Tyrosine-derived polycarbonate nerve guidance tubes elicit proregenerative extracellular matrix deposition when used to bridge segmental nerve defects in swine.

Promising biomaterials should be tested in appropriate large animal models that recapitulate human inflammatory and regenerative responses. Previous studies have shown tyrosine-derived polycarbonates (TyrPC) are versatile biomaterials with a wide range of applications across multiple disciplines. The library of TyrPC has been well studied and consists of thousands of polymer compositions with tunable mechanical characteristics and degradation and resorption rates that are useful for nerve guidance tubes (NGTs). NGTs made of different TyrPCs have been used in segmental nerve defect models in small animals. The current study is an extension of this work and evaluates NGTs made using two different TyrPC compositions in a 1 cm porcine peripheral nerve repair model. We first evaluated a nondegradable TyrPC formulation, demonstrating proof-of-concept chronic regenerative efficacy up to 6 months with similar nerve/muscle electrophysiology and morphometry to the autograft repair control. Next, we characterized the acute regenerative response using a degradable TyrPC formulation. After 2 weeks in vivo, TyrPC NGT promoted greater deposition of pro-regenerative extracellular matrix (ECM) constituents (in particular collagen I, collagen III, collagen IV, laminin, and fibronectin) compared to commercially available collagen-based NGTs. This corresponded with dense Schwann cell infiltration and axon extension across the lumen. These findings confirmed results reported previously in a mouse model and reveal that TyrPC NGTs were well tolerated in swine and facilitated host axon regeneration and Schwann cell infiltration in the acute phase across segmental defects - likely by eliciting a favorable neurotrophic ECM milieu. This regenerative response ultimately can contribute to functional recovery.

Immediate or Delayed Transplantation of a Vein Conduit Filled with Nasal Olfactory Stem Cells Improves Locomotion and Axogenesis in Rats after a Peroneal Nerve Loss of Substance.

Over the recent years, several methods have been experienced to repair injured peripheral nerves. Among investigated strategies, the use of natural or synthetic conduits was validated for clinical application. In this study, we assessed the therapeutic potential of vein guides, transplanted immediately or two weeks after a peroneal nerve injury and filled with olfactory ecto-mesenchymal stem cells (OEMSC). Rats were randomly allocated to five groups. A3 mm peroneal nerve loss was bridged, acutely or chronically, with a 1 cm long femoral vein and with/without OEMSCs. These four groups were compared to unoperated rats (Control group). OEMSCs were purified from male olfactory mucosae and grafted into female hosts. Three months after surgery, nerve repair was analyzed by measuring locomotor function, mechanical muscle properties, muscle mass, axon number, and myelination. We observed that stem cells significantly (i) increased locomotor recovery, (ii) partially maintained the contractile phenotype of the target muscle, and (iii) augmented the number of growing axons. OEMSCs remained in the nerve and did not migrate in other organs. These results open the way for a phase I/IIa clinical trial based on the autologous engraftment of OEMSCs in patients with a nerve injury, especially those with neglected wounds.

Vibration-induced depression in spinal loop excitability revisited.

KEY POINTS: While it has been well described that prolonged vibration locally applied to a muscle or its tendon (up to 1 h) decreases spinal loop excitability between homonymous Ia afferents and motoneurons, the involved mechanisms are not fully understood. By combining electrophysiological methods, this study aimed to provide new insights into the mechanisms involved in soleus decreased spinal excitability after prolonged local vibration. We report that prolonged vibration induces a decrease in motoneuron excitability rather than an increase in presynaptic mechanisms (as commonly hypothesized in the current literature). The present results may help to design appropriate clinical intervention and could reinforce the interest in vibration as a treatment for spastic patients who are characterized by spinal hyper-excitability responsible for spasms and long-lasting reflexes. ABSTRACT: The mechanisms that can explain the decreased spinal loop excitability in response to prolonged local vibration (LV), as assessed by the H-reflex, remain to be precisely determined. This study provides new insights into how prolonged Achilles' tendon LV (30 min, 100 Hz) acutely interacts with the spinal circuitry. The roles of presynaptic inhibition exerted on Ia afferents (Experiment A, n = 15), neurotransmitter release at the synapse level (Experiment B, n = 11) and motoneuron excitability (Experiment C, n = 11) were investigated in soleus. Modulation of presynaptic inhibition was assessed by conditioning the soleus H-reflex (tibial nerve electrical stimulation) with fibular nerve (D1 inhibition) and femoral nerve (heteronymous facilitation, HF) electrical stimulations. Potential vibration-induced changes in neurotransmitter depletion at the Ia afferent terminals was assessed through paired stimulations applied over the tibial nerve (HD). Intrinsic motoneuron excitability was assessed with thoracic motor evoked potentials (TMEPs) in response to electrical stimulation over the thoracic spine. Non-conditioned H-reflex was depressed by ∼60% after LV (P < 0.001), while D1 and HF H-reflexes increased by ∼75% after LV (P = 0.03 and 0.06, respectively). In Experiment B, HD remained unchanged after LV (P = 0.80). In Experiment C, TMEPs were reduced by ∼13% after LV (P = 0.01). Overall, presynaptic mechanisms do not seem to be involved in the depression of spinal excitability after LV. It rather seems to rely, at least in part, on a decrease in intrinsic motoneuron excitability. These results may have implications in reducing spinal hyper-excitability in spastic patients.

Thin-fibre receptors expressing acid-sensing ion channel 3 contribute to muscular mechanical hypersensitivity after exercise.

BACKGROUND: Delayed onset muscle soreness (DOMS) is characterized by mechanical hyperalgesia after lengthening contractions (LC). It is relatively common and causes disturbance for many people who require continuous exercise, yet its molecular and peripheral neural mechanisms are poorly understood. METHODS: We examined whether muscular myelinated Aδ-fibres, in addition to unmyelinated C-fibres, are involved in LC-induced mechanical hypersensitivity, and whether acid-sensing ion channel (ASIC)-3 expressed in thin-fibre afferents contributes to this type of pain using a rat model of DOMS. The peripheral contribution of ASIC3 was investigated using single-fibre electrophysiological recordings in extensor digitorum longus muscle-peroneal nerve preparations in vitro. RESULTS: Behavioural tests demonstrated a significant decrease of the muscular mechanical withdrawal threshold following LC to ankle extensor muscles, and it was improved by intramuscular injection of APETx2 (2.2 µM), a selective blocker of ASIC3. The lower concentration of APETx2 (0.22 µM) and its vehicle had no effect on the threshold. Intramuscular injection of APETx2 (2.2 µM) in naïve rats without LC did not affect the withdrawal threshold. In the ankle extensor muscles that underwent LC one day before the electrophysiological recordings, the mechanical response of Aδ- and C-fibres was significantly facilitated (i.e. decreased response threshold and increased magnitude of the response). The facilitated mechanical response of the Aδ- and C-fibres was significantly suppressed by selective blockade of ASIC3 with APETx2, but not by its vehicle. CONCLUSIONS: These results clearly indicate that ASIC3 contributes to the augmented mechanical response of muscle thin-fibre receptors in delayed onset muscular mechanical hypersensitivity after LC. SIGNIFICANCE: Here, we show that not only C- but also Aδ-fibre nociceptors in the muscle are involved in mechanical hypersensitivity after lengthening contractions, and that acid-sensing ion channel (ASIC)-3 expressed in the thin-fibre nociceptors is responsible for the mechanical hypersensitivity. ASIC3 might be a novel pharmacological target for pain after exercise.

Optogenetic Peripheral Nerve Immunogenicity.

Optogenetic technologies have been the subject of great excitement within the scientific community for their ability to demystify complex neurophysiological pathways in the central (CNS) and peripheral nervous systems (PNS). The excitement surrounding optogenetics has also extended to the clinic with a trial for ChR2 in the treatment of retinitis pigmentosa currently underway and additional trials anticipated for the near future. In this work, we identify the cause of loss-of-expression in response to transdermal illumination of an optogenetically active peroneal nerve following an anterior compartment (AC) injection of AAV6-hSyn-ChR2(H134R) with and without a fluorescent reporter. Using Sprague Dawley Rag2-/- rats and appropriate controls, we discover optogenetic loss-of-expression is chiefly elicited by ChR2-mediated immunogenicity in the spinal cord, resulting in both CNS motor neuron death and ipsilateral muscle atrophy in both low and high Adeno-Associated Virus (AAV) dosages. We further employ pharmacological immunosuppression using a slow-release tacrolimus pellet to demonstrate sustained transdermal optogenetic expression up to 12 weeks. These results suggest that all dosages of AAV-mediated optogenetic expression within the PNS may be unsafe. Clinical optogenetics for both PNS and CNS applications should take extreme caution when employing opsins to treat disease and may require concurrent immunosuppression. Future work in optogenetics should focus on designing opsins with lesser immunogenicity.

Molecular Mechanism of the "Babysitter" Procedure for Nerve Regeneration and Muscle Preservation in Peripheral Nerve Repair in a Rat Model.

OBJECTIVE: To investigate the molecular mechanism of nerve "babysitter" for nerve regeneration and muscle preservation in peripheral nerve repair. METHODS: Eighty rats were equalized into 4 groups: peroneal nerve transected, group A received no treatment; group B underwent end-to-end repair; group C underwent end-to-side "babysitter" with donor epineurial window; group D underwent end-to-side "babysitter" with 40% donor neurectomy. During second-stage procedure, end-to-end neurorrhaphies were executed in groups A, C, and D. Expression of Insulin-like growth factor (IGF)-1 in spinal cord and IGF-1, TNF-like weak inducer of apoptosis (TWEAK), and Fn14 in anterior tibial muscles were evaluated by histopathology at 4-, 8-, 12-, and 24-week timepoints postoperatively. RESULTS: At 4 weeks, group D expressed comparable IGF-1 with group B, and greater value than groups A and C in spinal cord. By 24 weeks, groups B and D showed higher values than groups A and C. Insulin-like growth factor 1 in muscles were greater in groups C and D than in groups A and B at 4 weeks, and comparable in all groups at 24 weeks. At 4 weeks, immunoreactive scores of TWEAK were 9.00 ± 0, 3.00 ± 0, 6.75 ± 0.75, and 6.75 ± 0.75, respectively. No differences were noticed in all groups by 24 weeks. At 4 weeks, Fn14 were similar in groups A, C, and D, but lower in group B. Group D showed comparable Fn14 with groups B and C, but lower value than group A at 24 weeks. CONCLUSIONS: End-to-side nerve "babysitter" in peripheral nerve could promote fiber regeneration and muscle preservation by regulating expression of IGF-1 and TWEAK-Fn14. End-to-side "babysitter" with partial donor neurectomy could achieve comparable effects with end-to-end repair.

Changes in VGLUT1 and VGLUT2 expression in rat dorsal root ganglia and spinal cord following spared nerve injury.

Disturbance of glutamate homeostasis is a well-characterized mechanism of neuropathic pain. Vesicular glutamate transporters (VGLUTs) determine glutamate accumulation in synaptic vesicles and their roles in neuropathic pain have been suggested by gene-knockout studies. Here, we investigated the spatio-temporal changes in VGLUT expression during the development of neuropathic pain in wild-type rats. Spared nerve injury (SNI) induced mechanical allodynia from postoperative day 1 to at least day 14. Expression of VGLUT1 and VGLUT2 in dorsal root ganglia and spinal cord was examined by western blot analyses on different postoperative days. We observed that VGLUT2 were selectively upregulated in crude vesicle fractions from the ipsilateral lumbar enlargement on postoperative days 7 and 14, while VGLUT1 was transiently downregulated in ipsilateral DRG (day 4) and contralateral lumbar enlargement (day 1). Upregulation of VGLUT2 was not accompanied by alterations in vesicular expression of synaptotagmin or glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Thus, VGLUTs expression, especially VGLUT2, is regulated following peripheral nerve injury. Temporal regulation of VGLUT2 expression in spinal cord may represent a novel presynaptic mechanism contributing to injury-induced glutamate imbalance and associated neuropathic pain.

A clinical and neuropathological study of Chinese patients with diabetic peripheral neuropathy.

OBJECTIVE: To examine whether the neuropathological and metabolic changes of peripheral nerves are correlated to clinical features in diabetes mellitus type 2 patients with peripheral neuropathy. METHODS: 147 type 2 diabetic patients with signs/symptoms of diabetic peripheralneuropathy (DPN) aged 53.4 ± 12.3 years and 134 healthy volunteers aged 55.5 ± 11.7 years were investigated for fasting plasma glucose (FPG), hemoglobin A1C (HbA1c), and red blood cell sorbitol (RBC sorbitol) in addition to nerve conduction velocity (NCV). Among the 147 diabetic patients, 10 patients underwent superficial peroneal nerve biopsy for light and electron microscopy. RESULTS: In the experimental group, the levels of HbA1c and RBC sorbitol showed significant increase compared with the controlled group, whereas motor nerve conduction velocity (MNCV) and sensory nerve conduction velocity (SNCV) both showed decline and SNCV decreased at a greater extent. Morphologically, there were various degrees of nerve fiber loss, associated with axon degeneration and capillary luminal narrowing in 10 patients undergone nerve biopsy. CONCLUSION: The metabolic change of sorbitol, the consequently observed changes in NCV and histopathology of peripheral nerves are positively correlated with the duration of diabetes and overall level of blood glucose.

An experimental study to determine and correlate choline acetyltransferase assay with functional muscle testing after nerve injury.

OBJECT Choline acetyltransferase (ChAT) is an enzyme synthesized within the body of a motor neuron whose role is to form the neurotransmitter acetylcholine. Quantification of ChAT levels in motor or mixed nerves has been proposed to provide information regarding the viability of a proximal nerve stump for motor neurotization following brachial plexus injury. To do so requires information regarding normal ChAT levels and those in injured nerves, as well as the correlation of ChAT level determined at surgery with eventual motor recovery. The purpose of this study was to determine ChAT activity in the normal and injured sciatic/peroneal nerve in a rat model, evaluate the correlation between ChAT and motor recovery, find the relationship between ChAT activity and isometric muscle force, and elucidate the parallel between ChAT activity and acetylcholinesterase (AChE) activity. METHODS Sixty animals were divided into 3 groups. The sciatic nerves in Group 1 were transected without repair. Nerves in Group 2 were transected and repaired. Nerves in Group 3 sustained a crush injury followed by transection and reconstruction. All animals were allowed 12 weeks of recovery followed by evaluation of ChAT levels in the peroneal nerve, correlated with measures of maximal isometric tibialis anterior muscle force and muscle weight (the operated side normalized to the control side). Karnovsky AChE staining of peroneal nerve segments was also compared with radiochemical assay of ChAT activity in the same nerve. RESULTS A significant difference in the tibialis anterior isometric tetanic force and the tibialis anterior muscle weight index (TAMI) was noted between Group 1 and Groups 2 and 3 (p < 0.0001); no significant difference was found comparing Group 2 with Group 3. The correlation between the force measurement and the TAMI was 0.382. Both AChE measurement and ChAT activity demonstrated significantly fewer fibers in the operated nerve compared with the contralateral nerve. Intergroup variability could also be illustrated using these tests. The correlation coefficient between the isometric tetanic force measurement and the ChAT analysis in Groups 1 and 2 was 0.468. The correlation for the AChE staining and the isometric tetanic force measurement was 0.111. The correlation between the TAMI and the ChAT levels was 0.773. The correlation between the TAMI and the AChE-stained fibers was 0.640. Correlating AChE staining to the ChAT analysis produced a correlation of 0.712. CONCLUSIONS The great variability in all groups and weak correlations to the functional muscle assessments and the ChAT radiochemical assay made this technique an unreliable method of determining motor nerve viability.

Hormone phase dependency of neural responses to chemoreflex-driven sympathoexcitation in young women using hormonal contraceptives.

Hormone fluctuations in women may influence muscle sympathetic nerve activity (MSNA) in a manner dependent on the severity of the sympathoexcitatory stimulus. This study examined MSNA patterns at rest and during chemoreflex stimulation in low- (LH) vs. high-hormone (HH) phases of contraceptive use in healthy young women (n = 7). We tested the hypothesis that MSNA would be greater in the HH phase at baseline and in response to chemoreflex stimulation. MSNA recordings were obtained through microneurography in LH and HH at baseline, during rebreathing causing progressive hypoxia and hypercapnia, and during a hypercapnic-hypoxic end-inspiratory apnea. Baseline MSNA burst incidence (P = 0.03) and burst frequency (P = 0.02) were greater in the HH phase, while MSNA burst amplitude distributions and hemodynamic measures were similar between phases. Rebreathing elicited increases in all MSNA characteristics from baseline (P < 0.05), but was not associated with hormone phase-dependent changes to MSNA patterns. Apnea data were considered in two halves, both of which caused large increases in all MSNA variables from baseline in each hormone phase (P < 0.01). Increases in burst incidence and frequency were greater in LH during the first half of the apnea (P = 0.03 and P = 0.02, respectively), while increases in burst amplitude and total MSNA were greater in LH during the second half of the apnea (P < 0.05). These results indicate that change in hormone phase brought on through use of hormonal contraceptives influences MSNA patterns such that baseline MSNA is greater in the HH phase, but responses to severe chemoreflex stimulation are greater in the LH phase.

Immunohistochemical, ultrastructural and functional analysis of axonal regeneration through peripheral nerve grafts containing Schwann cells expressing BDNF, CNTF or NT3.

We used morphological, immunohistochemical and functional assessments to determine the impact of genetically-modified peripheral nerve (PN) grafts on axonal regeneration after injury. Grafts were assembled from acellular nerve sheaths repopulated ex vivo with Schwann cells (SCs) modified to express brain-derived neurotrophic factor (BDNF), a secretable form of ciliary neurotrophic factor (CNTF), or neurotrophin-3 (NT3). Grafts were used to repair unilateral 1 cm defects in rat peroneal nerves and 10 weeks later outcomes were compared to normal nerves and various controls: autografts, acellular grafts and grafts with unmodified SCs. The number of regenerated ßIII-Tubulin positive axons was similar in all grafts with the exception of CNTF, which contained the fewest immunostained axons. There were significantly lower fiber counts in acellular, untransduced SC and NT3 groups using a PanNF antibody, suggesting a paucity of large caliber axons. In addition, NT3 grafts contained the greatest number of sensory fibres, identified with either IB4 or CGRP markers. Examination of semi- and ultra-thin sections revealed heterogeneous graft morphologies, particularly in BDNF and NT3 grafts in which the fascicular organization was pronounced. Unmyelinated axons were loosely organized in numerous Remak bundles in NT3 grafts, while the BDNF graft group displayed the lowest ratio of umyelinated to myelinated axons. Gait analysis revealed that stance width was increased in rats with CNTF and NT3 grafts, and step length involving the injured left hindlimb was significantly greater in NT3 grafted rats, suggesting enhanced sensory sensitivity in these animals. In summary, the selective expression of BDNF, CNTF or NT3 by genetically modified SCs had differential effects on PN graft morphology, the number and type of regenerating axons, myelination, and locomotor function.

Inhibition of augmented muscle vasoconstrictor drive following asphyxic apnoea in awake human subjects is not affected by relief of chemical drive.

Progressive asphyxia, produced by a prolonged voluntary breath hold (end-expiratory apnoea), evokes large bursts of muscle sympathetic nerve activity (MSNA). These bursts increase in amplitude until the asphyxic break point is reached, at which point the bursts are inhibited. We tested the hypothesis that lung inflation, rather than relief from hypoxia and hypercapnia, is responsible for the inhibition of MSNA. Multiunit MSNA was recorded from motor fascicles of the common peroneal nerve in 11 subjects. Following a period of quiet breathing, subjects were instructed to behave as follows: (i) to hold their breath in expiration for as long as they could (mean duration 32.3 ± 1.9 s); (ii) to take a single breath of room air, 100% N(2) or 10% CO(2) + 90% N(2) at the asphyxic break point; (iii) to exhale and continue the apnoea until the next break point; and then (iv) to resume breathing. All subjects reported relief during inhalation of any gas, and could continue holding their breath for a further 30.7 ± 2.8 s with room air, 18.6 ± 1.7 s with N(2) and 11.8 ± 1.8 s with 10% CO(2) + 90% N(2). Despite the exaggerated chemoreceptor drive in the latter two conditions (hence the significantly shorter latencies to the subsequent asphyxic break point), the inhibition still occurred; moreover, there was no significant difference in duration of the inhibition of MSNA following the single breath of room air (7.6 ± 0.7 s), N(2) (6.2 ± 0.6 s) or 10% CO(2) + 90% N(2) (5.5 ± 0.4 s). Following the resumption of breathing, however, the duration of MSNA inhibition (11.0 ± 1.0 s) was significantly longer than that following a single breath. To investigate the involvement of chemoreceptors in the respiratory modulation of MSNA further, the same gases were used during an inspiratory-capacity apnoea, which causes a brief inhibition of MSNA during the inflation phase and a sustained increase during the hold phase. The duration of the apnoea was shortest after a breath of 10% O(2) + 90% N(2), but the latency until the bursts resumed after the inspiratory breath hold were similar for all gases, which suggests that there is no chemoreceptor involvement during the sympathetic silence seen during the inflation phase of inspiratory-capacity apnoeas. We conclude that neither peripheral nor central chemoreceptors are responsible for the inhibition of muscle vasoconstrictor drive following an end-expiratory apnoea or an end-inspiratory apnoea. Rather, we suggest that the inhibition is evoked by stretch receptors in the lungs and/or chest wall, which may also contribute to the longer inhibition associated with the hyperventilation following the subsequent resumption of rhythmic breathing.

Modulation of nerve fibers in the rat thyroarytenoid muscle following recurrent laryngeal nerve injury.

CONCLUSION: Regeneration of nerve fibers in the thyroarytenoid (TA) muscle occurred actively after damage on the recurrent laryngeal nerve (RLN) compared with the vagus nerve (VN). However, remyelination did not occur after damage on the RLN. OBJECTIVES: To determine the regeneration process of nerve fibers in the TA muscle following transection and immediate anastomosis of the RLN or VN. METHODS: Three types of animal model were prepared: an RLN anastomosis model (RLNa), a VN anastomosis model (VNa), and a peroneal nerve anastomosis model (PNa). Animals were sacrificed at five time points following the procedure. The modulation of axons, myelin sheaths, Schwann cells (SCs), nerve terminals (NTs), and acetylcholine receptors (AchRs) in the TA or tibialis anterior muscles was examined by immunohistochemical analysis. The ratios of the expression areas in axons, myelin sheaths, and SCs, and the number of NTs and AchRs in the treated (T) and untreated (U) sides (T/U) were evaluated. RESULTS: At 18 weeks, the T/U ratios of expression in RLNa, VNa, and PNa were 68.5, 0, and 100.4%, respectively, in axons; 0, 0, and 97.6% in myelin sheaths; 53.7, 0, and 93.6% in SCs; 61.0, 0, and 96.4% in NTs; and 99.4, 67.0, and 101.2% in AchRs.

RhoE deficiency produces postnatal lethality, profound motor deficits and neurodevelopmental delay in mice.

Rnd proteins are a subfamily of Rho GTPases involved in the control of actin cytoskeleton dynamics and other cell functions such as motility, proliferation and survival. Unlike other members of the Rho family, Rnd proteins lack GTPase activity and therefore remain constitutively active. We have recently described that RhoE/Rnd3 is expressed in the Central Nervous System and that it has a role in promoting neurite formation. Despite their possible relevance during development, the role of Rnd proteins in vivo is not known. To get insight into the in vivo function of RhoE we have generated mice lacking RhoE expression by an exon trapping cassette. RhoE null mice (RhoE gt/gt) are smaller at birth, display growth retardation and early postnatal death since only half of RhoE gt/gt mice survive beyond postnatal day (PD) 15 and 100% are dead by PD 29. RhoE gt/gt mice show an abnormal body position with profound motor impairment and impaired performance in most neurobehavioral tests. Null mutant mice are hypoactive, show an immature locomotor pattern and display a significant delay in the appearance of the hindlimb mature responses. Moreover, they perform worse than the control littermates in the wire suspension, vertical climbing and clinging, righting reflex and negative geotaxis tests. Also, RhoE ablation results in a delay of neuromuscular maturation and in a reduction in the number of spinal motor neurons. Finally, RhoE gt/gt mice lack the common peroneal nerve and, consequently, show a complete atrophy of the target muscles. This is the first model to study the in vivo functions of a member of the Rnd subfamily of proteins, revealing the important role of Rnd3/RhoE in the normal development and suggesting the possible involvement of this protein in neurological disorders.

Muscle sympathetic nervous activity in depressed patients before and after treatment with sertraline.

BACKGROUND: Sympathetic hyperactivity is one of the mechanisms involved in the increased cardiovascular risk associated with depression, and there is evidence that antidepressants decrease sympathetic activity. OBJECTIVES: We tested the following two hypotheses: patients with major depressive disorder with high scores of depressive symptoms (HMDD) have augmented muscle sympathetic nervous system activity (MSNA) at rest and during mental stress compared with patients with major depressive disorder with low scores of depressive symptoms (LMDD) and controls; sertraline decreases MSNA in depressed patients. METHODS: Ten HMDD, nine LMDD and 11 body weight-matched controls were studied. MSNA was directly measured from the peroneal nerve using microneurography for 3 min at rest and 4 min during the Stroop color word test. For the LMDD and HMDD groups, the tests were repeated after treatment with sertraline (103.3 +/- 40 mg). RESULTS: Resting MSNA was significantly higher in the HMDD [29.1 bursts/min (SE 2.9)] compared with LMDD [19.9 (1.6)] and controls [22.2 (2.0)] groups (P = 0.026 and 0.046, respectively). There was a significant positive correlation between resting MSNA and severity of depression. MSNA increased significantly and similarly during stress in all the studied groups. Sertraline significantly decreased resting MSNA in the LMDD group and MSNA during mental stress in LMDD and HMDD groups. Sertraline significantly decreased resting heart rate and heart rate response to mental stress in the HMDD group. CONCLUSION: Moderate-to-severe depression is associated with increased MSNA. Sertraline treatment reduces MSNA at rest and during mental challenge in depressed patients, which may have prognostic implications in this group.

[Effects of lisinopril on diabetic peripheral neuropathy: experiment with rats].

OBJECTIVE: To investigate the effects of lisinopril, an angiotensin-converting enzyme inhibitor, on diabetic peripheral neuropathy (DNP). METHODS: Twenty-five Wistar rats underwent intravenous injection of streptozocin to establish diabetes models and 10 rats were injected with sodium citrate solution as normal controls. The diabetic rats were randomly divided into 2 groups: lisinopril group treated with gastric perfusion of lisinopril daily for 8 weeks, and diabetic control group. The diabetic controls and normal controls were treated with gastric perfusion of water. Sciatic nerve electrode penetration method was used to measure the motor nerve conduction velocity (MNCV) and sensory nerve conduction velocity (SNCV). Light and heat pain measuring apparatus was used to measure the pain threshold. Then the sciatic nerves were isolated. Electron microscopy was used to observe the ultra-structure. The contents of superoxide dismutase (SOD) and malonyldialdehyde (MDA), and Na(+)K(+)-ATPase activity were detected by chemical colorimetry. Immunohistochemistry was used to detect the CD34 in the sciatic nerve. The capillary density of sciatic nerve was calculated. RESULTS: The MNCV and SNCV levels of the lisinopril group were both lower than those of the 2 control groups (all P < 0.01). The potency of heat pain leg retraction response of the lisinopril group was significantly shorter than that pf the diabetic control group (P < 0.05). The pathological changes of the lisinopril group were milder than those of the other groups. The SOD level and the Na(+)K(+)-ATPase activity of the diabetic group were significantly lower than those of the normal control group, and the MDA of the diabetic group was significantly higher than those of the other 2 groups (all P < 0.05). And the SOD level and Na(+)K(+)-ATPase activity of the lisinopril group were significantly higher than those of the diabetic control group, and the MDA level of the lisinopril group was significantly lower than that of the diabetic control group. The capillary density of sciatic nerve of the diabetic control group was lower and the normal control group, and that of the lisinopril group was significantly higher than that of the diabetic control group (P < 0.01). CONCLUSION: ACE inhibitors effectively prevent and treat DPN.

Selective activation of microglia in spinal cord but not higher cortical regions following nerve injury in adult mouse.

Neuronal plasticity along the pathway for sensory transmission including the spinal cord and cortex plays an important role in chronic pain, including inflammatory and neuropathic pain. While recent studies indicate that microglia in the spinal cord are involved in neuropathic pain, a systematic study has not been performed in other regions of the central nervous system (CNS). In the present study, we used heterozygous Cx3cr1GFP/+mice to characterize the morphological phenotypes of microglia following common peroneal nerve (CPN) ligation. We found that microglia showed a uniform distribution throughout the CNS, and peripheral nerve injury selectively activated microglia in the spinal cord dorsal horn and related ventral horn. In contrast, microglia was not activated in supraspinal regions of the CNS, including the anterior cingulate cortex (ACC), prefrontal cortex (PFC), primary and secondary somatosensory cortex (S1 and S2), insular cortex (IC), amygdala, hippocampus, periaqueductal gray (PAG) and rostral ventromedial medulla (RVM). Our results provide strong evidence that nerve injury primarily activates microglia in the spinal cord of adult mice, and pain-related cortical plasticity is likely mediated by neurons.

Role of matrix metalloproteinases, proinflammatory cytokines, and oxidative stress-derived molecules in hepatitis C virus-associated mixed cryoglobulinemia vasculitis neuropathy.

OBJECTIVE: Mixed cryoglobulinemia (MC) is a systemic vasculitis, usually associated with hepatitis C virus (HCV) infection. The molecular mechanisms responsible for HCV-associated MC (HCV-MC) vasculitis are largely unknown. This study was undertaken to assess the expression profile of selected genes involved in inflammatory vascular damage in patients with HCV-MC vasculitis, patients with polyarteritis nodosa (PAN), and patients with noninflammatory idiopathic neuropathy. METHODS: The quantitative expression levels of 42 selected genes involved in inflammatory vascular damage were assessed in nerve lesions of patients with HCV-MC vasculitis, PAN (rheumatic disease controls), and noninflammatory idiopathic neuropathy (noninflammatory neuropathy controls), using real-time reverse transcriptase-polymerase chain reaction. Genes were considered to be differentially expressed when there was a >2-fold difference in mean expression levels between groups and the P value was less than 0.05. RESULTS: Expression levels of 8 genes were significantly increased in HCV-MC patients versus control patients with noninflammatory idiopathic neuropathy, with the highest increase for metallothionein 1 H (MT1H), a hypoxic and oxidative stress protein. Compared with PAN patients, HCV-MC patients had higher expression levels of genes encoding oxidative stress-derived molecules (MT1H, endothelial cell nitric oxide synthase 3, Hsp70, and Hsp90) and tissue plasminogen activator and lower expression levels of matrix metalloproteinase 7 (MMP-7). HCV-MC neuropathies were classified according to their morphologic pattern and the presence or absence of necrotizing arteritis. MMP-1, MMP-7, MMP-9, and interleukin-1beta were up-regulated in patients with necrotizing arteritis. CONCLUSION: This comprehensive molecular study of HCV-MC vasculitis provides strong evidence that MMPs, proinflammatory cytokines, and oxidative stress-derived molecules have a role in the pathogenesis of HCV-MC vasculitis neuropathy.

Involvement of chemokines and type 1 cytokines in the pathogenesis of hepatitis C virus-associated mixed cryoglobulinemia vasculitis neuropathy.

OBJECTIVE: To examine the expression profiles of a large number of genes within typical vasculitic nerve lesions in patients with mixed cryoglobulinemia (MC) vasculitis in order to better characterize the molecules involved in cellular tissue activation and trafficking. METHODS: The quantitative expression of 19 genes coding for cytokines, chemokines, and their receptors in the nerve lesions of 9 patients with hepatitis C virus (HCV)-associated MC vasculitis, 7 with idiopathic polyarteritis nodosa (PAN) (rheumatic disease controls), and 8 patients with noninflammatory idiopathic neuropathy (noninflammatory neuropathy controls) was assessed using a real-time reverse transcriptase-polymerase chain reaction procedure. RESULTS: Compared with the noninflammatory controls, HCV-MC vasculitis patients had a significantly higher expression of Th1 cytokines in vasculitic nerve lesions (mean +/- SEM fold increase 33.7 +/- 11.6 for interferon-gamma and 7.2 +/- 1.9 for tumor necrosis factor alpha), whereas Th2 cytokines were absent (interleukin-4 [IL-4], IL-5, and IL-13) or were not significantly different (IL-10). Chemokines involved in T cell and monocyte trafficking were also significantly up-regulated in the HCV-MC vasculitis patients (mean +/- SEM fold increase 27.4 +/- 8.3 for macrophage inflammatory protein 1alpha [MIP-1alpha], 19.9 +/- 5.7 for MIP-1beta, and 7.2 +/- 1.5 for CXCR3). Compared with patients with idiopathic PAN, there was a trend toward higher expression of MIP-1alpha and CXCR3 in HCV-MC vasculitis patients (mean +/- SEM fold increase 27.4 +/- 8.3 versus 5.3 +/- 3.4 for MIP-1alpha and 7.2 +/- 1.5 versus 2.5 +/- 0.9 for CXCR3). CONCLUSION: This study is the first to demonstrate a role of cellular immunity and Th1 lymphocytes in the pathogenesis of HCV-MC vasculitic nerve lesions.

Amyloid neuropathy: a retrospective study of 35 peripheral nerve biopsies.

We performed a retrospective study of 35 peripheral nerve biopsies (PNBs) with amyloid deposits in the endoneurium. In every case, nerve lesions were studied on paraffin-embedded fragments (PEFs) and by ultrastructural examination (USE). In addition, muscle fragments were taken and embedded in paraffin. Immunohistochemistry was performed with anti-transthyretin (TTR) serum on 19 nerve and 15 muscle PEFs. Direct immunofluorescence with anti-light-chain sera was performed on frozen nerve fragments in 19 cases. Endoneurial amyloid deposits were easily identified on routine PEF in 26 cases, after Congo red or thioflavine staining in three, and by USE in six. A dramatic myelinated fiber loss was evidenced in 34 cases (77-2970 per mm2), and features of axonal degeneration were present in every case. Segmental demyelination was observed in 10 cases. A mutation in the TTR gene was present in 14 cases, with Met30 mutation in 10 and Ala49 in four members of the same family. Amyloid deposits were strongly marked by the anti-TTR serum in 11 other cases, twice in the endoneurium, five around muscle fibers, and four in both locations. In eight patients, light-chain positivity was evidenced in endoneurial deposits, lambda in six and kappa in two. Two other patients with monoclonal gammopathy did not present any light-chain fixation. In 17 cases, amyloidosis was disclosed by PNB and 13 had a TTR pathology; eight of them, over 65 years old, correspond to a late-onset form of familial amyloid polyneuropathy which is an underdiagnosed condition.