Peripheral nerve ischemia: apolipoprotein E deficiency results in impaired functional recovery and reduction of associated intraneural angiogenic response

Apolipoprotein E-knockout (apoE KO) mice have peripheral sensory nerve defects, reduced and delayed response to noxious thermal stimuli, abnormal morphology of unmyelinated fibers, and impaired blood-nerve and blood-brain barriers. In this study, we show that, compared to wild-type mice, peripheral nerves of apoE KO mice have impaired ability to respond to ischemia, as demonstrated by measurement of motor and sensory conduction velocity. In addition, mice lacking apoE exhibit a deficit of reinnervation of ischemic epidermis, evaluated by immunofluorescent staining for the pan-neuronal marker PGP 9.5. Also regional nerve blood flow, measured by laser Doppler, and intraneural angiogenesis after ischemia are significantly compromised in apoE-deficient mice. Finally, upregulation of the angiogenic cytokine vascular endothelial growth factor (VEGF), which physiologically occurs after ischemia in the peripheral nerve of wild-type mice, is severely impaired in apoE KO mice. Among the several neural defects that have already been described in mice lacking apoE, this is the first demonstration that functional recovery to ischemia is impaired in the peripheral nerves of these animals. This deficit is mirrored by the inability of upregulating VEGF and mounting an appropriate intraneural angiogenic response following injury. These findings provide new evidence of possible interdependent relationships between VEGF, angiogenesis, and nerve function and regeneration and may provide new important information on the role of apoE in the nervous system.     

The blood-nerve barrier in Wallerian degeneration: a sequential long-term study

The blood-nerve barrier (BNB) for serum proteins was studied after a crush lesion of the murine sciatic nerve or after transsection with persistent Wallerian degeneration. Using single intraperitoneal injections of biotinylated human albumin, transferrin, IgG, and complement components as tracers, the integrity of the BNB during degeneration and regeneration was determined over time. In Wallerian degeneration induced by crush the BNB became increasingly leaky, with a maximum in the distal stump 8 days after crush (i.e., during early regeneration). When regeneration potentials could first be elicited from the small foot muscles and when thinly myelinated nerve fibers were present, the BNB gradually regained its barrier function and was nearly intact on day 30 after crush. After transsection breakdown of the BNB persisted beyond 30 days. The BNB leakage may foster repair by allowing exchange of trophic factors of large molecular size during nerve regeneration.     

The mouse blood-brain barrier and blood-nerve barrier for IgG: A tracer study by use of the avidin-biotin system

Summary To study the permeability of the blood-brain barrier (BBB) and the blood-nerve barrier (BNB) for immunoglobulin G (IgG) we adapted the avidin-biotin system for postembedding demonstration of the tracer IgG in the central and peripheral nervous system (CNS, PNS). Normal mouse and human IgG were biotinylated and injected daily into the intraperitoneal (i.p.) space of adult BDF₁ mice. After 24 h, IgG was detected in blood vessels and in the interstitium of various organs, but staining was restricted to the dura mater in the CNS, to the spinal ganglia, and to the perineurium of peripheral nerves. After 4 days, IgG was also present in the endoneurial connective tissue of peripheral nerves, while the brain, spinal cord, and spinal roots remained free of IgG. Our results show a partial permeability of the normal mouse BNB for homologous and heterologous IgG.Supported by the Deutsche Forschungsgemeinschaft SFB 200, B 5 and C 1, and by the Ministerium MWF (North-Rhine Westphalia)     

Temporal relationship of blood-nerve barrier breakdown to the metabolic and morphologic alterations of tellurium neuropathy

The appearance of endoneurial edema early in the evolution of tellurium neuropathy raises the possibility that a breakdown of the blood-nerve barrier (BNB) plays a role in the pathogenesis of the tellurium-induced demyelination. To investigate this possibility, we correlated the temporal onset of breakdown of the BNB with inhibition of cholesterol synthesis and ultrastructural abnormalities in sciatic nerve of weanling Long-Evans rats fed a diet containing 1.1% elemental tellurium. Permeability of the BNB was assessed with [125I]-albumin and horseradish peroxidase (HRP); cholesterol synthesis was assessed by incubating segments of sciatic nerve in vitro with [1-14C]acetate. Cholesterol synthesis was severely inhibited and labeled squalene was accumulating in sciatic nerve at 12 hr of tellurium exposure. The permeability of the BNB progressively increased between 24 hr and 72 hr of tellurium exposure. Membrane-delimited vacuoles, lipid droplets and cytoplasmic excrescences appeared in myelinating Schwann cells at 24 hr; demyelinating axons appeared at 48 hr of tellurium exposure. These observations suggest that factors other than BNB breakdown and vasogenic endoneurial edema are responsible for the initial Schwann-cell injury in tellurium neuropathy. However, the early onset of BNB breakdown may have a synergistic role in the pathogenesis of tellurium-induced demyelination.     

Polyneuropathy in Waldenström s macroglobulinaemia. Passive transfer from man to mouse

ABSTRACT— To support the hypothesis of an immunopathogenesis of polyneuropathy in Waldenström's macroglobulinaemia (MW), serum IgM fractions of MW patients were applied intraperitoneally to mice for 17 days. Sections of liver, kidney, M. glutaeus maximus, central nervous system (CNS) and both Nn. ischiadici were examined for IgM, IgG, C3 and as control IgD with PAP-immunostaining. IgM deposits were found in every organ except the CNS. In peripheral nerves larger amounts were visualized in perineurium and endoneural space, whereas myelin lamellae and periaxon did not stain. Therefore, perhaps our investigation reveals a greater permeability of the blood-nerve barrier (BNB) compared with the blood-brain barrier (BBB). The involvement of the monoclonal IgM of MW, which has been shown to react in vitro with peripheral nerve constituents, appears possible in the pathogenesis of polyneuropathy.     

p21Cip1/WAF1 regulates radial axon growth and enhances motor functional recovery in the injured peripheral nervous system

Recent studies have provided evidence that p21Cip1/WAF1 has not only cell cycle-associated activities but also other biological activities like neurite elongation. To investigate the role of p21Cip1/WAF1 in the in vivo axonal regeneration in the peripheral nervous system, we developed a p21Cip1/WAF1 knockout (KO) mice sciatic nerve injury model. We performed quantitative assessments of the functional, histological, and electrophysiological recoveries after sciatic nerve injury in p21Cip1/WAF1 KO mice and compared the results with those of the wild-type mice. p21Cip1/WAF1 KO mice showed a significant delay of the motor functional recovery between 21 and 42 days after sciatic nerve injury. The values of motor conduction velocity in p21Cip1/WAF1 KO mice were significantly lower than those in the wild-type mice on postoperative day 28. The mean percent neural tissue and the mean nerve axon width of p21Cip1/WAF1 KO mice were significantly less than those of the wild-type mice, which was caused by hyperphosphorylation of neurofilaments. Therefore, p21Cip1/WAF1 was considered to be involved in radial axon growth and to be essential for the motor functional recovery following peripheral nervous system injury.     

Enhanced permeabilities of cationized-bovine serum albumins at the blood-nerve and blood-brain barriers in awake rats.

Permeability-surface area products (PAs) of the blood-nerve barrier (BNB) and blood-brain barrier (BBB) to 125I-labeled native bovine serum albumin (nBSA, pI approximately 4), and to 2 cationized albumins (cBSA) of differing pI (pI approximately 8 and 11), were quantitatively determined in awake rats, using an i.v. bolus injection technique. Mean PAs of the BNB and BBB to 125I-nBSA, after a circulation time of up to 120 min, were (0.17 +/- 0.23) and (0.09 +/- 0.05) x 10(-5) ml/s.g. wet wt, respectively (n = 12 rats), and were not significantly different from 0 (P greater than 0.05). Mean PAs of the BNB and BBB to 125I-cBSA (pI approximately 8), after circulation time of 12 min, were (1.9 +/- 0.1) and (1.7 +/- 0.1) x 10(-5) ml/s.g wet wt, respectively (n = 8). Significant greater PAs, at both the BNB and BBB to 125I-cBSA (pI approximately 11) [(8.2 +/- 1.8) and (3.0 +/- 0.6) x 10(-5) ml/s.g wet wt, respectively (n = 12)], than both PA's of nBSA and cBSA (pI approximately 8) were found. The accumulation of 125I-cBSA in epi-perineurial tissues also was higher than that of 125I-nBSA, and was related to the degree of cationization. Our results indicate that, as at the BBB, the transfer of cationized serum albumin is enhanced over that of native albumin at the BNB of the mammalian peripheral nerve.     

Leukaemia inhibitory factor is required for normal inflammatory responses to injury in the peripheral and central nervous systems in vivo and is chemotactic for macrophages in vitro

The cytokine leukaemia inhibitory factor (LIF) is up-regulated in glial cells after injury to the peripheral and central nervous systems. In addition, LIF is required for the changes in neuropeptide expression that normally occur when the axons of sympathetic and sensory neurons are transected. We investigated whether LIF is also necessary for the initial inflammatory response that follows mechanical injury to the sciatic nerve and cerebral cortex of adult mice. We find that inflammatory cell infiltration into crushed sciatic nerve is significantly slower in LIF knock-out (KO) mice compared with wild-type (WT) mice. Similarly, the microglial and astroglial responses to surgical injury of the cortex are significantly slower in LIF KO mice compared with WT mice. Consistent with these in vivo results, LIF is chemotactic for peritoneal macrophages in a microchamber culture assay. Thus, LIF is a key regulator of neural injury in vivo, where it is produced by glia and can act directly on neurons, glia and inflammatory cells. We also find that the initial inflammatory response to cortical injury is diminished in interleukin (IL)-6 KO mice. Surprisingly, however, the inflammatory response in LIF-IL-6 double KO mice is very similar to that of the single KO mice, suggesting that these cytokines may act in series rather than in parallel in this response.     

Blood-nerve barrier: structure and function

The blood-nerve barrier (BNB) is a dynamic interface between the endoneurial microenvironment and surrounding extracellular space or blood contents, and is localized the innermost layer of multilayered ensheathing perineurium and endoneurial microvessels. Since the BNB is a key structure controlling the internal milieu of the peripheral nerve parenchyma, adequate understanding of the BNB is crucial for developing treatment strategies for human peripheral nervous system disorders, including Guillain-Barré syndrome, chronic inflammatory demyelinating polyneuropathy, and diabetic and various metabolic/toxic neuropathies. However, fewer studies have been conducted on the BNB, if we compare against the number of studies on the blood-brain barrier. This is because of the lack of adequate human cell lines originating from the BNB. In our laboratory, human immortal cell lines from the BNB, namely, the endothelial cell line and pericyte cell line, have recently been established and vigorous investigations of their biological and physiological properties are now underway. Pericytes constituting the BNB were found to possess robust ability of controlling BNB integrity via secretion of various cytokines and growth factors including bFGF, VEGF, GDNF, BDNF, and angiopoietin-1. Unknown soluble factors secreted by pericytes also contribute to the upregulation of claudin-5 in endothelial cells in the BNB and thus, strengthen the barrier function of the BNB. In diabetic neuropathy, pericytes were shown to regulate the vascular basement membrane, while AGEs were shown to induce basement membrane hypertrophy and disrupt the BNB by increasing the autocrine secretion of VEGF and TGF-beta from pericytes. In this review article, we discuss the macroscopic and microscopic anatomy of the human BNB as well as the molecular mechanisms of mononuclear cell infiltration across the BNB.     

Reconstitution of the olfactory epithelium following injury in ApoE-deficient mice

ApoE, a protein component of lipoproteins, is extensively expressed in the primary olfactory pathway. Because apoE has been shown to play a vital role in nerve repair and remodeling, we hypothesized that apoE expression will increase in the injured olfactory epithelium (OE), and that apoE deficiency in apoE knockout (KO) mice will lead to delayed/incomplete reconstitution of the OE following injury. To directly test this hypothesis, we compared OE regeneration in wild-type (WT) and KO mice following injury induced by intranasal irrigation of Triton X-100. OE was collected at 0, 3, 7, 21, 42, and 56 days post lesion. The amount and distribution of apoE in the regenerating OE was measured by immunoblotting and immunohistochemistry. Rate of OE reconstitution in WT and KO mice was assessed by using three independent measures: (1) OE thickness was measured in cresyl-violet stained sections, (2) basal cell proliferation was determined by using bromodeoxyuridine (BrdU) staining, and (3) differentiation and maturation of olfactory sensory neurons were measured by immunoblotting and immunohistochemical analysis of growth associated protein (GAP) 43 and olfactory marker protein (OMP). The results revealed that apoE expression in the OE is highly regulated during the entire course of OE reconstitution post injury, and that apoE deficiency in apoE KO mice leads to delayed recovery of mature OMP⁺ cells in the reconstituting OE. The data suggest that apoE production increases in the injured OE to facilitate maturation of olfactory sensory neurons.     

Macrophage depletion alters the blood-nerve barrier without affecting Schwann cell function after neural injury

Previous work has shown that, during the early phases of chronic nerve compression (CNC) injury, axonal pathology is absent while Schwann cells undergo a dramatic process of cellular turnover with marked proliferation. It is known that macrophages may release Schwann cell mitogens, so we sought to explore the role of macrophages in CNC injury by selectively depleting the population of hematogenously derived macrophages in nerves undergoing CNC injury by injecting clodronate liposomes at days 1, 3, and 6 postinjury and evaluating both the integrity of the blood-nerve barrier (BNB) and Schwann cell function. Integrity of the BNB was evaluated by intravenously injecting Evans blue albumin (EBA), and Schwann cell number was determined via stereologic techniques. The BNB was clearly altered by 2 weeks postinjury and continued to disintegrate at later time points. Macrophage depletion attenuated this response at all observed time points. Quantification of Schwann cell nuclei in CNC nerves showed no differences between compressed sections of macrophage-depleted and nondepleted animals. Although macrophages are largely responsible for the increased vascular permeability associated with CNC injury, it is likely that the Schwann cell response to CNC injury is not influenced by macrophage-derived mitogenic signals but rather must be mediated via alternative mechanisms.     

Blood-nerve transfer of albumin and its implications for the endoneurial microenvironment

Blood-nerve transfer of plasma albumin was studied by measuring the permeability coefficient-surface area (PS) product of the blood-nerve barrier (BNB) to 125I-albumin in rat sciatic nerve using the i.v. bolus injection method. The calculated PS was 6.3 +/- 0.5 (S.E.M.) x 10(-7) ml.g-1.s-1. This value is smaller by more than an order of magnitude of that measured for sucrose and confirms the relative impermeance of the BNB to blood-borne solutes. From a review of the available evidence, it is concluded that normal blood-nerve exchange occurs predominantly across the endoneurial microvasculature, and the PS of the BNB reflects the permeability of capillaries to a greater extent than that of the perineurium. The only capillaries found to be less permeable than these are the cerebral capillaries. Proximo-distal differences (sciatic vs tibial) of the PS could not be detected. Blood-nerve albumin transfer was calculated at 1.2 mg.g-1.day-1, and the daily turnover of endoneurial albumin to be about 30%. It is postulated that small increases in PS of BNB to albumin lead to an elevation of endoneurial albumin concentration and, through the operation of Starling forces, subsequently produce endoneurial oedema. A major question posed by the results of this study is the identity of pathways for clearance of albumin and other macromolecules from the endoneurium.     

The distribution of serum albumin in rat peripheral nerve

To clarify the nature of the blood-nerve barrier (BNB) in excluding serum proteins from peripheral nerve endoneurium, we studied the ultrastructural distribution of albumin in peripheral nerve using light and electron microscopic immunocytochemistry. We found that serum albumin is commonly found within the endoneurium, but that it is not normally found intraaxonally. After subperineurial injection of labeled albumin, the albumin redistributed within intrafascicular clefts towards the perivascular spaces and the perineurium. These results suggest that, contrary to previous evidence, axons are exposed to serum proteins in normal nerves, and implies that they may be subject to interaction with abnormal proteins or altered serum proteins without any disruption of the BNB.     

Calcium transfer at the blood-nerve barrier of the frog sciatic nerve

Calcium transfer across the blood-nerve barrier of the frog sciatic nerve was studied using an in situ perfusion technique and an in vivo i.v. bolus injection technique. The permeability-surface area product of 45Ca at the blood-nerve barrier, (PA)BNB, calculated from radioactivity in the desheathed nerve segment after 5 min of circulation of tracer, and corrected for the residual radioactivity in the blood space, equaled 4.4 +/- 0.4 (S.E.M.) X 10(-5) ml.s-1.g-1 wet wt. The (PA)BNB of 45Ca was independent of [Ca2+] in the perfusion medium between 0.18 and 18 mM. The permeability-surface area products of 45Ca across the perineurium [(PA)per] also was measured by an in situ incubation technique, and equaled 1.45 +/- 0.41 X 10(-5) ml.s-1.g-1 wet wt. (n = 8). The half time (t 1/2) for nerve calcium to equilibrate with plasma calcium was calculated to be 60 min. The low, passive permeability to calcium of the blood-nerve barrier probably limits marked calcium concentration changes in nerve endoneurium following transient changes of plasma calcium, but should not alter steady-state responses.     

Blood-nerve barrier studies in experimental allergic neuritis

Summary The integrity of the blood-nerve barrier (BNB) was studied during the development of experimental allergic neuritis (EAN). Lewis rats immunized with bovine nerve or myelin plus complete Freund's adjuvant developed histological lesions of EAN in nerve roots by 10–12 days and in sciatic nerves by 12–14 days. Evans blue-albumin (EBA) and horseradish peroxidase (HRP) were injected i.v. 1 h prior to killing on days 6–18. Perivascular and diffuse endoneurial leakage of the tracers was seen in nerve roots by 10–12 days post immunization (p.i.) and in sciatic nerves by 12–14 days. This coincided with the appearance of endoneurial infiltration with inflammatory cells and endoneurial proteinaceous edema at a time when Schwann cell and myelin changes were still minimal. Therefore, an alteration in BNB permeability occurs early in EAN, coincident with inflammatory cell infiltration. This could be an expression of delayed hypersensitivity, yet it would also facilitate the entry of anti-myelin antibodies into the endoneurium where they could initiate demyelination.Supported by a research grant of the Muscular Dystrophy Association of CanadaA preliminary report was present at the American Association of Neuropathologists Meeting, San Diego, June 14–17, 1984     

Immunocytochemical studies of serum proteins and immunoglobulins in human sural nerve biopsies

Summary Post-embedding immunocytochemical studies on immunoglobulins (Ig) and other serum proteins were carried out on 38 human sural nerve biopsies using the PAP method. In addition to toxic, hereditary, metabolic, dysproteinemic, and vasculiticneuritic neuropathies, morphologically normal sural nerves were included as controls. The intensity of the immunocytochemical reactions was strong for proteins, such as IgG, the light chains of Igs, and albumin, but weak or absent for others like complement component C3, IgA, ceruloplasmin, and alpha-1-antitrypsin (AAT) in normal nerve biopsies and in all pathologic groups. IgG, the light chains of immunoglobulins, and albumin could readily be detected in perineurium, endoneurial interstitium, and blood vessel walls. IgM, C3, and beta-lipoprotein (BLP) were largely confined to the walls of blood vessels and perineurium, thus indicating that they do not penetrate the blood nerve barrier. Only in a few cases, in vasculitic-neuritic and dysproteinemic neuropathies, staining of the endoneurial intersitium for IgM and C3 was observed. Increased staining for the corresponding heavy or light chains was not detected in the endoneurium in any of the neuropathies associated with gammopathy.The results stress that PAP immunocytochemistry is suitable for studying the blood-nerve barrier (BNB) and provides new aspects to the concept of the BNB with respect to the steady state of serum proteins between endoneurial and vascular spaces. It is suggested that, in addition to serum concentration and molecular weight of serum proteins, the permeability of the BNB is influenced by other yet undefined factors.     

Delayed olfactory nerve regeneration in ApoE-deficient mice

Apolipoprotein E (apoE), a lipid transporting protein, is extensively expressed in the primary olfactory pathway, but its function is unknown. We previously reported increased apoE levels in the olfactory bulb (OB) following olfactory epithelium (OE) lesion in mice, and hypothesized that apoE may play a vital role in olfactory nerve (ON) regeneration. To directly test this hypothesis, we examined the rate of ON regeneration following OE lesion in apoE deficient/knockout (KO) and wild-type (WT) mice. OE was lesioned in 2- to 3-month-old mice by intranasal irrigation with Triton X-100 (TX). OB were collected at 0, 3, 7, 21, 42, and 56 days post-lesion. OB recovery was measured by both immunoblotting and immunohistochemical analysis of growth cone associated protein (GAP) 43 and olfactory marker protein (OMP). The results revealed that (1) OMP recovery in the OB was significantly slower in apoE KO compared to WT mice; (2) recovery of glomerular area was similarly slower; and (3) GAP43 increases and return to prelesion levels in the OB were slower in KO mice. Together, these results show that olfactory nerve regeneration is significantly slower in KO mice as compared to WT mice, suggesting apoE facilitates olfactory nerve regeneration.     

Restoration of blood-nerve barrier in neuropathy is associated with axonal regeneration and remyelination.

We investigated the temporal course of blood-nerve barrier (BNB) breakdown during the evolution of tellurium neuropathy, ricin neuropathy, and Wallerian degeneration following nerve transection or nerve crush. Blood-nerve barrier permeability was assessed with a 4,000-molecular weight fluoresceinated dextran from three days to 19 weeks after onset of neuropathy. Blood-nerve barrier breakdown was present during the first two weeks in all four models of neuropathy. Restoration of the BNB to the dextran began within four weeks and was complete by 14 weeks in tellurium neuropathy, a model of demyelinating neuropathy characterized by rapid remyelination, and after nerve crush, a model of Wallerian degeneration characterized by rapid axonal regeneration into distal stump. In contrast, there was persistence of BNB breakdown beyond 14 weeks in ricin neuropathy, a model of neuropathy with no axonal regeneration or remyelination, and after nerve transection, a model of Wallerian degeneration characterized by minimal axonal regeneration into distal stump. We conclude from these data that alterations in the BNB over the course of neuropathy differ among various types of neuropathy, and that these alterations are dependent on the form of nerve fiber injury. The lack of regenerating or remyelinating axons in ricin neuropathy and after nerve transection may be responsible for the persistent BNB breakdown found in these neuropathies.     

The recovery of blood-nerve barrier in crush nerve injury--a quantitative analysis utilizing immunohistochemistry

The purpose of this study is to reveal whether the application of immunohistochemical examinations to the peripheral nervous system (PNS) can be a reliable method for the quantitative analysis of the blood-nerve barrier (BNB) and the relationship between restoration of BNB and nerve regeneration. Sciatic nerves in rats were examined after nerve crush. Immunohistochemical staining with anti-rat endothelial cell antigen-1 (anti-RECA-1) that recognizes endothelial cells and anti-endothelial barrier antigen (anti-EBA) for the detection of barrier-type endothelial cells were used. Neurofilament for staining axons was also performed. A quantitative analysis of the BNB was assessed using the ratio of EBA positive cells and RECA-1 positive cells. The ratio of EBA/RECA-1 decreased significantly 3 days postoperatively and reached its lowest level at day 7 in the segment 5 mm proximal and the entire distal stump. The ratio gradually recovered from the proximal and the regeneration of axons started a week earlier than BNB. The ratio of EBA/RECA-1 applied to the PNS can be a reliable method for the quantitative analysis of BNB. In crush injuries, the breakdown of BNB occurred simultaneously in the segment 5 mm proximal and the entire distal stump; restoration began from the proximal to distal and followed a week later to nerve regeneration.     

Dicer-microRNA pathway is critical for peripheral nerve regeneration and functional recovery in vivo and regenerative axonogenesis in vitro

Both central and peripheral axons contain pivotal microRNA (miRNA) proteins. While recent observations demonstrated that miRNA biosynthetic machinery responds to peripheral nerve lesion in an injury-regulated pattern, the physiological significance of this phenomenon remains to be elucidated. In the current paper we hypothesized that deletion of Dicer would disrupt production of Dicer-dependent miRNAs and would negatively impact regenerative axon growth. Taking advantage of tamoxifen-inducible CAG-CreERt:Dicer(fl/fl) knockout (Dicer KO), we investigated the results of Dicer deletion on sciatic nerve regeneration in vivo and regenerative axon growth in vitro. Here we show that the sciatic functional index, an indicator of functional recovery, was significantly lower in Dicer KO mice in comparison to wild-type animals. Restoration of mechanical sensitivity recorded in the von Frey test was also markedly impaired in Dicer mutants. Further, Dicer deletion impeded the recovery of nerve conduction velocity and amplitude of evoked compound action potentials in vitro. Histologically, both total number of regenerating nerve fibers and mean axonal area were notably smaller in the Dicer KO mice. In addition, Dicer-deficient neurons failed to regenerate axons in dissociated dorsal root ganglia (DRG) cultures. Taken together, our results demonstrate that knockout of Dicer clearly impedes regenerative axon growth as well as anatomical, physiological and functional recovery. Our data suggest that the intact Dicer-dependent miRNA pathway is critical for the successful peripheral nerve regeneration after injury.