T-cell immunosuppression in sepsis is augmented by sciatic denervation-induced skeletal muscle atrophy.

Skeletal muscle atrophy is a known risk factor for immunosuppressive conditions and for a poor prognosis in sepsis. However, its immunopathology has not been experimentally elucidated. This study investigated the effects of skeletal muscle atrophy on the immunopathology of sepsis. Male C57BL/6J mice were subjected to sciatic denervation (DN) and caecal ligation and puncture (CLP) to induce muscle atrophy or sepsis. The macrophages, myeloid-derived suppressor cells (MDSC), and T-cells in peritoneal and spleen were analysed using flow cytometry. DN-induced muscle atrophy did not affect macrophage and MDSC populations. In contrast, the percentage of cytotoxic T-lymphocyte-associated antigen (CTLA)-4+ CD4+ T-cells, programmed death (PD)-1+ CD8+ T-cells, regulatory T-cells, and the CTLA-4+ regulatory T-cells was statistically significantly higher in DN-CLP mice than in sham-CLP mice. Skeletal muscle atrophy before sepsis triggers excessive T cell immunosuppression, which may contribute to the exacerbation of sepsis under skeletal muscle atrophy.

Pathogenic TNF-α drives peripheral nerve inflammation in an Aire-deficient model of autoimmunity.

Immune cells infiltrate the peripheral nervous system (PNS) after injury and with autoimmunity, but their net effect is divergent. After injury, immune cells are reparative, while in inflammatory neuropathies (e.g., Guillain Barré Syndrome and chronic inflammatory demyelinating polyneuropathy), immune cells are proinflammatory and promote autoimmune demyelination. An understanding of immune cell phenotypes that distinguish these conditions may, therefore, reveal new therapeutic targets for switching immune cells from an inflammatory role to a reparative state. In an autoimmune regulator (Aire)-deficient mouse model of inflammatory neuropathy, we used single-cell RNA sequencing of sciatic nerves to discover a transcriptionally heterogeneous cellular landscape, including multiple myeloid, innate lymphoid, and lymphoid cell types. Analysis of cell-cell ligand-receptor interactions uncovered a macrophage-mediated tumor necrosis factor-α (TNF-α) signaling axis that is induced by interferon-γ and required for initiation of autoimmune demyelination. Developmental trajectory visualization suggested that TNF-α signaling is associated with metabolic reprogramming of macrophages and polarization of macrophages from a reparative state in injury to a pathogenic, inflammatory state in autoimmunity. Autocrine TNF-α signaling induced macrophage expression of multiple genes (Clec4e, Marcksl1, Cxcl1, and Cxcl10) important in immune cell activation and recruitment. Genetic and antibody-based blockade of TNF-α/TNF-α signaling ameliorated clinical neuropathy, peripheral nerve infiltration, and demyelination, which provides preclinical evidence that the TNF-α axis may be effectively targeted to resolve inflammatory neuropathies.

CX3CR1 But Not CCR2 Expression Is Required for the Development of Autoimmune Peripheral Neuropathy in Mice.

One hallmark of Guillain-Barre syndrome (GBS), a prototypic autoimmune peripheral neuropathy (APN) is infiltration of leukocytes (macrophages and T cells) into peripheral nerves, where chemokines and their receptors play major roles. In this study, we aimed to understand the potential contribution of chemokine receptors CCR2 and CX3CR1 in APN by using a well-established mouse model, B7.2 transgenic (L31) mice, which possesses a predisposed inflammatory background. We crossbred respectively CCR2KO and CX3CR1KO mice with L31 mice. The disease was initiated by partial ligation on one of the sciatic nerves. APN pathology and neurological function were evaluated on the other non-ligated sciatic nerve/limb. Our results revealed that L31/CX3CR1KO but not L31/CCR2KO mice were resistant to APN. CX3CR1 is needed for maintaining circulating monocyte and CD8+ T cell survival. While migration of a significant number of activated CD8+ T cells to peripheral nerves is essential in autoimmune response in nerve, recruitment of monocytes into PNS seems optional. Disease onset is independent of CCR2 mediated blood-derived macrophage recruitment, which can be replaced by compensatory proliferation of resident macrophages in peripheral nerve. CX3CR1 could also contribute to APN via its critical involvement in maintaining nerve macrophage phagocytic ability. We conclude that blockade of CX3CR1 signaling may represent an interesting anti-inflammatory strategy to improve therapeutic management for GBS patients.

Effect of glutaraldehyde-crosslinked cartilage acellular matrix film on anti-adhesion and nerve regeneration in a rat sciatic nerve injury model.

Decellularized extra-cellular matrix (ECM) has been studied as an alternative to anti-adhesive biomaterials and cartilage acellular matrix (CAM) has been shown to inhibit postoperative adhesion in several organs. This study aimed to evaluate the suitability of glutaraldehyde (GA) crosslinked CAM-films as anti-adhesion barriers for peripheral nerve injury. The films were successfully fabricated and showed improved physical properties such as mechanical strength, swelling ratio, and lengthened degradation period while maintaining the microstructure and chemical composition after GA crosslinking. In the in vitro study of CAM-film, the dsDNA content met the recommended limit of decellularization and more than 70% of the major ECM components were preserved after decellularization. The adhesion and proliferation of seeded human umbilical vein endothelial cells and fibroblasts were significantly lower in CAM-film than in control, but similar with Seprafilm. However, the CAM-film extract did not show cytotoxicity. In the in vivo study, the peri-neural fibrosis was thicker, adhesion score higher, and peri-neural collagen fibers more abundant in the control group than in the CAM-film group. The total number of myelinated axons was significantly higher in the CAM-film group than in the control group. The inflammatory marker decreased with time in the CAM-film group compared to that in the control group, whereas the nerve regenerative marker expression was maintained. Moreover, the ankle angles at contracture and toe-off were higher in the CAM film-treated rats than in the control rats. GA-crosslinked CAM films may be used during peripheral nerve surgery to prevent peri-neural adhesion and enhance nerve functional recovery.

Spinal Ninjurin2 contributes to the neuropathic pain via NF-κB-mediated neuroinflammation in the spared sciatic nerve injury rats.

OBJECT: Ninjurin2 (nerve injury induced protein 2, NINJ2) is a molecule which mediates cell-to-cell and cell-to-extracellular matrix interactions in the nervous system. Clinical study shows NINJ2 is associated with the development of postherpetic neuralgia. However, it is lack of direct evidence that NINJ2 participated in neuropathic pain. In this study, we aim to investigate the role of NINJ2 in the development of neuropathic pain in spared sciatic nerve injury rats and the underlying mechanism. METHOD: Spared sciatic nerve injury (SNI) models were established. The level of NINJ2 and p-p65 (a NF-κB family member) were measured in SNI rats by western blots and immunofluorescent staining. Lentivirus encoding small interfering RNA targeting NINJ2 (RNAi) was intrathecally injected into rats. Then the change of pain behavior of rats induced by NINJ2 RNAi was tested by Von-Frey hairs. The change of p-p65 in the spinal cord in rats after NINJ2 RNAi treatment was also measured by western blots. inhibitor of p-p65-induced change of TNF-α, IL-1ß, and IL-6 levels were measured by ELISA. RESULTS: NINJ2 and p-p65 were increased in the spinal cord of SNI rats on the 3, 7, 14th days after modeling. NINJ2 were mainly expressed in neurons, and co-located with p-p65 in the spinal dorsal horn. When down regulating the level of NINJ2 by RNAi, the development of pain in SNI rats was partially blocked. Phosphorylation of p65 was also inhibited by NINJ2 RNAi. Blocking the phosphorylation of NF-κB pathway could inhibit the increase of TNF-α, IL-1ß, and IL-6 in the spinal cord of SNI rats. CONCLUSION: NINJ2 protein was increased in the spinal cord of SNI rats. It participated in the development of nerve injury-induced neuropathic pain by activating neuroinflammation in the spinal cord via NF-κB pathway. This study provides a new target to investigate the mechanism of neuropathic pain.

The Expression of Chemokines Is Downregulated in a Pre-Clinical Model of TTR V30M Amyloidosis.

Inflammation is a hallmark of several neurodegenerative disorders including hereditary amyloidogenic transthyretin amyloidosis (ATTRv). ATTRv is an autosomal dominant neurodegenerative disorder with extracellular deposition of mutant transthyretin (TTR) aggregates and fibrils, particularly in nerves and ganglia of the peripheral nervous system. Nerve biopsies from ATTRv patients show increased cytokine production, but interestingly no immune inflammatory cellular infiltrate is observed around TTR aggregates. Here we show that as compared to Wild Type (WT) animals, the expression of several chemokines is highly downregulated in the peripheral nervous system of a mouse model of the disease. Interestingly, we found that stimulation of mouse Schwann cells (SCs) with WT TTR results in the secretion of several chemokines, a process that is mediated by toll-like receptor 4 (TLR4). In contrast, the secretion of all tested chemokines is compromised upon stimulation of SCs with mutant TTR (V30M), suggesting that V30M TTR fails to activate TLR4 signaling. Altogether, our data shed light into a previously unappreciated mechanism linking TTR activation of SCs and possibly underlying the lack of inflammatory response observed in the peripheral nervous system of ATTRv patients.

Loss of direct adrenergic innervation after peripheral nerve injury causes lymph node expansion through IFN-γ.

Peripheral nerve injury can cause debilitating disease and immune cell-mediated destruction of the affected nerve. While the focus has been on the nerve-regenerative response, the effect of loss of innervation on lymph node function is unclear. Here, we show that the popliteal lymph node (popLN) receives direct neural input from the sciatic nerve and that sciatic denervation causes lymph node expansion. Loss of sympathetic, adrenergic tone induces the expression of IFN-γ in LN CD8 T cells, which is responsible for LN expansion. Surgery-induced IFN-γ expression and expansion can be rescued by ß2 adrenergic receptor agonists but not sensory nerve agonists. These data demonstrate the mechanisms governing the pro-inflammatory effect of loss of direct adrenergic input on lymph node function.

UVB irradiation induces contralateral changes in galanin, substance P and c-fos immunoreactivity in rat dorsal root ganglia, dorsal horn and lateral spinal nucleus.

The selection of control group is crucial, as the use of an inadequate group may strongly affect the results. In this study we examine the effect on contralateral tissue protein levels, in a model of unilateral UVB irradiation, as the contralateral side is commonly used as a control. Previous studies have shown that UVB irradiation increases immunoreactivity for inflammatory regulated neuropeptides. Unilateral UVB irradiation of rat hind paw was performed and corresponding contralateral spinal cord and dorsal root ganglia (DRG) were collected 2-96 h after and investigated for changes in galanin, substance P and c-fos immunoreactivity. Control tissue was collected from naïve rats. Measurement of skin blood flow from contralateral heel hind paws (Doppler), revealed no change compared to naïve rats. However, UVB irradiation caused a significant reduction in the contralateral proportion of galanin immunopositive DRG neurons, at all-time points, as well as an increase in the contralateral spinal cord dorsal horn, around the central canal and in the lateral spinal nucleus (2-48 h). The contralateral proportion of SP positive DRG neurons and dorsal horn immunoreactivity was unchanged, whereas the lateral spinal nucleus area showed increased immunoreactivity (48 h). UVB irradiation also induced a slight contralateral upregulation of c-fos in the dorsal horn/central canal area (24 and 48 h). In summary, unilateral UVB irradiation induced contralateral changes in inflammatory/nociceptive neuropeptides in spinal cord and afferent pathways involved in pain signaling already within 24 h, a time point when also ipsilateral neurochemical/physiological changes have been reported for rats and humans.

Very Early Involvement of Innate Immunity in Peripheral Nerve Degeneration in SOD1-G93A Mice.

Recent preclinical and clinical evidence suggest that immune system has a role in the progression and prognosis of Amyotrophic Lateral Sclerosis (ALS), but the identification of a clear mechanism and immune players remains to be elucidated. Here, we have investigated, in 30 and 60 days (presymptomatic) and 120 days (symptomatic) old SOD1-G93A mice, systemic, peripheral, and central innate and adaptive immune and inflammatory response, correlating it with the progression of the neurodegeneration in neuromuscular junction, sciatic nerves, and spinal cord. Surprisingly, we found a very initial (45-60 days) presence of IgG in sciatic nerves together with a gradual enhancement of A20/TNFAIP3 (protein controlling NF-κB signalling) and a concomitantly significant increase and activation of circulating mast cells (MCs) as well as MCs and macrophages in sciatic nerve and an enhancement of IL-6 and IL-10. This immunological frame coincided with a myelin aggregation. The 30-60 days old SOD1-G93A mice didn't show real elements of neuroinflammation and neurodegeneration in spinal cord. In 120 days old mice macrophages and monocytes are widely diffused in sciatic nerves, peripheral neurodegeneration reaches the tip, high circulating levels of TNFα and IL-2 were found and spinal cord exhibits clear signs of neural damage and infiltrating immune cells. Our results underpin a clear immunological disorder at the origin of ALS axonopathy, in which MCs are involved in the initiation and sustaining of inflammatory events. These data cannot be considered a mere epiphenomenon of motor neuron degeneration and reveal new potential selective immune targets in ALS therapy.

Analysis of the immune response to sciatic nerve injury identifies efferocytosis as a key mechanism of nerve debridement.

Sciatic nerve crush injury triggers sterile inflammation within the distal nerve and axotomized dorsal root ganglia (DRGs). Granulocytes and pro-inflammatory Ly6Chigh monocytes infiltrate the nerve first and rapidly give way to Ly6Cnegative inflammation-resolving macrophages. In axotomized DRGs, few hematogenous leukocytes are detected and resident macrophages acquire a ramified morphology. Single-cell RNA-sequencing of injured sciatic nerve identifies five macrophage subpopulations, repair Schwann cells, and mesenchymal precursor cells. Macrophages at the nerve crush site are molecularly distinct from macrophages associated with Wallerian degeneration. In the injured nerve, macrophages 'eat' apoptotic leukocytes, a process called efferocytosis, and thereby promote an anti-inflammatory milieu. Myeloid cells in the injured nerve, but not axotomized DRGs, strongly express receptors for the cytokine GM-CSF. In GM-CSF-deficient (Csf2-/-) mice, inflammation resolution is delayed and conditioning-lesion-induced regeneration of DRG neuron central axons is abolished. Thus, carefully orchestrated inflammation resolution in the nerve is required for conditioning-lesion-induced neurorepair.

Modifying macrophages at the periphery has the capacity to change microglial reactivity and to extend ALS survival.

Microglia and peripheral macrophages have both been implicated in amyotrophic lateral sclerosis (ALS), although their respective roles have yet to be determined. We now show that macrophages along peripheral motor neuron axons in mouse models and patients with ALS react to neurodegeneration. In ALS mice, peripheral myeloid cell infiltration into the spinal cord was limited and depended on disease duration. Targeted gene modulation of the reactive oxygen species pathway in peripheral myeloid cells of ALS mice, using cell replacement, reduced both peripheral macrophage and microglial activation, delayed symptoms and increased survival. Transcriptomics revealed that sciatic nerve macrophages and microglia reacted differently to neurodegeneration, with abrupt temporal changes in macrophages and progressive, unidirectional activation in microglia. Modifying peripheral macrophages suppressed proinflammatory microglial responses, with a shift toward neuronal support. Thus, modifying macrophages at the periphery has the capacity to influence disease progression and may be of therapeutic value for ALS.

Human Wharton's Jelly Mesenchymal Stem Cell-Mediated Sciatic Nerve Recovery Is Associated with the Upregulation of Regulatory T Cells.

The acceleration of peripheral nerve regeneration is crucial for functional nerve recovery. Our previous study demonstrated that human Wharton's jelly-derived mesenchymal stem cells (hWJ-MSC) promote sciatic nerve recovery and regeneration via the direct upregulation and release of neurotrophic factors. However, the immunomodulatory role of hWJ-MSC in sciatic nerve recovery remains unclear. The effects of hWJ-MSC on innate immunity, represented by macrophages, natural killer cells, and dendritic cells, as well as on adaptive immunity, represented by CD4+ T, CD8+ T, B, and regulatory T cells (Tregs), were examined using flow cytometry. Interestingly, a significantly increased level of Tregs was detected in blood, lymph nodes (LNs), and nerve-infiltrating cells on POD7, 15, 21, and 35. Anti-inflammatory cytokines, such as IL-4 and IL-10, were significantly upregulated in the LNs and nerves of hWJ-MSC-treated mice. Treg depletion neutralized the improved effects of hWJ-MSC on sciatic nerve recovery. In contrast, Treg administration promoted the functional recovery of five-toe spread and gait stance. hWJ-MSC also expressed high levels of the anti-inflammatory cytokines TGF-ß and IL-35. This study indicated that hWJ-MSC induce Treg development to modulate the balance between pro- and anti-inflammation at the injured sciatic nerve by secreting higher levels of anti-inflammatory cytokines.

Imaging resident and recruited macrophage contribution to Wallerian degeneration.

Wallerian degeneration (WD) is a process of autonomous distal degeneration of axons upon injury. Macrophages (MPs) of the peripheral nervous system (PNS) are the main cellular agent controlling this process. Some evidence suggests that resident PNS-MPs along with MPs of hematogenous origin may be involved, but whether these two subsets exert distinct functions is unknown. Combining MP-designed fluorescent reporter mice and coherent anti-Stokes Raman scattering (CARS) imaging of the sciatic nerve, we deciphered the spatiotemporal choreography of resident and recently recruited MPs after injury and unveiled distinct functions of these subsets, with recruited MPs being responsible for efficient myelin stripping and clearance and resident MPs being involved in axonal regrowth. This work provides clues to tackle selectively cellular processes involved in neurodegenerative diseases.

Profiling peripheral nerve macrophages reveals two macrophage subsets with distinct localization, transcriptome and response to injury.

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: Relmα+Mgl1+ snMacs in the epineurium and Relmα-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.

Efecto del ácido láctico en la respuesta celular del sistema inmunitario durante la fase inflamatoria temprana en el nervio isquiático dañado por compresión / Effect of lactic acid on the cellular response of the immune system during the early inflammatory phase in the sciatic nerve damaged by compression

En órganos dañados, el ácido láctico (AL) modifica la respuesta inmune innata e inflamatoria, induciendo una menor expresión de citoquinas pro-inflamatorias, que provocan, la modulación del reclutamiento de células inmunes. El daño por compresión del nervio isquiático (NI) desencadena una respuesta inflamatoria y un aumento exponencial del infiltrado inflamatorio de células inmunes, produciendo la destrucción de axones y pérdida funcional del nervio. El objetivo de este estudio es evaluar el efecto agudo de la inyección de AL, sobre la proporción de células inmunes en la fase inflamatoria temprana, en el sitio de lesión del NI post compresión. Para ello, se utilizaron 15 ratas machos Sprague Dawley adultas, en tres grupos de compresión nerviosa. Un grupo control, un grupo control negativo con placebo (100 µL PBS) y un grupo experimental con inyección de 100 µL de AL [20mM]. Al tercer día los NI se analizaron histológicamente y se estableció la proporción de células inmunes en el sitio de lesión. Los resultados muestran que la inyección intraneural de AL provoca una disminución en el porcentaje de linfocitos y un aumento en el porcentaje de macrófagos. Este es el primer trabajo de inyección intraneural de AL y demuestra el efecto modulador del AL sobre las células inmunes en el sistema nervioso periférico.

In damaged organs, lactic acid (LA) modifies the innate and inflammatory immune response, inducing a lower expression of pro-inflammatory cytokines, which provoke the modulation of immune cell recruitment. Damage by compression of the sciatic nerve (SN) triggers an inflammatory response and an exponential increase in the inflammatory infiltrate of immune cells, producing the destruction of axons and functional loss of the nerve. The objective of this study is to evaluate the acute effect of the injection of LA, on the proportion of immune cells in the early inflammatory phase, in the site of SN post-compression injury. For this, 15 adult Sprague Dawley rats were used in three groups of nervous compression. A control group, a negative control group with placebo (100 mL PBS) and an experimental group with injection of 100 mL of LA [20mM]. On the third day, the SNs were histologically analyzed and the proportion of immune cells at the injury site was established. The results show that the intraneural injection of LA causes a decrease in the percentage of lymphocytes and an increase in the percentage of macrophages. This is the first work of intraneural injection of LA and demonstrates the modulating effect of LA on immune cells in the peripheral nervous system.

Inflammatory Macrophages in the Sciatic Nerves Facilitate Neuropathic Pain Associated with Type 2 Diabetes Mellitus.

Despite the requirement for effective medication against neuropathic pain associated with type 2 diabetes mellitus (T2DM), mechanism-based pharmacotherapy has yet to be established. Given that long-lasting neuroinflammation, driven by inflammatory macrophages in the peripheral nerves, plays a pivotal role in intractable pain, it is important to determine whether inflammatory macrophages contribute to neuropathic pain associated with T2DM. To generate an experimental model of T2DM, C57BL/6J mice were fed a high-fat diet (HFD) ad libitum. Compared with control diet feeding, obesity and hyperglycemia were observed after HFD feeding, and the mechanical pain threshold evaluated using the von Frey test was found to be decreased, indicating the development of mechanical allodynia. The expression of mRNA markers for macrophages, inflammatory cytokines, and chemokines were significantly upregulated in the sciatic nerve (SCN) after HFD feeding. Perineural administration of saporin-conjugated anti-Mac1 antibody (Mac1-Sap) improved HFD-induced mechanical allodynia. Moreover, treatment of Mac1-Sap decreased the accumulation of F4/80+ macrophages and the upregulation of inflammatory mediators in the SCN after HFD feeding. Inoculation of lipopolysaccharide-activated peritoneal macrophages in tissue surrounding the SCN elicited mechanical allodynia. Furthermore, pharmacological inhibition of inflammatory macrophages by either perineural or systemic administration of TC-2559 [4-(5-ethoxy-3-pyridinyl)-N-methyl-(3E)-3-buten-1-amine difumarate], a α4ß2 nicotinic acetylcholine receptor-selective agonist, relieved HFD-induced mechanical allodynia. Taken together, inflammatory macrophages that accumulate in the SCN mediate the pathophysiology of neuropathic pain associated with T2DM. Inhibitory agents for macrophage-driven neuroinflammation could be potential candidates for novel pharmacotherapy against intractable neuropathic pain.

scAAV9-VEGF-165 inhibits neuroinflammatory responses and invasion of macrophages into the peripheral nervous system of ALS transgenic mice.

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disorder that leads to paralysis and death within 3-5 years. Although the vast majority of studies have focused on vulnerable neurons, growing evidence has shown that non-neuronal cells contribute to the pathogenesis and disease progression. Here, we showed that intrathecal injection of scAAV9-VEGF at 60 days of age significantly reduced the number of microglia and inhibited the neuroinflammatory response in the CNS. Meanwhile, we found that administration of VEGF inhibited the invasion of macrophages into the PNS, including ventral nerve roots, sciatic nerves and muscles. Overall, our study indicated the anti-inflammation effect of VEGF in the CNS and PNS of ALS mice when delivered by intrathecal injection.

GLP­1R agonists ameliorate peripheral nerve dysfunction and inflammation via p38 MAPK/NF­κB signaling pathways in streptozotocin­induced diabetic rats.

The present study aimed to investigate the mechanism of glucagon­like peptide­1 receptor (GLP­1R) agonists in the progression of diabetic peripheral neuropathy (DPN) in streptozotocin (STZ)­induced diabetic rats, through inflammatory signaling pathways. The DPN rat model was generated by intraperitoneal injection of STZ and then treated with the GLP­1R agonist liraglutide or saline for 8 weeks. These animals were randomly divided into 4 groups (10 rats in each): The normal control + saline group, the normal control + liraglutide group, the diabetic + saline (DM) group and the diabetic + liraglutide (DML) group. The nerve conduction velocity (NCV) in the sciatic nerves of the rats was monitored over a period of 8 weeks. Peripheral serum was obtained for the measurement of blood glucose, tumor necrosis factor­α (TNF­α), interleukin­6 (IL­6) and IL­1ß level. The protein levels of phosphorylated (p­) and total extracellular signal­regulated kinase, c­Jun NH2­terminal kinases, p38 mitogen­activated protein kinases (MAPK), and nuclear and cytoplasmic nuclear factor­κB (NF­κB) were measured through western blot analysis. Sciatic nerve mRNA expression levels of proinflammatory chemokines (TNF­α, IL­6 and IL­1ß), chemokines [monocyte chemoattractant protein­1 (MCP­1)], adhesion molecules [intercellular adhesion molecule 1 (ICAM­1)], neurotrophic factors [neuritin, nerve growth factor (NGF) and neuron­specific enolase (NSE)] and NADPH oxidase 4 (NOX4) were evaluated by reverse transcription-quantitative polymerase chain reaction. Subsequent to 8 weeks of treatment with liraglutide, the density of myelin nerve fibers was partially restored in the DML group. The delayed motor NCV and sensory NCV in the DML group were improved. The IOD value of NOX4 staining in the DML group (24.43±9.01) was reduced compared with that in the DM group (56.60±6.91). The levels of TNF­α, IL­1ß and IL­6 in the peripheral serum of the DML group were significantly suppressed compared with those of the DM group. It was also observed that the mRNA expression levels of TNF­α, IL­6, IL­1ß, MCP­1, ICAM­1 and NOX4 in the sciatic nerve were attenuated in the DML group. The mRNA expression of neuritin and NGF was significantly increased in the DML group compared with that of the DM group; NSE was reduced in the sciatic nerves of the DML group compared with that of the DM group. Additionally, the protein expression of p­p38 MAPK and NF­κB in the DML group was significantly suppressed. These data demonstrated that GLP­1R agonists may prevent nerve dysfunction in the sciatic nerves of diabetic rats via p38 MAPK/NF­κB signaling pathways independent of glycemic control. GLP­1R agonists may be a useful therapeutic strategy for slowing the progression of DPN.

IGF1 affects macrophage invasion and activation and TNF-α production in the sciatic nerves of female SOD1G93A mice.

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease leading to paralysis and death within 3-5 years of its diagnosis. The SOD1G93A mouse is used extensively as an ALS animal model. Increasing evidence shows that non-neuronal cellscontribute to the pathogenesis and progression of ALS. Among them, many studies focus on microgliosis in the spinal cord (SC); while few on macrophage activation in the sciatic nerves. Substantial evidence shows that insulin-like growth factor 1 (IGF1) delays disease progression and increases the lifespan of SOD1G93A mice, and some studies indicate that IGF1 reduces inflammation in the SC of ALS mice. However, no studies have focused on the effect of IGF on sciatic nerve inflammation. Here, we find that ALS progression is characterized by increasing macrophage invasion and activation accompanied by significant TNF-α production in the sciatic nerve. Furthermore, IGF1 treatment and knockdown alleviate and aggravate these responses, respectively. Collectively, our findings show the first time that IGF1 has an anti-inflammatory effect in the sciatic nerves of SOD1G93A mice.

Aluminum exposure for 60days at an equivalent human dietary level promotes peripheral dysfunction in rats.

Aluminum (Al) is a neurotoxic associated with a number of chronic human diseases. We investigated the effects of Al exposure at doses similar to human dietary levels and at a high level exposure to Al on the peripheral nervous system. Wistar male rats were divided into two major groups and received orally: 1) First group - Low level - rats were subdivided and treated for 60days: a) Control - received ultrapure water; b) AlCl3 - received Al at 8.3mg/kg body weight (bw) for 60days; and 2) Second group - High level - rats were subdivided and treated for 42days: C) Control - received ultrapure water through oral gavage; d) AlCl3 - received Al at 100mg/kg bw for 42days. Von Frey hair test, plantar test, the presence of catalepsy and the spontaneous motor activity were investigated. Reactive oxygen species, lipid peroxidation and total antioxidant capacity, immunohistochemistry to investigate the nerve inflammation and, the specific presence of Al in the sciatic nerve fibers were investigated. Al exposure at a representative human dietary level promotes the development of mechanical allodynia, catalepsy, increased inflammation in the sciatic nerve, systemic oxidative stress and, is able to be retained in the sciatic nerve. The effects of low-dose Al were similar to those found in rats exposed to Al at a dose much higher (100mg/kg). Our findings suggest that Al may be considered toxic for the peripheral nervous system, thus inducing peripheral dysfunction.