Nerve Conduction Velocity Deficits Research Articles

Glycyrrhizic acid promotes sciatic nerves recovery in type 1 diabetic rats and protects Schwann cells from high glucose-induced cytotoxicity.

The present study aims to investigate the therapeutic effect and mechanism of glycyrrhizic acid (GA) in diabetic peripheral neuropathy (DPN). GA significantly mitigated nerve conduction velocity (NCV) deficit and morphological abnormality and reduced high-mobility group box-1 (HMGB1) expression in the sciatic nerves of diabetic rats independent of blood glucose and body weight. Notably, GA alleviated the increase of HMGB1 and the decrease of cell viability in high glucose-stimulated RSC96 cells. Furthermore, GA obviously reduced the concentration of inflammatory cytokines in the sciatic nerves of diabetic rats and supernatants of high glucose-exposed RSC96 cells, then restored the decreased expression levels of nerve growth factor (NGF) and neuritin-1, and the increased expression levels of cleaved caspase-3 and neuron-specific enolase. Additionally, GA markedly inhibited receptor for advanced glycation end products (RAGE) expression, p38MAPK phosphorylation, and the nuclear translocation of NF-κBp65 in diabetic rats and high glucose-exposed RSC96 cells. The promotional effect of high glucose in RSC96 cells was diminished following Hmgb1 siRNA treatment. Our findings indicate that GA may exert neuroprotection on DPN by suppressing HMGB1, which lead to extenuation of inflammation response, balance of NGF, neuritin-1 and caspase-3, as well as inactivation of RAGE/p38MAPK/NF-κBp65 signaling pathway.

Allantoin attenuates deficits of behavioural and motor nerve conduction in an animal model of cisplatin-induced neurotoxicity in rats.

BackgroundThe aim of the present study was to evaluate the neuroprotective effect of allantoin in cisplatin‐induced toxicity in rats.MethodsAdult male Wistar rats weighing 160‐200 g were used. Neuropathy was induced by injecting cisplatin (2 mg/kg, ip, twice a week for 6 weeks) and the rats were concurrently treated with allantoin (200 and 400 mg/kg, po) for 8 weeks. At the end of the study, body weight and hemogram were measured. Behavioural tests were performed, including tests for cold and hot hyperalgesia, motor co‐ordination, locomotor activity, mechano‐tactile allodynia and mechanical hyperalgesia. The rats were then sacrificed and sciatic nerve conduction velocity was determined. The antioxidant enzyme and nitric oxide levels in sciatic nerve homogenates were measured.ResultsIn this study, allantoin restored the motor nerve conduction velocity deficits induced by cisplatin, and the allantoin‐treated rats showed improvement in cold and thermal hyperalgesia, mechano‐tactile allodynia, and mechanical hyperalgesia. Allantoin treatment also improved the rats’ hematological status, increasing haemoglobin, platelet and RBC counts compared to the cisplatin‐treated group. Allantoin treatment also mitigated the functional abnormalities seen in the cisplatin neuropathy group, protecting neurons from the neurotoxic effects of cisplatin.ConclusionAllantoin shows promise for use as an adjuvant drug in cancer treatment to protect against cisplatin‐induced neuropathy.

Ghrelin agonist HM01 attenuates chemotherapy-induced neurotoxicity in rodent models

Chemotherapy-Induced Peripheral Neurotoxicity (CIPN) is often dose-limiting and impacts life quality and survival of cancer patients. Ghrelin agonists have neuroprotectant effects and may have a role in treating or preventing CIPN. We evaluated the CNS-penetrant ghrelin agonist HM01 in three experimental models of CIPN at doses of 3–30 mg/kg p.o. daily monitoring orexigenic properties, nerve conduction, mechanical allodynia, and intra-epidermal nerve fiber density (IENFD). In a cisplatin-based study, rats were dosed daily for 3 days (0.5 mg/kg i.p.) + HM01. Cisplatin treatment induced mechanical hypersensitivity which was significantly reduced by HM01. In a second study, oxaliplatin was administered to mice (6 mg/kg i.p. 3 times/week for 4 weeks) resulting in significant digital nerve conduction velocity (NCV) deficits and reduction of IENFD. Concurrent HM01 dose dependently prevented the decline in NCV and attenuated the reduction in IENFD. Pharmacokinetic studies showed HM01 accumulation in the dorsal root ganglia and sciatic nerves which reached concentrations > 10 fold that of plasma. In a third model, HM01 was tested in preventive and therapeutic paradigms in a bortezomib-based rat model (0.2 mg/kg i.v., 3 times/week for 8 weeks). In the preventive setting, HM01 blocked bortezomib-induced hyperalgesia and IENFD reduction at all doses tested. In the therapeutic setting, significant effect was observed, but only at the highest dose. Altogether, the robust peripheral nervous system penetration of HM01 and its ability to improve multiple oxaliplatin-, cisplatin-, and bortezomib-induced neurotoxicities suggest that HM01 may be a useful neuroprotective adjuvant for CIPN.

Abstract 4503: Pharmacokinetics and pharmacodynamic analysis of eribulin mesylate and paclitaxel in mouse

Abstract Neuropathy arising from chemotherapy (CIPN) is a major clinical problem representing the dose-limiting side effect of many antineoplastic drugs. At their respective MTDs, we previously reported paclitaxel and ixabepilone produced more severe deficits in nerve conduction velocity, amplitude and degenerative changes in dorsal root ganglia (DRG) and sciatic nerve (SN) versus eribulin mesylate (Wozniak et al 2011). Similar trends for eribulin to cause less neuropathy have also been reported in the clinic (Cigler and Vadhat 2010, Jain and Cigler 2012, Vadhat et al 2013). The underlying reason for this differential effect remains elusive. Differences in tubulin binding may, in part, explain the differential effect (Perez et al 2009, Jordan et al 2005). Another potential explanation could reside in different pharmacokinetic (PK) and nervous tissue distribution of these agents. Toward this latter point, we conducted tissue distribution and pharmacokinetic studies following acute dosing of eribulin mesylate, paclitaxel and ixabepilone. We reported that while all three drugs rapidly cleared from plasma, they distributed into and cleared slowly (>3 days) from peripheral nervous system tissues (Wozniak et al 2014). We have now extended these studies to look out to 42 days following both acute and 2 week MTD-dosing paradigms evaluating drug pharmacokinetics in plasma, DRG and SN as well as pharmacodynamic measurements of nerve conduction and amplitude. We found plasma levels of eribulin mesylate and paclitaxel rapidly declined over 24h after infusion. In contrast, the levels of eribulin mesylate and paclitaxel persisted at above quantification levels (ranging from 5-10 ng/g) for 7 to 14 days post cessation of 2 week MTD dosing. Eribulin mesylate consistently showed greater distribution into DRG and SN over paclitaxel. When comparing the induced nerve conduction deficits, we found paclitaxel and eribulin mesylate caused reduction of nerve conduction parameters similar to our previous studies, most notably in caudal nerve amplitude (by 80.5% and 54.9% respectively). However these deficits, which were sustained for up to several weeks after cessation of dosing, were more severe with paclitaxel. The findings demonstrate that despite eribulin mesylate showing higher nervous tissue distribution than paclitaxel, eribulin mesylate caused less neurotoxicity as evidenced by milder reductions in nerve conduction and amplitude. This work was financially supported by Eisai. Citation Format: Krystyna Wozniak, Ying Wu, Bruce A. Littlefield, Kenichi Nomoto, Christopher DesJardins, Yanke Yu, George Lai, Larisa Reyderman, Barbara S. Slusher. Pharmacokinetics and pharmacodynamic analysis of eribulin mesylate and paclitaxel in mouse. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4503. doi:10.1158/1538-7445.AM2015-4503

Antioxidation and anti-inflammatory activity of Tang Bi Kang in rats with diabetic peripheral neuropathy.

BackgroundTang Bi Kang (TBK) is a traditional Chinese medicine granule. It has been shown to have effects on nerve conduction velocity deficits, blood-related factors and oxidative stress. This study was undertaken to evaluate proposed antioxidative and anti-inflammatory activity of Tang Bi Kang in rats with diabetic peripheral neuropathy (DPN).MethodsDPN was induced in male Wistar rats by intraperitoneal administration of streptozocin (STZ) (60 mg/kg.b.w) for 8 weeks. Fasting blood glucose (FBG) levels were measured in the blood obtained by clipping the tails of the rats. Tail-flick tests were conducted with a tail-flick analgesic meter. Motor and sensory nerve conduction velocities (MNCV and SNCV) of sciatic nerve were measured directly at two sites using a Functional Experiment System. Oxidative stress makers such as malondialdehyde (MDA), superoxide-dismutase (SOD) and glutathione peroxidase (GSH-Px), inflammatory cytokines such as interleukin (IL)-6, and tumour necrosis factor (TNF)-α were estimated. The statistical analysis of results was carried out using Student t-test and one-way analysis of variance (ANOVA), followed by least-significant difference post hoc with SPSS.ResultsThe administration of TBK for 4 weeks in DPN rats resulted in a significant decrease in FBG levels compared to untreated DPN rats. There was a significant increase in MNCV and SNCV in the DPN rats compared to untreated DPN rats. Serum level of MDA was significantly reduced while the activities of SOD and GSH-pX were significantly increased in the TBK treated DPN rats. TBK prevented DPN-induced increase in the serum levels of IL-6 and TNF-α.ConclusionThe results of this study demonstrate that the therapeutic effect of TBK on DPN rats may be associated with the antioxidative and anti-inflammatory responses.

Peroxynitrite and protein nitration in the pathogenesis of diabetic peripheral neuropathy

Peroxynitrite, a product of the reaction of superoxide with nitric oxide, causes oxidative stress with concomitant inactivation of enzymes, poly(ADP-ribosylation), mitochondrial dysfunction and impaired stress signalling, as well as protein nitration. In this study, we sought to determine the effect of preventing protein nitration or increasing peroxynitrite decomposition on diabetic neuropathy in mice after an extended period of untreated diabetes. C57Bl6/J male control and diabetic mice were treated with the peroxynitrite decomposition catalyst Fe(III) tetramesitylporphyrin octasulfonate (FeTMPS, 10 mg/kg/day) or protein nitration inhibitor (-)-epicatechin gallate (20 mg/kg/day) for 4 weeks, after an initial 28 weeks of hyperglycaemia. Untreated diabetic mice developed motor and sensory nerve conduction velocity deficits, thermal and mechanical hypoalgesia, tactile allodynia and loss of intraepidermal nerve fibres. Both FeTMPS and epicatechin gallate partially corrected sensory nerve conduction slowing and small sensory nerve fibre dysfunction without alleviation of hyperglycaemia. Correction of motor nerve conduction deficit and increase in intraepidermal nerve fibre density were found with FeTMPS treatment only. Peroxynitrite injury and protein nitration are implicated in the development of diabetic peripheral neuropathy. The findings indicate that both structural and functional changes of chronic diabetic peripheral neuropathy can be reversed and provide rationale for the development of a new generation of antioxidants and peroxynitrite decomposition catalysts for treatment of diabetic peripheral neuropathy.

Targeting apoptosis signalling kinase-1 (ASK-1) does not prevent the development of neuropathy in streptozotocin-induced diabetic mice.

Apoptosis signal-regulating kinase-1 (ASK1) is a mitogen-activated protein 3 kinase (MAPKKK/MAP3K) which lies upstream of the stress-activated MAPKs, JNK and p38. ASK1 may be activated by a variety of extracellular and intracellular stimuli. MAP kinase activation in the sensory nervous system as a result of diabetes has been shown in numerous preclinical and clinical studies. As a common upstream activator of both p38 and JNK, we hypothesised that activation of ASK1 contributes to nerve dysfunction in diabetic neuropathy. We therefore wanted to characterize the expression of ASK1 in sensory neurons, and determine whether the absence of functional ASK1 would protect against the development of neuropathy in a mouse model of experimental diabetes. ASK1 mRNA and protein is constitutively expressed by multiple populations of sensory neurons of the adult mouse lumbar DRG. Diabetes was induced in male C57BL/6 and transgenic ASK1 kinase-inactive (ASK1n) mice using streptozotocin. Levels of ASK1 do not change in the DRG, spinal cord, or sciatic nerve following induction of diabetes. However, levels of ASK2 mRNA increase in the spinal cord at 4 weeks of diabetes, which could represent a future target for this field. Neither motor nerve conduction velocity deficits, nor thermal or mechanical hypoalgesia were prevented or ameliorated in diabetic ASK1n mice. These results suggest that activation of ASK1 is not responsible for the nerve deficits observed in this mouse model of diabetic neuropathy.

Abstract 4653: Pharmacokinetics and biodistribution of eribulin mesylate, paclitaxel, and ixabepilone in mouse

Abstract Chemotherapy-induced peripheral neurotoxicity is a major clinical problem representing the dose-limiting side effect of many antineoplastic drugs. The degree and type of neuropathy incurred is dependent on the chemotherapeutic drug, dose intensity, and cumulative dose. In an earlier study by our group, we found that paclitaxel and ixabepilone, administered to mice at their respective MTDs, produced dose dependent deficits in caudal nerve conduction velocity, caudal amplitude and digital nerve amplitudes, as well as moderate to severe degenerative pathologic changes in dorsal root ganglia (DRG) and sciatic nerve (SN). In contrast, eribulin mesylate produced no significant deleterious effects on any nerve conduction parameter measured and caused milder, less frequent effects on morphology. Similar trends for eribulin to cause less neuropathy (including later onset and resolution) in the clinic have also been reported (Cigler and Vadhat, 2010; Jain and Cigler 2012, Vadhat et al., 2013). Differences in tubulin binding may, in part, potentially explain the differential effect (Perez et al 2009; Jordan et al 2005). Another potential explanation may reside in different pharmacokinetic (PK) and peripheral nerve distribution of these agents. To elucidate this, mice received a single dose of paclitaxel, ixabepilone or eribulin mesylate (30, 3 and 1.75mg/kg respectively, based on an MTD regimen), administered IV into the caudal vein. Plasma, DRG and SN samples were obtained at 0.25, 1, 3, 6, 8, 24, 48, and 72 h post dose. Tissues were homogenized and after extraction, concentrations of eribulin mesylate, ixabepilone and paclitaxel were determined in each matrix using LC/MS/MS. PK parameters were calculated using noncompartmental analysis in WinNonLin v 5.0.3. We report that all 3 chemotherapies rapidly penetrated both SN and DRG with maximal concentrations seen at, or within 15 mins after administration. Concentrations of all 3 agents in DRG and SN then declined by 30-50% over the first 1-3h and thereafter remained relatively stable in the tissue for up to 72 h, despite clearing rapidly from plasma to reach levels below quantification, (<5 ng/ml), by 24 h. Overall, dose normalized AUC in SN was similar for eribulin mesylate and ixabepilone and higher than for paclitaxel (1927 vs 2451 vs 793 ng/h/per g tissue, respectively). The ranking for the dose normalized AUC in DRG was eribulin mesylate < paclitaxel < ixabepilone (3894 vs 7217 vs 22800 ng/h/per g tissue, respectively). These findings suggest that peripheral nerve exposure is likely not the underlying mechanism for the differential propensity of eribulin mesylate, paclitaxel and ixabepilone to produce neurotoxicity. This work was financially supported by Eisai. Citation Format: Krystyna Wozniak, Ying Wu, Kenichi Nomoto, Bruce A. Littlefield, Christopher DesJardins, Edgar Schuck, Phil Saxton, Nancy Wong, Barbara S. Slusher. Pharmacokinetics and biodistribution of eribulin mesylate, paclitaxel, and ixabepilone in mouse. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4653. doi:10.1158/1538-7445.AM2014-4653

Matrix metalloproteinase-2 is downregulated in sciatic nerve by streptozotocin induced diabetes and/or treatment with minocycline: Implications for nerve regeneration

Minocycline is an inhibitor of matrix metalloproteinases (MMPs) and has been shown to have analgesic effects. Whilst increased expression of MMPs is associated with neuropathic pain, MMPs also play crucial roles in Wallerian degeneration and nerve regeneration. In this study we examined the expression of MMP-2, MMP-9 and tissue inhibitor of metalloproteinase (TIMP)-1/-2 in the sciatic nerve of control and streptozotocin-induced diabetic rats treated with either vehicle or minocycline by quantitative PCR and gelatin zymography. We assessed the effects of minocycline on nerve conduction velocity and intraepidermal nerve fibre (IENF) deficits in diabetic neuropathy and investigated the effects of minocycline or MMP-2 on neurite outgrowth from primary cultures of dissociated adult rat sensory neurons.We show that MMP-2 is expressed constitutively in the sciatic nerve in vivo and treatment with minocycline or diabetes leads to downregulation of MMP-2 expression and activity. The functional consequence of this is IENF deficits in minocycline-treated nondiabetic rats and an unsupportive microenvironment for regeneration in diabetes. Minocycline reduces levels of MMP-2 mRNA and nerve growth factor-induced neurite outgrowth. Furthermore, in vivo minocycline treatment reduces preconditioning-induced in vitro neurite outgrowth following a sciatic nerve crush. In contrast, the addition of active MMP-2 facilitates neurite outgrowth in the absence of neurotrophic support and pre-treatment of diabetic sciatic nerve substrata with active MMP-2 promotes a permissive environment for neurite outgrowth. In conclusion we suggest that MMP-2 downregulation may contribute to the regenerative deficits in diabetes. Minocycline treatment also downregulates MMP-2 activity and is associated with inhibitory effects on sensory neurons. Thus, caution should be exhibited with its use as the balance between beneficial and detrimental outcomes may be critical in assessing the benefits of using minocycline to treat diabetic neuropathy.

Differential expression of neuregulin-1 isoforms and downregulation of erbin are associated with Erb B2 receptor activation in diabetic peripheral neuropathy

BackgroundAberrant neuron/glia interactions can contribute to a variety of neurodegenerative diseases and we have previously demonstrated that enhanced activation of Erb B2, which is a member of the epidermal growth factor receptor (EGFR) family, can contribute to the development of diabetic peripheral neuropathy (DPN). In peripheral nerves, Erb B receptors are activated by various members of the neuregulin-1 (NRG1) family including NRG1 Type I, NRG1 Type II and NRG1 Type III to regulate Schwann cell (SC) growth, migration, differentiation and dedifferentiation. Alternatively, Erb B2 activity can be negatively regulated by association with the Erb B2-interacting protein, erbin. Since the effect of diabetes on the expression of NRG1 isoforms and erbin in peripheral nerve are unknown, the current study determined whether changes in NRG1 isoforms and erbin may be associated with altered Erb B2 signaling in DPN.ResultsSwiss Webster mice were rendered diabetic with streptozotocin (STZ) and after 12 weeks of diabetes, treated with erlotinib, an inhibitor of Erb B2 activation. Inhibition of Erb B2 signaling partially reversed several pathophysiologic aspects of DPN including a pronounced sensory hypoalgesia, nerve conduction velocity deficits and the decrease in epidermal nerve fiber innervation. We also observed a decrease of NRG1 Type III but an increase of NRG1 Type I level in diabetic sural nerves at early stage of diabetes. With disease progression, we detected reduced erbin expression and enhanced MAPK pathway activity in diabetic mice. Inhibition of Erb B2 receptor suppressed MAPK pathway activity in treated-diabetic sural nerves.ConclusionsThese results support that hyperglycemia may impair NRG1/Erb B2 signaling by disrupting the balance between NRG1 isoforms, decreasing the expression of erbin and correspondingly activating the MAPK pathway. Together, imbalanced NRG1 isoforms and downregulated erbin may contribute to the dysregulation of Erb B2 signaling in the development of DPN.

Endoplasmic reticulum stress contributes to prediabetic peripheral neuropathy

Growing evidence suggests that prediabetes and metabolic syndrome are associated with increased risk for the development of microvascular complications including retinopathy, nephropathy, and, most commonly, peripheral painful neuropathy and/or autonomic neuropathy. The etiology of these disabling neuropathies is unclear, and several clinical and experimental studies implicated obesity, impaired fasting glycemia/impaired glucose tolerance, elevated triglyceride and non-esterified fatty acids, as well as oxidative–nitrative stress. Endoplasmic reticulum stress resulting from abnormal folding of newly synthesized proteins and leading to the impairment of metabolism, transcriptional regulation, and gene expression, is emerging as a key mechanism of metabolic diseases including obesity and diabetes. We evaluated the role for this phenomenon in prediabetic neuropathy using two animal models i.e., Zucker (fa/fa) rats and high-fat diet fed mice which displayed obesity and impaired glucose tolerance in the absence of overt hyperglycemia. Endoplasmic reticulum stress manifest in upregulation of the glucose-regulated proteins BiP/GRP78 and GRP94 of unfolded protein response was identified in the sciatic nerve of Zucker rats. A chemical chaperone, trimethylamine oxide, blunted endoplasmic reticulum stress and alleviated sensory nerve conduction velocity deficit, thermal and mechanical hypoalgesia, and tactile allodynia. A selective inhibitor of eukaryotic initiation factor-2α dephosphorylation, salubrinal, improved glucose intolerance and alleviated peripheral nerve dysfunction in high-fat diet fed mice. Our findings suggest an important role of endoplasmic reticulum stress in the neurobiology of prediabetic peripheral neuropathy, and identify a new therapeutic target.

The Orally Active Glutamate Carboxypeptidase II Inhibitor E2072 Exhibits Sustained Nerve Exposure and Attenuates Peripheral Neuropathy

Peripheral neuropathy from nerve trauma is a significant problem in the human population and often constitutes a dose-limiting toxicity in patients receiving chemotherapy. (3-2-Mercaptoethyl)biphenyl-2,3-dicarboxylic acid (E2072) is a potent (K(i) = 10 nM), selective, and orally available inhibitor of glutamate carboxypeptidase II (GCPII). Here, we report that E2072 attenuates hyperalgesia and nerve conduction velocity deficits in preclinical rodent models of neuropathic pain and oxaliplatin-induced neuropathy. In the chronic constrictive injury model, orally administered E2072 reversed pre-existing thermal hyperalgesia in rats in a dose-dependent fashion with a minimally effective dose of 0.1 mg/kg/day. It is noteworthy that multiple days of dosing of E2072 were required before analgesia was realized even though GCPII inhibitory exposures were achieved on the first day of dosing. In addition, analgesia was found to persist for up to 7 days after cessation of dosing, consistent with E2072's pharmacokinetic profile and sustained exposure. Furthermore, in a chronic oxaliplatin-induced neuropathy model (6 mg/kg i.p. oxaliplatin twice weekly for 4 weeks), female BALB/c mice receiving daily oral E2072 at 1.0 and 0.1 mg/kg displayed no deficits in either caudal or digital velocity compared with significant deficits observed in mice treated with oxaliplatin alone (12 ± 3 and 9 ± 2%, respectively). Similar findings were seen with oxaliplatin-induced digital and caudal amplitude deficits. It is noteworthy that E2072 showed no interference with the antineoplastic efficacy of oxaliplatin in mice bearing leukemia (L1210), even at doses 100 times its neuroprotective/analgesic dose, indicating a selective effect on neuropathy. These data support the therapeutic utility of GCPII inhibitors in neuropathy and neuropathic pain.

Modulating molecular chaperones improves sensory fiber recovery and mitochondrial function in diabetic peripheral neuropathy

Quantification of intra-epidermal nerve fibers (iENFs) is an important approach to stage diabetic peripheral neuropathy (DPN) and is a promising clinical endpoint for identifying beneficial therapeutics. Mechanistically, diabetes decreases neuronal mitochondrial function and enhancing mitochondrial respiratory capacity may aid neuronal recovery from glucotoxic insults. We have proposed that modulating the activity and expression of heat shock proteins (Hsp) may be of benefit in treating DPN. KU-32 is a C-terminal Hsp90 inhibitor that improved thermal hypoalgesia in diabetic C57Bl/6 mice but it was not determined if this was associated with an increase in iENF density and mitochondrial function. After 16weeks of diabetes, Swiss Webster mice showed decreased electrophysiological and psychosensory responses and a >30% loss of iENFs. Treatment of the mice with ten weekly doses of 20mg/kg KU-32 significantly reversed pre-existing deficits in nerve conduction velocity and responses to mechanical and thermal stimuli. KU-32 therapy significantly reversed the pre-existing loss of iENFs despite the identification of a sub-group of drug-treated diabetic mice that showed improved thermal sensitivity but no increase in iENF density. To determine if the improved clinical indices correlated with enhanced mitochondrial activity, sensory neurons were isolated and mitochondrial bioenergetics assessed ex vivo using extracellular flux technology. Diabetes decreased maximal respiratory capacity in sensory neurons and this deficit was improved following KU-32 treatment. In conclusion, KU-32 improved physiological and morphologic markers of degenerative neuropathy and drug efficacy may be related to enhanced mitochondrial bioenergetics in sensory neurons.

Triglyceride, nonesterified fatty acids, and prediabetic neuropathy: role for oxidative–nitrosative stress

Peripheral neuropathy develops in human subjects with prediabetes and metabolic syndrome before overt hyperglycemia. The contributions of impaired glucose tolerance and insulin signaling, hypertriglyceridemia and/or increased nonesterified fatty acids (NEFA), and hypercholesterolemia to this condition remain unknown. Niacin and its derivatives alleviate dyslipidemia with a minor effect on glucose homeostasis. This study evaluated the roles of impaired glucose tolerance versus dyslipidemia in prediabetic neuropathy using Zucker fatty (fa/fa) rats and the niacin derivative acipimox, as well as the interplay of hypertriglyceridemia, increased NEFA, and oxidative–nitrosative stress. Sixteen-week-old Zucker fatty rats with impaired glucose tolerance, obesity, hyperinsulinemia, hypertriglyceridemia, hypercholesterolemia, and increased NEFA displayed sensory nerve conduction velocity deficit, thermal and mechanical hypoalgesia, and tactile allodynia. Acipimox (100mgkg−1day−1, 4weeks) reduced serum insulin, NEFA, and triglyceride concentrations without affecting glucose tolerance and hypercholesterolemia. It alleviated sensory nerve conduction velocity deficit and changes in behavioral measures of sensory function and corrected oxidative–nitrosative stress, but not impaired insulin signaling, in peripheral nerve. Elevated NEFA increased total and mitochondrial superoxide production and NAD(P)H oxidase activity in cultured human Schwann cells. In conclusion, hypertriglyceridemia and/or increased NEFA concentrations cause prediabetic neuropathy through oxidative–nitrosative stress. Lipid-lowering agents and antioxidants may find a use in the management of this condition.

LOX-1-mediated injury in sensory neurons in type 2 diabetes

Our laboratory recently identified several differentially expressed genes (DEG) through microarray analysis of the sciatic nerves of 24-week-old BKS db/db (diabetic) and db/+ (nondiabetic) mice. We noted prominent increases in lipid metabolism genes, consistent with another model, the high fat-fed C57BL6/J mouse, where we postulated that oxidized low-density lipoproteins (oxLDL) activate a receptor-mediated inflammatory pathway of neuronal injury. Our findings support clinical observations that plasma lipids associate with neuropathy, and suggest that glucose and lipids together will be responsible for the most aggressive development of diabetic neuropathy. A role for oxLDL-induced injury remains to be proved in vivo, and this is the goal of the present study. First, we performed biostatistical network analysis to identify molecular interactions of our LOX-1 gene of interest, Olr1. Next, nondiabetic BKSdb/+ and diabetic BKSdb/db littermates received intraperitoneal injections of LOX-1 neutralizing antibody or control non-immune IgG every 48 h from 6-12 wk of age. At 12 wk of age, blood and tissues were harvested for metabolic and neuropathy phenotyping. We confirmed our network analysis by demonstrating that NFkappaB, a gene closely linked to Olr1 in our network, was decreased in anti-LOX-1 treated mice. Most metabolic parameters (blood glucose, glycated hemoglobin, plasma triglycerides and cholesterol) and also the sciatic motor nerve conduction velocity deficit were unchanged in the presence of anti-LOX-1 treatment compared with control IgG. However, there was a striking preservation of the sural nerve conduction velocity in anti-LOX-1-treated mice, demonstrating that this receptor is involved in sural nerve injury in type 2 diabetes.

Beneficial Effects of PKF275-055, a Novel, Selective, Orally Bioavailable, Long-Acting Dipeptidyl Peptidase IV Inhibitor in Streptozotocin-Induced Diabetic Peripheral Neuropathy

1-[(2-adamantyl)amino]acetyl-2-cyano-(S)-pyrrolidine, monohydrochloride (PKF275-055), a vildagliptin analog, is a novel, selective, potent, orally bioavailable, and long-acting dipeptidyl peptidase IV inhibitor. We studied the effect of PKF275-055 administration on the prevention, protection, and treatment of diabetic neuropathy in the streptozotocin-induced diabetic rat. PKF275-055 improved body and muscle weight. Oral glucose tolerance tests showed a marked improvement in glucose metabolism under all treatment schedules. When tested in prevention and protection experiments, PKF275-055 completely averted the decrease of Na⁺/K⁺-ATPase activity and partially counteracted the nerve conduction velocity (NCV) deficit observed in untreated diabetic rats but had no effects on abnormal mechanical and thermal sensitivity. When used in a therapeutic setting, PKF275-055 induced a significant correction in the alteration in Na⁺,K⁺-ATPase activity and NCV present in untreated diabetics. Diabetic rats developed mechanical hyperalgesia within 2 weeks after streptozotocin injection and exhibited significantly longer thermal response latencies. It is noteworthy that PKF275-055 treatment restored mechanical sensitivity thresholds by approximately 50% (p < 0.01) and progressively improved the alteration in thermal responsiveness. In conclusion, PKF275-055 showed an anabolic effect, improved oral glucose tolerance, and counteracted the alterations in Na⁺,K⁺-ATPase activity, NCV, and nociceptive thresholds in diabetic rats. The present data support a potential therapeutic effect of PKF275-055 in the treatment of rodent diabetic neuropathy.

Comparison of Neuropathy-Inducing Effects of Eribulin Mesylate, Paclitaxel, and Ixabepilone in Mice

Chemotherapy-induced neurotoxicity is a significant problem associated with successful treatment of many cancers. Tubulin is a well-established target of antineoplastic therapy; however, tubulin-targeting agents, such as paclitaxel and the newer epothilones, induce significant neurotoxicity. Eribulin mesylate, a novel microtubule-targeting analogue of the marine natural product halichondrin B, has recently shown antineoplastic activity, with relatively low incidence and severity of neuropathy, in metastatic breast cancer patients. The mechanism of chemotherapy-induced neuropathy is not well understood. One of the main underlying reasons is incomplete characterization of pathology of peripheral nerves from treated subjects, either from patients or preclinically from animals. The current study was conducted to directly compare, in mice, the neuropathy-inducing propensity of three drugs: paclitaxel, ixabepilone, and eribulin mesylate. Because these drugs have different potencies and pharmacokinetics, we compared them on the basis of a maximum tolerated dose (MTD). Effects of each drug on caudal and digital nerve conduction velocity, nerve amplitude, and sciatic nerve and dorsal root ganglion morphology at 0.25 × MTD, 0.5 × MTD, 0.75 × MTD, and MTD were compared. Paclitaxel and ixabepilone, at their respective MTDs, produced significant deficits in caudal nerve conduction velocity, caudal amplitude and digital nerve amplitudes, as well as moderate to severe degenerative pathologic changes in dorsal root ganglia and sciatic nerve. In contrast, eribulin mesylate produced no significant deleterious effects on any nerve conduction parameter measured and caused milder, less frequent effects on morphology. Overall, our findings indicate that eribulin mesylate induces less neuropathy in mice than paclitaxel or ixabepilone at equivalent MTD-based doses.

Poly(ADP-ribose)polymerase inhibition counteracts renal hypertrophy and multiple manifestations of peripheral neuropathy in diabetic Akita mice

Poly(ADP-ribose) polymerase (PARP) activation has been implicated in the pathogenesis of diabetic complications, including nephropathy and peripheral neuropathy. This study aimed at evaluating the manifestations of both complications in diabetic Akita mice, a model of Type 1 (insulin-dependent) diabetes, and their amenability to treatment with the potent PARP inhibitor, 10-(4-methyl-piperazin-1-ylmethyl)-2H-7-oxa-1,2-diaza-benzo[de] anthracen-3-one (GPI-15427). Male non-diabetic C57Bl6/J and diabetic C57Bl/6-Ins2Akita/J (Akita) mice were maintained with or without treatment with GPI-15427, 30 mg/kg/day, for 4 weeks starting from 16 weeks of age. Sixteen week-old Akita mice displayed sensory nerve conduction velocity (SNCV) deficit, whereas the motor nerve conduction velocity (MNCV) tended to decrease, but the difference with controls did not achieve statistical significance. They also developed thermal and mechanical hypoalgesia and tactile allodynia. SNCV deficit, mechanical hypoalgesia, and tactile allodynia progressed with age whereas the severity of thermal hypoalgesia was similar in 16- and 20-week-old Akita mice. PARP inhibition alleviated, although it did not completely reverse, SNCV deficit, thermal and mechanical hypoalgesia and tactile allodynia. Sixteen-week-old Akita mice displayed MNCV deficit (41.3±2.5 vs. 51.0±1.2 m/sec in non-diabetic controls, P<0.01), axonal atrophy of myelinated fibers, kidney hypertrophy, and albuminuria. MNCV slowing, axonal atrophy, and kidney hypertrophy, but not albuminuria, were less severe in GPI-15427-treated age-matched Akita mice. Neuroprotective and nephroprotective effects of PARP inhibition were not due to alleviation of diabetic hyperglycemia, or peripheral nerve p38 mitogen-activated protein kinase activation. GPI-15427 did not affect any variables in control mice. In conclusion, the findings support an important role for PARP activation in diabetic peripheral neuropathy and kidney hypertrophy associated with Type 1 diabetes, and provide rationale for development and further studies of PARP inhibitors, for the prevention and treatment of these complications.

Interactions between the 12/15‐lipoxygenase pathway and MAPK activation in diabetic peripheral neuropathy

Both MAPK activation and, recently, the 12/15-lipoxygenase (12/15-LO) pathway, have been implicated in the pathogenesis of diabetic peripheral neuropathy (DPN). We evaluated the relationship between the two mechanisms using the 12/15-LO inhibitor cinnamyl-3,4-dihydroxy-á-cyanocinnamate (CDC) and 12/15-LO−/− mice. Male wild-type (WT) and 12/15-LO−/− mice were made diabetic with streptozotocin. Wild-type mice with 12–14 wk duration of STZ diabetes displayed motor and sensory nerve conduction velocity deficits, thermal hypoalgesia, tactile allodynia, and intraepidermal nerve fiber loss. They also displayed 12/15-LO overexpression, 12(S)-HETE accumulation (a sign of increased 12/15-LO activity), and increased phosphorylation of p38, ERK, and SAP/JNK in the sciatic nerve. 12/15-LO inhibition with CDC (8 mgkg-1d-1, s.c., for 4 wks after initial 12 wks without treatment) or 12/15-LO gene deficiency alleviated peripheral nerve dysfunction and reduced p38 and ERK phosphorylation, without affecting SAP/JNK phosphorylation or intraepidermal nerve fiber loss, a sign of small sensory nerve fiber degeneration. In conclusion, the findings suggest interaction between the 12/15-LO pathway and p38 and ERK, but not SAP/JNK, in the development of peripheral nerve dysfunction characteristic for DPN. Neither 12/15-LO nor MAPK activation contribute to diabetes-induced intraepidermal nerve fiber loss.

PARP inhibition alleviates diabetes-induced systemic oxidative stress and neural tissue 4-hydroxynonenal adduct accumulation: Correlation with peripheral nerve function

This study evaluated the role of poly(ADP-ribose) polymerase (PARP) in systemic oxidative stress and 4-hydoxynonenal adduct accumulation in diabetic peripheral neuropathy. Control and streptozotocin-diabetic rats were maintained with or without treatment with the PARP inhibitor, 1,5-isoquinolinediol, 3mg kg−1 day−1, for 10weeks after an initial 2weeks. Treatment efficacy was evaluated by poly(ADP-ribosyl)ated protein content in peripheral nerve and spinal cord (Western blot analysis) and dorsal root ganglion neurons and nonneuronal cells (fluorescence immunohistochemistry), as well as by indices of peripheral nerve function. Diabetic rats displayed increased urinary isoprostane and 8-hydroxy-2′-deoxyguanosine excretion (ELISA) and 4-hydroxynonenal adduct accumulation in endothelial and Schwann cells of the peripheral nerve, neurons, astrocytes, and oligodendrocytes of the spinal cord and neurons and glial cells of the dorsal root ganglia (double-label fluorescence immunohistochemistry), as well as motor and sensory nerve conduction velocity deficits, thermal hypoalgesia, and tactile allodynia. PARP inhibition counteracted diabetes-induced systemic oxidative stress and 4-hydroxynonenal adduct accumulation in peripheral nerve and spinal cord (Western blot analysis) and dorsal root ganglion neurons (perikarya, fluorescence immunohistochemistry), which correlated with improvement of large and small nerve fiber function. The findings reveal the important role of PARP activation in systemic oxidative stress and 4-hydroxynonenal adduct accumulation in diabetic peripheral neuropathy.