expression of the irreversible bacterial pathway for AdoHcy degradation in yeast allowed us to dissect the impact of AdoHcy accumulation on lipid metabolism from the impact of elevated Hcy. Expression of this pathway fully suppressed the growth deficit of sah1 mutants as well as the deregulation of lipid metabolism in both the sah1 mutant and Hcy-exposed wildtype, showing that AdoHcy accumulation mediates the deregulation of lipid metabolism in response to elevated Hcy in yeast. Furthermore, Hcy supplementation in yeast led to increased resistance to cerulenin, an inhibitor of fatty acid synthase, as well as to a concomitant decline of condensing enzymes involved in very long-chain fatty acid synthesis, in line with the observed shift in fatty acid content and composition.

Protease activated receptor 2 controls myelin development, resiliency and repair.

Oligodendrocytes are essential regulators of axonal energy homeostasis and electrical conduction and emerging target cells for restoration of neurological function. Here we investigate the role of protease activated receptor 2 (PAR2), a unique protease activated G protein-coupled receptor, in myelin development and repair using the spinal cord as a model. Results demonstrate that genetic deletion of PAR2 accelerates myelin production, including higher proteolipid protein (PLP) levels in the spinal cord at birth and higher levels of myelin basic protein and thickened myelin sheaths in adulthood. Enhancements in spinal cord myelin with PAR2 loss-of-function were accompanied by increased numbers of Olig2- and CC1-positive oligodendrocytes, as well as in levels of cyclic adenosine monophosphate (cAMP), and extracellular signal related kinase 1/2 (ERK1/2) signaling. Parallel promyelinating effects were observed after blocking PAR2 expression in purified oligodendrocyte cultures, whereas inhibiting adenylate cyclase reversed these effects. Conversely, PAR2 activation reduced PLP expression and this effect was prevented by brain derived neurotrophic factor (BDNF), a promyelinating growth factor that signals through cAMP. PAR2 knockout mice also showed improved myelin resiliency after traumatic spinal cord injury and an accelerated pattern of myelin regeneration after focal demyelination. These findings suggest that PAR2 is an important controller of myelin production and regeneration, both in the developing and adult spinal cord.