Abstracts of the International Diabetes Federation Congress Fifth Diabetic Neuropathy Satellite Symposium

s Journal of the Peripheral Nervous System 5:169–188 (2000) 170 Alpha-lipoic acid (LPA) has marked effects in experimental diabetic neuropathy. Treatment for 2 weeks after 8 weeks of diabetes in rats corrected motor and sensory conduction velocity and endoneurial perfusion deficits with an ED 50 of approximately 40 mg/kg. There was no difference in efficacy between Rand S-enantiomers of LPA. When combined with the n-6 essential fatty acid, gamma-linolenic acid (GLA), either as a drug mixture or joint compound, there was a synergistic effect on neurovascular function, resulting in a 6-fold increase in treatment efficacy. LPA has wide-ranging actions on markers of endothelial damage, the coagulation system and lipids that are important risk factors for neuropathy and vascular disease in patients. Thus, 2 months of diabetes in rats caused 2.5to 5.4-fold elevations of von Willebrand factor, factor VII, triglycerides and LDL cholesterol, which were 41-64% attenuated by 2 weeks of LPA treatment. Vascular effects are not restricted to endoneurial perfusion. Thus, the time taken for a fixed-volume tail-bleed was 4.2-fold greater with diabetes and this was 79% attenuated by LPA treatment. Nitric oxide and endothelium-derived hyperpolarizing factor dependent vasodilation were markedly reduced by diabetes in corpus cavernosum and mesenteric vasculature, LPA giving a high degree of protection. The function of small nerve fibres, exemplified by the nitrergic vasodilator innervation to corpus cavernosum, is greatly diminished by diabetes, high-dose LPA having a protective effect. When LPA was combined with GLA, as for large fibres, there was a marked synergy for both the innervation and the endothelium of corpus cavernosum. Thus LPA alone, and particularly combined with GLA, has a broad spectrum of action highly relevant for the treatment of diabetic neuropathy and micro and macrovascular disease. SIGNAL TRANSDUCTION BY STRESS-ACTIVATED MAP KINASES Davis RJ. Howard Hughes Medical Institute & Program in Molecular Medicine, University of Massachusetts Medical School, 373 Plantation Street, Worcester, MA 01605. The JNK group of stress-activated MAP kinases consists of ten protein kinases that phosphorylate the NH 2 -terminal activation domain of c-Jun on Ser-63 and Ser-73 causing increased transcriptional activity. JNK protein kinase activity is increased in response to treatment of cells with pro-inflammatory cytokines or exposure to environmental stress. Activated JNK is phosphorylated on Thr and Tyr within the tripeptide motif Thr-Pro-Tyr located in kinase subdomain VIII. Mutational analysis demonstrates that JNK activation requires the phosphorylation of both Thr and Tyr within this motif. This phosphorylation is mediated by dual specificity protein kinases, including MKK4 and MKK7. The function of the JNK signaling pathway has been studied using a combination of biochemical and genetic approaches. Genetic analysis of JNK signaling in Drosophila demonstrates that the JNK signaling pathway is required for early embryonic morphogenesis. Similarly, disruption of the JNK signaling pathway in mice using homologous recombination demonstrates that JNK signaling is required for embryonic viability. In contrast, mice with genetically engineered selective defects in JNK signaling are viable, but exhibit changes in stress-induced gene expression and apoptosis. These studies provide insight into the role of the JNK stress-activated MAP kinase pathway in the cellular response to environmental stress, including radiation. THE RISK FACTORS FOR DIABETIC COMPLICATIONS Dyck PJ. Mayo Clinic, 200 First Street SW, Rochester, MN 55905. The major late complications of diabetes mellitus can be classified into those attributable to atherosclerosis (heart attacks, stroke, and peripheral vascular disease) and to microvascular disease (diabetic polyneuropathy [DPN], retinopathy, and nephropathy). The risk factors for diabetic polyneuropathy had not previously been adequately assessed by: 1) assessing representative population-based cohorts; 2) assessing DPN comprehensively and quantitatively; 3) expressing nerve tests as a percentile and normal deviate value correcting for anthropometric characteristics as based on study of a randomly selected normal subject cohort from which neurologic disease had been excluded; 4) assessing risk factors serially over time; and 5) performing multivariate analysis of risk factors. For DPN we have found that the risk factors are: 1) markers of diabetic microvessel disease (diabetic retinopathy, 24-hour proteinuria, or 24-hour microalbuminuria); 2) chronic hyperglycemic exposure; and 3) type of DM. Excluding markers of microvessel disease and type of DM we found that the highest correlation coefficients were obtained by glycated hemoglobin (GH) 3 duration of DM 1/2 3 18/age of onset DM (lowest score 5 1) 3 total plasma antioxidant activity. Similarly, major determinants for diabetic retinopathy and nephropathy also are GH 3 duration of DM. These results provide strong supportive evidence that the product of chronic hyperglycemic (especially GH) exposure 3 duration of DM are the main determinants of the diabetic complications of retinopathy, nephropathy, and neuropathy. NEUROTROPHINS IN DIABETIC NEUROPATHY Fernyhough P. Division of Neuroscience, School of Biological Sciences, University of Manchester, UK. The recent Genentech/Roche sponsored FDA phase III clinical trial of nerve growth factor (NGF) for the treatment of diabetic neuropathy failed. The aim of this talk is to discuss the reasons for this failure and based upon the lessons learned identify if there remain any other viable avenues for studies on neurotrophins in diabetic neuropathy. The presentation will briefly introduce the basic biology of the neurotrophins and their putative role in the aetiology and/or treatment of diabetic neuropathy. Some details of the NGF clinical trials will be presented and reasons for failure assessed. The main part of the talk will focus on future strategies for neurotrophin work with an emphasis on the possible roles of neurotrophin-3 and brain-derived neurotrophic factor. Alternative approaches based upon regulation of endogenous neurotrophin expression and use of neurotrophin trk receptor agonists will also be discussed. The aim of the talk is to show that lessons learned from the failed NGF trial should lead to a re-invigoraAbstracts Journal of the Peripheral Nervous System 5:169–188 (2000)s Journal of the Peripheral Nervous System 5:169–188 (2000) 171 tion of studies directed at determining alternative methods for optimising neurotrophic support of sensory neurones in diabetes. THE MOLECULAR MECHANISM OF THE ORGANELLE TRANSPORTS IN NEURONS; KINESIN SUPERFAMILY PROTEINS (KIFS), STRUCTURE, GENE, AND FUNCTION Hirokawa N. Department of Cell Biology and Anatomy, University of Tokyo, Graduate School of Medicine. Many proteins are transported to their proper destination as membranous organelles or protein complexes after synthesis. Especially in highly polarized cells such as epithelial cells and neurons this transport is very important for the proper targeting of proteins to distinct parts of cells. Actually we visualized dynamics of membrane organelles carrying certain kinds of proteins in living cells using GFP technology. In order to understand the mechanism of this transport we identified 26 new members of microtubule based molecular motors, KIFs. Using multiple molecular cell biological and molecular genetical approaches we have characterized these new members. In this lecture I will focus on some of KIF members such as KIF1A, 1B, 3, C2 and 17. KIF1A is the fastest (1.5 m m/sec) anterograde monomeric motor for transport of precursor of synaptic vesicles and essential for neuronal function and survival while KIF1B is a unique monomeric anterograde motor (0.5 m m/sec) for transport of mitochondria. KIF3A and KIF3B, expressed ubiquitously, form a heterodimer associated with a protein, KAP3, and work as a new anterograde transporter for membranous organelles which are different from synaptic vesicle precursors and essential for neurite extension. KIFC2 is a neuron specific Cterminal motor domain type KIF transporting multivesicular body-like organelles to dendrites. Very recently we have identified new KIF17 which is expressed specifically in neurons and transport NMDA receptors in nerve dendrites. Thus, our studies revealed that transport of important functional molecules as various kinds of membrane organelles and protein complexes is accomplished very precisely by these new molecular motors. DIABETIC NEUROPATHY: PATHOGENETIC MECHANISMS AND TREATMENT IMPLICATIONS Low PA 1 , Nickander KK 1 , Kihara M 1 , Schmelzer JD 1 , Nagamatsu M 1 , Sasaki H 1 , Tritschler H-J 2 . 1 Dept of Neurology, Mayo Clinic, 2 Rochester MN, USA, 2 Asta Medica, FRG. Chronic hyperglycemia underlies the pathogenesis of diabetic neuropathy (DN). Recent research emphasizes the roles of ischemia-hypoxia, oxidative stress, glycation, v 6 fatty acid metabolism and the interacting roles of excessive lipolysis, hyperactive polyol pathway and PKC activity and deficiencies of growth factors. In DN, oxidative stress is due primarily to nerve ischemia and auto-oxidation/glycation, but all the above mechanisms are interactive and all can cause oxidative stress. Oxidative stress appears to be a particularly important mechanism in the pathogenesis of DN. Antioxidant enzymes are reduced in peripheral nerve and are further reduced in DN. That lipid peroxidation will cause neuropathy is supported by the development of neuropathy de novo when normal nerve is rendered a -tocopherol deficient. Recent studies emphasize the prominent pathological alterations in dorsal root ganglion (DRG) and nerve roots of experimental DN. DRG mitochondrial cytopathy is suggested to result. Histochemical studies of DRG shows defective respiratory enzyme activity, TUNEL staining and caspase-3 immunostaining, suggesting apoptosis, especially apoptosis lente, a process that results in impairment of funtion with modest and slow cellular loss. These pathogenetic observations have spawned clinical trials with aldose reductase inhibitors, growth factor, protein glycation, g -linolenic acid, PKCb , and a -lipoic acid. Clinical trials on human diabetic neuropathy have been underpowered, especially with the short duration of the studies. Treatment with antioxidants have been especially efficacious in experimental diabetes, mainly in a preventive role. In established human DN, a minimum of 2 years is likely required to demonstrate benefit. A 4 year study of a -lipoic acid in human DN is underway. AUTONOMIC NEUROPATHY IN SORBITOL DEHYDROGENASE INHIBITOR (SDI)-TREATED STREPTOZOTOCIN-DIABETIC RATS Schmidt RE. Washington University School of Medicine, Saint Louis, MO USA. We have developed an animal model of diabetic autonomic neuropathy characterized by neuroaxonal dystrophy (NAD) involving ileal mesenteric nerves and prevertebral sympathetic superior mesenteric ganglia (SMG) in chronic streptozotocin-diabetic rats. We have examined the effect of SDI-158, which interrupts the conversion of sorbitol to fructose (and reactions dependent on the second step of the polyol pathway), on NAD in control and diabetic rats. SDItreatment (SDI-Rx) did not produce NAD in control animals despite the fact that sciatic nerve sorbitol levels reached the same levels as untreated diabetic animals. SDI-Rx resulted in a dramatically increased frequency of NAD in ileal mesenteric nerves and SMG. SDI-Rx diabetic rats developed lesions prematurely, after only one month of diabetes, and in greater numbers than untreated diabetics. SDI-Rx for 1,2,3 and 6 months showed a statistically significant but progressively smaller effect on NAD. The ultrastructural appearance of SDIRx ganglia and mesenteric nerves were identical to that previously reported in long-term untreated diabetics; however, short term SDI-Rx resulted in less compacted tubulovesicular elements in the SMG. Three month SDI-Rx superior cervical ganglia failed to develop NAD in the same animals in which large numbers of lesions were found in the SMG. Dystrophic axons involved ileal mesenteric nerves while sparing those to the jejunum. Dystrophic tyrosine hydroxylase immunoreactive sympathetic axons were distributed to myenteric and submucosal ganglia within the gut wall but not to the mesenteric vasculature. Therefore, although SDI-Rx diabetic rats show an exaggerated severity and accelerated time course of NAD compared to untreated diabetics, lesion appearance, immunoreactivity and distribution are comparable to those found in untreated STZ-diabetic rats of much longer durations. Abstracts Journal of the Peripheral Nervous System 5:169–188 (2000)s Journal of the Peripheral Nervous System 5:169–188 (2000) 172 EVALUATION OF DIABETIC NEUROPATHY BY SKIN BIOPSY Yasuda H. Third Department of Medicine, Shiga University of Medical Science, Otsu, Shiga 520-2192, Japan. Instead of nerve biopsy which is most reliable method for diagnosing diabetic neuropathy but rather harmful, we have examined the usefulness of skin biopsy to evaluate the grade of diabetic neuropathy and therapeutic effects of given compounds. Nerve fibers immunostained by antibodies against protein gene product (PGP)9.5 and labeled with streptavidin fluorescein isothiocyanate and measured under confocal laser scanning microscopy were significantly shorter in the epidermis and dermis and around sweat glands in diabetic patients than in healthy subjects. Sural nerve conduction velocity was significantly correlated with dermal nerve fiber length (NFL) (p , 0.05) in diabetic patients. Patients with higher aldose reductase (AR) level of erythrocytes ( . 10.8 ng/ Hg) showed shorter dermal NFL than those with lower AR level ( , 10.8)(p , 0.05). Using this technique, we evaluated the therapeutic effect of AR inhibitor (ARI) epalrestat during approximately 13 months in diabetic patients with neuropathy (19, epalrestat-treated; 12, control). The number of patients whose percent change of dermal NFL exceeded its mean 6 2SD in the control group was significantly higher in the ARI group than in the control group (p , 0.05, 8/9 vs 1/12). These results suggest that quantitation of cutaneous nerves using biopsied skin samples may provide important information about the severity of diabetic neuropathy and that epalrestat can elongate dermal unmyelinated nerve fibers in at least part of diabetic patients. CURRENT EVIDENCE FOR TREATING DIABETIC NEUROPATHY Ziegler D. German Diabetes Research Institute at the Heinrich Heine University, Düsseldorf, Germany. At least one of three diabetic patients is affected by polyneuropathy which represents a major health problem, as it may present with partly excruciating neuropathic pain and is responsible for substantial morbidity, increased mortality, and impaired quality of life. Treatment is based on four cornerstones: 1.) causal treatment aimed at (near)-normoglycaemia, 2.) treatment based on pathogenetic mechanisms, 3.) symptomatic treatment, and 4.) avoidance of risk factors and complications. Recent experimental studies suggest a multifactorial pathogenesis of diabetic neuropathy. Several pathogenetic mechanisms are being discussed which, however, in contrast to previous years are no longer regarded as separate hypotheses but in the first place as a complex interplay with multiple interactions between metabolic and vascular factors. From the clinical point of view it is important to note that, based on these pathogenetic mechanisms, therapeutic approaches could be derived, some of which are currently being evaluated in clinical studies. These treatments include the inhibition of the increased flux through the polyol pathway by aldose reductase inhibitors, correction of the deficits in essential fatty acid and prostanoid metabolism by substitution of g -linolenic acid contained in evening primrose oil, administration of antioxidants ( a -lipoic acid) to reduce the enhanced formation of reactive oxygen species that induce increased oxidative stress, improvement in endoneurial blood flow and resulting hypoxia by vasodilating agents such as ACE inhibitors and protein kinase C (PKC) ß inhibitors, neurotrophic support by administration of NGF, inhibition of non-enzymatic glycation and formation of AGEs, correction of C-peptide deficiency by substitution of C-peptide, and immunosuppressive treatment. Currently only a -lipoic acid is available for treatment in several countries. Randomized controlled clinical trials using this agent have demonstrated that 1.) short-term treatment for 3 weeks (600 mg/day i.v.) reduces the chief symptoms and deficits due to diabetic polyneuropathy. 2.) Oral treatment for 4-7 months tends to ameliorate neuropathic deficits and cardiac autonomic neuropathy. 3.) Preliminary data over 2 years indicate possible long-term improvement in motor and sensory nerve conduction in the lower limbs. Clinical and postmarketing surveillance studies have revealed a highly favorable safety profile of the drug. Despite the recently accelerating publication rate for controlled trials demonstrating significant pain relief with several agents, the pharmacologic treatment of chronic painful diabetic neuropathy remains a challenge for the physician. A simple measure of therapeutic efficacy (number needed to treat: NNT) permits to estimate the risk-benefit-ratio for each agent on the basis of the available controlled trials. In recent meta-analyses, the NNTs have been calculated for several drugs employed in the treatment of painful diabetic neuropathy which may serve the physician in deciding for the individual treatment. Epidemiological data indicate that not only increased alcohol consumption but also the traditional cardiovascular risk factors such as hypertension, smoking, and cholesterol play a role in development and progression of diabetic neuropathy and, hence, need to be prevented or treated. METABOLIC VS. VASCULAR ABNORMALITIES IN DIABETIC NEUROPATHY