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Hh signaling has been shown to be activated in intact and injured peripheral nerve. However, the role of Hh signaling in peripheral nerve is not fully understood. In the present study, we observed that Hh signaling responsive cells [Gli1(+) cells] in both the perineurium and endoneurium. In the endoneurium, Gli1(+) cells were classified as blood vessel associated or non-associated. After injury, Gli1(+) cells around blood vessels mainly proliferated to then accumulate into the injury site along with endothelial cells. Hh signaling activity was retained in Gli1(+) cells during nerve regeneration. To understand the role of Hedgehog signaling in Gli1(+) cells during nerve regeneration, we examined mice with Gli1(+) cells-specific inactivation of Hh signaling (Smo cKO). After injury, Smo cKO mice showed significantly reduced numbers of accumulated Gli1(+) cells along with disorganized vascularization at an early stage of nerve regeneration, which subsequently led to an abnormal extension of the axon. Thus, Hh signaling in Gli1(+) cells appears to be involved in nerve regeneration through controlling new blood vessel formation at an early stage.

Peripheral nerves are organized into discrete cellular compartments. Axons, Schwann cells (SCs), and endoneurial fibroblasts (EFs) reside within the endoneurium and are surrounded by the perineurium - a cellular sheath comprised of layers of perineurial glia (PNG). SC secretion of Desert Hedgehog (Dhh) regulates this organization. In Dhh nulls, the perineurium is deficient and the endoneurium is subdivided into small compartments termed minifascicles. Human Dhh mutations cause a peripheral neuropathy with similar defects. Here we examine the role of Gli1, a canonical transcriptional effector of hedgehog signaling, in regulating peripheral nerve organization. We identify PNG, EFs, and pericytes as Gli1-expressing cells by genetic fate mapping. Although expression of Dhh by SCs and Gli1 in target cells is coordinately regulated with myelination, Gli1 expression unexpectedly persists in Dhh null EFs. Thus, Gli1 is expressed in EFs non-canonically i.e., independent of hedgehog signaling. Gli1 and Dhh also have non-redundant activities. In contrast to Dhh nulls, Gli1 nulls have a normal perineurium. Like Dhh nulls, Gli1 nulls form minifascicles, which we show likely arise from EFs. Thus, Dhh and Gli1 are independent signals: Gli1 is dispensable for perineurial development but functions cooperatively with Dhh to drive normal endoneurial development. During development, Gli1 also regulates endoneurial extracellular matrix production, nerve vascular organization, and has modest, non-autonomous effects on SC sorting and myelination of axons. Finally, in adult nerves, induced deletion of Gli1 is sufficient to drive minifascicle formation. Thus, Gli1 regulates the development and is required to maintain the endoneurial architecture of peripheral nerves.SIGNIFICANCE STATEMENTPeripheral nerves are organized into distinct cellular/ECM compartments: the epineurium, perineurium and endoneurium. This organization, with its associated cellular constituents, are critical for the structural and metabolic support of nerves and their response to injury. Here, we show Gli1 - a transcription factor normally expressed downstream of hedgehog signaling - is required for the proper organization of the endoneurium but not the perineurium. Unexpectedly, Gli1 expression by endoneurial cells is independent of, and functions non-redundantly with, Schwann Cell-derived Desert Hedgehog in regulating peripheral nerve architecture. These results further delineate how peripheral nerves acquire their distinctive organization during normal development and highlight mechanisms that may regulate their reorganization in pathologic settings including peripheral neuropathies and nerve injury.

The sheaths of the damaged peripheral nerve of Wistar-Kyoto rats were studied after single subperineural administration of bromodeoxyuridine (BrdU)-labeled bone marrow mesenchymal stem cells (MSC) from the same rats. The sciatic nerve was damaged by ligation for 40 sec directly before MSC administration. BrdU+ MSC were identified in the recipient nerve within 1 week after transplantation and were detected not only in the endoneurium, but also in the epineurium and perineurium. It was found that single administration of MSC into the damaged nerve trunk led to an almost 2-fold increase in the thickness of its sheaths (perineurium and epineurium) in comparison with the control group (ligation). It can be hypothesized that MSC induce thickening of nerve sheaths through the production of factors that stimulate angiogenesis and adipogenesis.

Background A perineural (Tarlov) cyst is a fluid-filled lesion occurring between the perineurium and the endoneurium of spinal nerve roots. The aim of the study was to evaluate the prevalence and morphology of perineural cysts, detected incidentally in patients with symptomatic degenerative disk disease. Materials/methods The study was based on the retrospective data gathered during magnetic resonance imaging (MRI) examinations. Results and Conclusions Out of 3,128 spinal MRI examinations, perineural cysts were detected in 286 patients (9%). The cysts were most commonly observed in the sacral region, followed by thoracic, cervical, and lumbar regions. Cysts were more common in women than in men and the average age of patients was 54.8 years. In the majority, a single cyst was found. The average longest dimension of the lesion was 11.72 mm.

The aim of this study was to analyze the expression of extracellular matrix (ECM) proteins in human endoneurium during aging. We harvested 15 cadaveric sural nerves, distributed in 3 age groups (I: 25-44, II: 45-64, III: 65-86 years old). Histological sections were stained immunohistochemically for the presence of collagen type I, type IV and laminin, and the ImageJ processing program was used in morphometrical analysis to determine the percentages of these endoneurial proteins. In two younger groups, the endoneurial matrix of the sural nerve was composed from about equal proportions of these proteins, which may be considered a favorable microenvironment for the regeneration of nerve fibers. Linear regression analysis showed a significant increase in endoneurial collagen type IV with age, while collagen type I and laminin significantly decreased during the aging process. In cases older than 65 years, remodeling of the endoneurial matrix was observed to be significantly higher for the presence of collagen type IV, and lower for the expression of collagen type I and laminin. This age-related imbalance of ECM proteins could represent a disadvantageous microenvironment for nerve fiber regeneration in older adults. Our findings contribute to the development of therapeutic approaches for peripheral nerve regeneration.

Patients with transthyretin (TTR)-type familial amyloid polyneuropathy (FAP) typically exhibit sensory dominant polyneuropathy and autonomic neuropathy. However, the molecular pathogenesis of the neuropathy remains unclear. In this study, we characterize the features of FAP TTR the substitution of lysine for glutamic acid at position 61 (E61K). This FAP was late-onset, with sensory dominant polyneuropathy, autonomic neuropathy, and cardiac amyloidosis. Interestingly, no amyloid deposits were found in the endoneurium of the four nerve specimens examined. Therefore, we examined the amyloidogenic properties of E61K TTR in vitro. Recombinant wild-type TTR, the substitution of methionine for valine at position 30 (V30M) TTR, and E61K TTR proteins were incubated at 37°C for 72 hr, and amyloid fibril formation was assessed using the thioflavin-T binding assay. Amyloid fibril formation by E61K TTR was less than that by V30M TTR, and similar to that by wild-type TTR. E61K TTR did not have an inhibitory effect on neurite outgrowth from adult rat dorsal root ganglion (DRG) neurons, but V30M TTR did. Furthermore, we studied the sural nerve of our patient by terminal deoxynucleotidyl transferase dUTP nick end labeling and electron microscopy. A number of apoptotic cells were observed in the endoneurium of the nerve by transferase dUTP nick end labeling. Chromatin condensation was confirmed in the nucleus of non-myelinating Schwann cells by electron microscopy. These findings suggest that E61K TTR is low amyloidogenic, in vitro and in vivo. However, TTR aggregates and amyloid fibrils in the DRG may cause sensory impairments in FAP because the DRG has no blood-nerve barrier. Moreover, Schwann cell apoptosis may contribute to the neurodegeneration.

Nerve defects are challenging to address clinically without satisfactory treatments. As a reliable alternative to autografts, decellularized nerve matrix scaffolds (DNM-S) have been widely used in clinics for surgical nerve repair. However, DNM-S remain inferior to autografts in their ability to support nerve regeneration for long nerve defects. In this study, we systematically and clearly presented the nano-architecture of nerve-specific structures, including the endoneurium, basement membrane and perineurium/epineurium in DNM-S. Furthermore, we modified the DNM-S by supplementing decellularized nerve matrix hydrogel (DNMG) and glial-derived neurotrophic factor (GDNF) and then bridged a 50-mm sciatic nerve defect in a beagle model. Fifteen beagles were randomly divided into three groups (five per group): an autograft group, DNM-S group and GDNF-DNMG-modified DNM-S (DNM-S/GDNF@DNMG) group. DNM-S/GDNF@DNMG, as optimized nerve grafts, were used to bridge nerve defects in the same manner as in the DNM-S group. The repair outcome was evaluated by behavioural observations, electrophysiological assessments, regenerated nerve tissue histology and reinnervated target muscle examinations. Compared with the DNM-S group, limb function, electrophysiological responses and histological findings were improved in the DNM-S/GDNF@DNMG group 6 months after grafting, reflecting a narrower gap between the effects of DNM-S and autografts. In conclusion, modification of DNM-S with DNMG and GDNF enhanced nerve regeneration and functional recovery, indicating that noncellular modification of DNM-S is a promising method for treating long nerve defects.

In inflammatory peripheral demyelinating disorders, demyelination represents segmental demyelination in which the myelin sheath of a myelinating Schwann cell (SC) is completely removed by macrophages or a partial myelin degeneration in the paranode occurring due to autoantibodies attacking the node/paranode. For the segmental demyelination from living myelin-forming SCs, macrophages infiltrate within the endoneurium and insinuate between myelin lamellae and the cytoplasm of SCs, and the myelin is then removed via phagocytosis. During the macrophage invasion into the SC cytoplasm from the node of Ranvier and internodal areas, the attacked SCs do not remain quiescent but transdifferentiate into inflammatory demyelinating SCs (iDSCs), which exhibit unique demyelination pathologies, such as myelin uncompaction from Schmidt-Lanterman incisures with myelin lamellae degeneration. The longitudinal extension of this self-myelin clearance process of iDSCs into the nodal region is associated with the degeneration of nodal microvilli and paranodal loops, which provides a potential locus for macrophage infiltration. In addition to the nodal intrusion, macrophages appear to be able to invade fenestrated internodal plasma membrane or the degenerated outer mesaxon of iDSC. These SC demyelination morphologies indicate that the SC reprogramming to iDSCs may be a prerequisite for macrophage-mediated inflammatory demyelination. In contrast, paranodal demyelination caused by autoantibodies to nodal/paranodal antigens does not result in iDSC-dependent macrophage infiltration and subsequent segmental demyelination. In the context of inflammatory demyelination, the novel perspective of iDSCs provides an important viewpoint to understand the pathophysiology of demyelinating peripheral neuropathies and establish diagnostic and therapeutic strategies.

Objective To investigate the pathological features of blood stasis syndrome (BSS) in non-diabetic peripheral neuropathy. Methods Clinical data of 31 patients with non-diabetic peripheral neuropathy who had undergone nerve biopsy during December 2004 and December 2010 in Xuanwu Hospital Capital Medical University were retrospectively analyzed. According to Chinese medicine (CM) syndrome differentiation and signs, 26 patients were blood stasis type and 5 patients were non-blood stasis type. Clinical and pathological data were compared in detail. Results Clinically, although both groups shared similar symptoms of limb numbness, weakness and sensory disturbances, the prevalence of neuralgia was much grievous in BSS group (73.1%, 26/31) compared with the non-BSS group (0%, 0/5). As for signs, dermal nutrients disturbance (84.6%, 22/26), dark or purple tongue (100.0%, 26/26), and sublingual varices (80.7%, 21/26) were more common in the BSS group than the non-BSS group (0%, 60%, 20%, respectively). The prevalence of qi deficiency cases (19/26) in the BSS group was significantly higher compared with the non-BSS group (1/5). The unique histological manifestations of BSS were axonal degeneration (16/26 vs 2/5 in non-BSS group), which was the hallmark of ischemia. Cases with BSS had prominent microangiopathy (61.5%, 16/26), manifested as epineurium vasculitis (inflammatory cell infiltrated to the vessel wall, obliteration and recanalization, vascular proliferation, extravascular hemosiderin deposition), angiotelectasis, proliferation and hyaline degeneration of endoneurium capillary. In the BSS group, impaired blood-nerve barrier was indicated by sub-perineurial edema (46.2%, 11/26) and endoneurial edema (15.4%, 4/26). The Renaut body (15.4%, 4/26) and amyloid deposition (3.8%, 1/26) found in the BSS group were absent in the non-BSS group. Conclusions BBS was common in non-diabetic peripheral neuropathies. The nerves exhibited ischemic alteration of primary axon degeneration and secondary demyelination. The interstitial tissue revealed microcirculation impairment, blood-nerve barrier disturbance, amyloid deposition and proliferation changes. The high prevalence of qi deficiency also highlights the therapy of promotion of blood circulation and removal of blood stasis.

The blood-nerve barrier (BNB) formed by tight junction-forming endoneurial microvessels located in the innermost compartment of peripheral nerves, and the perineurium serve to maintain the internal microenvironment required for normal signal transduction. The specific molecular components that define the normal adult human BNB are not fully known. Guided by data derived from the adult human BNB transcriptome, we evaluated the in situ expression of 25 junctional complex, transporter, cell membrane, and cytoskeletal proteins in four histologically normal adult sural nerves by indirect fluorescent immunohistochemistry to determine proteins specifically expressed by restrictive endoneurial microvascular endothelium. Using Ulex Europaeus Agglutinin-1 expression to detect endothelial cells, we ascertained that the selected proteins were uniformly expressed in ≥90% of endoneurial microvessels. P-glycoprotein (also known as adenosine triphosphate-binding cassette subfamily B member 1) and solute carrier family 1 member 1 demonstrated restricted expression by endoneurial endothelium only, with classic tight junction protein claudin-5 also expressed on fenestrated epineurial macrovessels, and vascular-specific adherens junction protein cadherin-5 also expressed by the perineurium. The expression profiles of the selected proteins provide significant insight into the molecular composition of normal adult peripheral nerves. Further work is required to elucidate the human adult BNB molecular signature in order to better understand its development and devise strategies to restore function in peripheral neuropathies.

Background Although most severe peripheral nerve injuries result from high-speed mechanisms, there is no laboratory model to replicate this clinical condition. Objective To qualitatively and quantitatively describe microanatomical injury of rapid stretch. Methods The sciatic nerves of 36 Sprague-Dawley rats were subjected to rapid-stretch nerve injury, using fixed-direction strain produced via constrained weight drop applied to an intact nerve. Nerve injury severity was categorized by biomechanical parameters. Injury to nerve microarchitecture was quantified with serial longitudinal sectioning, with specific focus on the endoneurium, perineurium, and epineurium. Results Four grades of stretch injury severity were determined by mathematical cluster analysis: sham, elastic stretch, inelastic stretch, and stretch rupture. Two patterns of injury to endoneurial architecture were quantified: loss of fiber undulation (straightened fibers) and rupturing of individual fibers ("microruptures"). Straightening of nerve fibers was the earliest accommodation to stretch injury and accounted for elongation during elastic stretch. Microruptures were distributed along the length of the nerve and were more severe and involved greater volume of the nerve at higher biomechanical severity. Epineurium and perineurium disruption increased in frequency with progressive injury severity, yet did not predict transition from one injury grade to another (P = .3), nor was it a hallmark of severe injury. Conversely, accumulation of microruptures provided strong correlation to nerve injury severity (Pearson's R = .9897) and progression to mechanical failure. Conclusion Nerve architecture is injured in a graded fashion during stretch injury, which likely reflects tissue biomechanics. This study suggests new considerations in the theoretical framework of nerve stretch trauma.

Objective To review pathomorphological characteristics of the nerves of the palmar aponeurosis in Dupuytren’s contracture and develop a hypothesis of injury mechanisms. Material and methods The study included retrospective analysis of medical charts and surgical records of 123 patients with Dupuytren’s disease grades 2-4 who underwent partial aponeurectomy, light microscopic micrography of histological preparations of nerve trunks ofpalmar aponeurosis stained with hematoxylin and eosin. Results Three major types of nerve injury to palm aponeurosis were identified in Dupuytren’s disease including (1) active and residual perineuritis (41.6 %), (2) necrosis of endoneurial blood vessels and endoneurium (22%), and (3) fibromatosis of nerve sheath (37.4%). Higher rate of bilateral fascial fibromatosis and right-side involvement in unilateral fibromatosis (p less 0.05) was observed in Group 2 as compared to Group 3. The mean time interval of contracture formation was shorter in Group 3 by 21/2 years as compared to Groups 1 and 2 (p less 0.05). Discussion Perineurial lymphocyte and histiocyte infiltration was shown to be an additional criterion of fascial fibromatosis. Necrosis of endoneurial blood vessels was easily suspected in combination of bilateral fibromatosis (strong evidence for familial predisposition) and chronic hand injury of manual handling. Fibromatosis of nerve sheath indicated to aggressive course of the disease. Conclusion Pathomorphological characteristics of neuropathy in Dupuytren’s disease allowed assessment of fibromatosis and prognosis of the course of the disease in an individual patient to administer postoperative immunocorrective and neurally mediated therapy for optimization of wound healing and recurrence prevention.

Peripheral nerves are organ-like structures containing diverse cell types to optimize function. This interactive assembly includes mostly axon-associated Schwann cells, but also endothelial cells of supporting blood vessels, immune system-associated cells, barrier-forming cells of the perineurium surrounding and protecting nerve fascicles, and connective tissue-resident cells within the intra-fascicular endoneurium and inter-fascicular epineurium. We have established transcriptional profiles of mouse sciatic nerve-inhabitant cells to foster the fundamental understanding of peripheral nerves. To achieve this goal, we have combined bulk RNA sequencing of developing sciatic nerves up to the adult with focused bulk and single-cell RNA sequencing of Schwann cells throughout postnatal development, extended by single-cell transcriptome analysis of the full sciatic nerve both perinatally and in the adult. The results were merged in the transcriptome resource Sciatic Nerve ATlas (SNAT: https://www.snat.ethz.ch). We anticipate that insights gained from our multi-layered analysis will serve as valuable interactive reference point to guide future studies., eLife, 10, ISSN:2050-084X

A 31-year-old female patient presented with swelling on the anteromedial aspect of her right elbow with a sensory deficit in the median nerve territory. Biopsies led to the diagnosis of intraneural perineurioma (INP). Surgical excision was performed and followed by an intercalary graft. INP is a rare benign tumor of the peripheral nerves characterized by a multiplication of perineural cells invading the endoneurium. This lesion is often unknown, under-diagnosed, and its treatment is poorly defined. Few cases have been described in the literature; the majority involve the median nerve at the wrist and no references have been found about its localization to the median nerve at the elbow.

We studied telocytes/CD34+ stromal cells in the normal and pathological peripheral nervous system (PNS), for which we reviewed the literature and contributed our observations under light and electron microscopy in this field. We consider the following aspects: (A) general characteristics of telocytes and the terminology used for these cells (e.g., endoneurial stromal cells) in PNS; (B) the presence, characteristics and arrangement of telocytes in the normal PNS, including (i) nerve epi-perineurium and endoneurium (e.g., telopodes extending into the endoneurial space); (ii) sensory nerve endings (e.g., Meissner and Pacinian corpuscles, and neuromuscular spindles); (iii) ganglia; and (iv) the intestinal autonomic nervous system; (C) the telocytes in the pathologic PNS, encompassing (i) hyperplastic neurogenic processes (neurogenic hyperplasia of the appendix and gallbladder), highly demonstrative of telocyte characteristics and relations, (ii) PNS tumours, such as neurofibroma, schwannoma, granular cell tumour and nerve sheath myxoma, and interstitial cell of Cajal-related gastrointestinal stromal tumour (GIST), (iii) tumour-invaded nerves and (iv) traumatic, metabolic, degenerative or genetic neuropathies, in which there are fewer studies on telocytes, e.g., neuroinflammation and nerves in undescended testicles (cryptorchidism), Klinefelter syndrome, crush injury, mucopolysaccharidosis II (Hunter’s syndrome) and Charcot–Marie–Tooth disease.

The role of the plantar nerve in the pathogenesis of macrodactyly of the foot is unknown. We investigated the distribution of affected toes and forefoot in 27 feet of 26 patients with pedal macrodactyly, and how this relates to innervation of the affected plantar nerve. A preoperative ultrasound examination was performed to determine the diameter and structure of the plantar nerve. Histologic findings were recorded during surgery. The microstructure of affected plantar nerves was evaluated by hematoxylin-eosin staining, while S100 expression was assessed by immunofluorescence analysis. Phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA) gene mutation in the affected nerve tissue was detected by Sanger DNA sequencing. The affected toes and forefoot involved innervation of the medial plantar nerve in 25/27 feet, the lateral plantar nerve in one foot, and both medial and lateral plantar nerves in one foot. All affected plantar nerves, which were accompanied by a fatty strip, were surrounded by or infiltrated with fat. The affected plantar nerves showed enlargement, a tortuous course, fatty infiltration, or a combination of these. Pathologic changes in affected plantar nerves involved only the epineurium and not the perineurium or endoneurium. Expression of the Schwann cell marker S100 was absent in some areas of affected nerves. Sequencing of PIK3CA exons identified a gain-of-function mutation (p.His1047Arg) in affected plantar nerves. These results indicate that pathologic impairment of the plantar nerve can lead to macrodactyly of the foot, which may be considered as a nerve trunk disease.

Diabetic neuropathy has an incidence as high as 50% of diabetic patients and is characterized by damage to neurons, Schwann cells and blood vessels within the peripheral nervous system. The low-affinity neurotrophin receptor p75 (p75NTR ), particularly expressed by the Schwann cells in the peripheral nerve, has previously been reported to play a role in developmental myelination and cell survival/death. Increased levels of p75NTR , in the endoneurium and plasma from diabetic patients and rodent models of disease, have been observed, proposing that this receptor might be involved in the pathogenesis of diabetic neuropathy. Therefore, in this study, we addressed this hypothesis by utilizing a mouse model of selective nerve growth factor receptor (Ngfr) deletion in Schwann cells (SC-p75NTR -KO). Electron microscopy of sciatic nerves from mice with high fat diet induced obesity demonstrated how loss of Schwann cell-p75NTR aggravated axonal atrophy and loss of C-fibers. RNA sequencing disclosed several pre-clinical signaling alterations in the diabetic peripheral nerves, dependent on Schwann cell p75NTR signaling, specially related with lysosome, phagosome, and immune pathways. Morphological and biochemical analyses identified abundant lysosomes and autophagosomes in the C-fiber axoplasm of the diabetic SC-p75NTR -KO nerves, which together with increased Cathepsin B protein levels corroborates gene upregulation from the phagolysosomal pathways. Altogether, this study demonstrates that Schwann cell p75NTR deficiency amplifies diabetic neuropathy disease by triggering overactivation of immune-related pathways and increased lysosomal stress.

A highly regulated endoneurial microenvironment is required for normal axonal function in peripheral nerves and nerve roots, which structurally consist of an outer collagenous epineurium, inner perineurium consisting of multiple concentric layers of specialized epithelioid myofibroblasts that surround the innermost endoneurium, which consists of myelinated and unmyelinated axons embedded in a looser mesh of collagen fibers. Endoneurial homeostasis is achieved by tight junction-forming endoneurial microvessels that control ion, solute, water, nutrient, macromolecule and leukocyte influx and efflux between the bloodstream and endoneurium, and the innermost layers of the perineurium that control interstitial fluid component flux between the freely permeable epineurium and endoneurium. Strictly speaking, endoneurial microvascular endothelium should be considered the blood-nerve barrier (BNB) due to direct communication with circulating blood. The mammalian BNB is considered the second most restrictive vascular system after the blood-brain barrier (BBB) based on classic in situ permeability studies. Structural alterations in endoneurial microvessels or interactions with hematogenous leukocytes have been described in several human peripheral neuropathies; however major advances in BNB biology in health and disease have been limited over the past 50 years. Guided by transcriptome and proteome studies of normal and pathologic human peripheral nerves, purified primary and immortalized human endoneurial endothelial cells that form the BNB and leukocytes from patients with well-characterized peripheral neuropathies, validated by in situ or ex vivo protein expression studies, data are emerging on the molecular and functional characteristics of the human BNB in health and in specific peripheral neuropathies, as well as chronic neuropathic pain. These early advancements have the potential to not only increase our understanding of how the BNB works and adapts or fails to adapt to varying insult, but provide insights relevant to pathogenic leukocyte trafficking, with translational potential and specific therapeutic application for chronic peripheral neuropathies and neuropathic pain.

The authors identify two subsets of peripheral nerve macrophages residing in the endoneurium and the epineurium and displaying a distinct transcriptome and response to injury. These cells lack the main microglia identity and have a distinct origin. While CNS microglia have been extensively studied, relatively little is known about macrophages populating the peripheral nervous system. Here we performed ontogenic, transcriptomic and spatial characterization of sciatic nerve macrophages (snMacs). Using multiple fate-mapping systems, we show that snMacs do not derive from the early embryonic precursors colonizing the CNS, but originate primarily from late embryonic precursors and become replaced by bone-marrow-derived macrophages over time. Using single-cell transcriptomics, we identified a tissue-specific core signature of snMacs and two spatially separated snMacs: Relm alpha(+)Mgl1(+) snMacs in the epineurium and Relm alpha(-)Mgl1(-) snMacs in the endoneurium. Globally, snMacs lack most of the core signature genes of microglia, with only the endoneurial subset expressing a restricted number of these genes. In response to nerve injury, the two resident snMac populations respond differently. Moreover, and unlike in the CNS, monocyte-derived macrophages that develop during injury can engraft efficiently in the pool of resident peripheral nervous system macrophages.

A nerve is composed of axon (neural tissue) and connective tissue. Each axon in myelinated nerve fibers is surrounded by the endoneurium. Groups of nerve fibers form fascicles, surrounded by the perineurium, and groups of fascicles are surrounded by the internal and external epineurium. Impairments of the peripheral nervous system (PNS) lead to peripheral neuropathies that could be whether acquired (traumatic, drug toxicity, infection, etc.) or inherited, due to genetic abnormalities.

Recent anatomical discoveries indicate the importance of identifying membranes and compartments surrounding peripheral nerves into which local anesthetic agents can be injected and continuous nerve block catheters placed during regional anesthetic procedures. However, current markers used in anatomical studies have multiple drawbacks, specifically extravasation into noninjected locations, which can result in inadequate treatment. We studied a readily-available new marker, heparinized blood solution (HBS), which is easy to identify by microscopy and can remain in the nerve compartment into which it is deposited without distorting the tissue. We collected blood from 22 patients and prepared it as HBS. This was then injected into four fresh cadavers as in routine clinical practice for ultrasound-guided nerve blocks to form a so-called "doughnut" by "hydro-dissecting" at 32 sites. All samples, including nerves and neighboring tissues, were then prepared and examined by light microscopy. Although no deliberate intraneural injection was attempted, the marker was identified inside all the nerve compartments except the fascicles. Apart from leaking through the needle entry site in some instances, there was no extravasation of the HBS into neighboring nerve compartments in either direction. The tissues were not distorted and the erythrocytes did not form a thrombus. Nerve membranes and compartments could be clearly identified with routine staining. This technique enabled us to study the longitudinal and circumferential spread in all nerve compartments and to collect data for better interpretation of factors influencing an anesthetic nerve block and situations in which complications could possibly arise. HBS seemed superior to other markers because it did not leave the compartments into which it had been injected, did not distort the tissue, and was easily visible under the light microscope. Clin. Anat., 31:1050-1057, 2018. © 2018 Wiley Periodicals, Inc.

Heterotopic ossification (HO), or de novo bone formation in soft tissue, is often observed following traumatic injury. Recent studies suggest that peripheral nerves may play a key functional role in this process. The results supporting a neurological basis for HO are examined in this article. Evidence supports the fact that BMPs released from bone matrix possess the capacity to induce HO. However, the process cannot be recapitulated using recombinant proteins without extremely high doses suggesting other components are required for this process. Study of injuries that increase risk for HO, i.e. amputation, hip replacement, elbow fracture, burn, and CNS injury suggests that a likely candidate is traumatic injury of adjacent peripheral nerves. Recent studies suggest neuroinflammation may play a key functional role, by its ability to open the blood-nerve barrier (BNB). Barrier opening is characterized by a change in permeability and is experimentally assessed by the ability of Evans blue dye to enter the endoneurium of peripheral nerves. A combination of BMP and barrier opening is required to activate bone progenitors in the endoneurial compartment. This process is referred to as "neurogenic HO".

Objective The most common oral neurogenic lesions are traumatic neuroma, neurilemmoma, and neurofibroma. They share clinical and histological features, but have a different prognosis and treatment. Our aim was to use a small panel of immunohistochemistry antibodies to distinguish among them. Methods Eight cases of traumatic neuroma, 9 cases of neurilemmoma, and 11 cases of neurofibroma were selected and stained with primary antibodies against S-100 protein, laminin, and epithelial membrane antigen (EMA) using the streptavidin-biotin-peroxidase method of immunohistochemistry. Results The S-100 protein was positive in the endoneurium of traumatic neuroma, Schwann cells, and Verocay bodies of the neurilemmomas and revealed a variable number of positive cells amongst spindle-shaped cells that compose neurofibromas. Laminin was positive in the endoneurium and in the perineurium of traumatic neuromas. In neurilemmomas, all neoplasic cells and the capsule were positive for laminin. For neurofibromas, laminin positivity followed the patterns observed for S-100 protein. EMA was positive in the perineurium of traumatic neuroma and in the capsule of neurilemmomas. Most cases of neurofibroma were completely negative for EMA, but three cases showed rare, scattered positive cells. Conclusion The use of this small immunohistochemical panel to evaluate the presence and localization of S-100 protein, laminin, and EMA can help to distinguish among these three lesions and provide a more specific diagnosis. Consequently, this will provide a better prognosis for patients.

Familial amyloid polyneuropathy (FAP) is an autosomal dominant hereditary systemic amyloidosis caused by mutation of the transthyretin (TTR) gene, and usually shows sensory-dominant polyneuropathy and autonomic neuropathy at the initial stage. The pathogenesis of this neuropathy remains unknown, although several mechanisms, including mechanical compression, vessel occlusion, TTR toxicity and Schwann cell dysfunction have been proposed. We describe a patient with late-onset FAP caused by a TTR E61K mutation. Amyloid deposits were not detected in the endoneurium or perineurium of the sural nerve 7years after the onset of the disease, but a marked loss of nerve fibers was observed in the sural nerve. TTR-derived amyloid deposits were confirmed in the peroneus brevis muscle, salivary gland and heart tissue. DNA analysis revealed a heterozygous mutation in TTR. These findings suggest that proximal parts of the peripheral nervous system might be strongly affected by TTR aggregates or amyloid fibrils, and that the blood-nerve barrier in distal parts of peripheral nerves are initially preserved in this patient. This case indicates that several biopsy sites other than nerves may be helpful and necessary for the diagnosis of TTR amyloidosis in mild or late-onset FAP.