CHAPTER 9 • PATHOPHYSIOLOGY OF DIABETIC NEUROPATHY 89 Insulin like growth factors (IGFs) In cultured Schwann cells and the STZ-diabetic rat, IGF-1 demonstrates a pro- tective effect via PI 3-kinase, in preventing glucose-mediated neuronal and Schwann cell apoptosis. Both the STZ-diabetic and BB/W rat develop severe hyperglycaemia and a deficiency in circulating IGF-I levels with neuroaxonal dystrophy (NAD) in the nerve terminals of the prevertebral sympathetic gan- glia and the distal portions of noradrenergic ileal mesenteric nerves. In contrast the Zucker Diabetic Fatty (ZDF) rat, an animal model of type 2 diabetes, also develops severe hyperglycaemia but maintains normal levels of plasma IGF-I and does not demonstrate NAD in sympathetic ganglia and ileal mesenteric nerves. However, IGF-I and IGF-I receptor mRNA levels have not been shown to differ in the sural nerve of diabetic patients compared with control subjects. C-Peptide In experimental studies C-peptide has demonstrated effects on Na(+)/K(+)- ATPase activity, endothelial nitric oxide synthase, expression of neurotrophic factors and regulation of molecules controlling degeneration of the nodal apparatus in Type 1 diabetic nerves, as well as DNA binding of transcription factors and modulation of apoptotic phenomena. These findings have recent- ly been effectively translated into benefits in patients with Type 1 diabetes with the demonstration of a significant improvement in sural sensory nerve con- duction velocity and vibration perception but without a benefit in either cold or heat perception after 12 weeks of daily subcutaneous C-peptide treatment. Vascular endothelial growth factor (VEGF) VEGF was originally discovered as an endothelial-specific growth factor with a predominant role in angiogenesis. However, recent observations indicate that VEGF also has direct effects on neurones and glial cells stimulating their growth, survival and axonal outgrowth. Thus with its potential for a dual impact on both the vasculature and neurones it could represent an important therapeutic inter- vention in diabetic neuropathy. Although immunohistochemistry of sciatic nerves and dorsal root ganglia from STZ-diabetic rats demonstrates intense VEGF staining in cell bodies and nerve fibers with no or very little VEGF expres- sion in controls. Intramuscular gene transfer of plasmid DNA encoding VEGF- 1 or VEGF-2 in the STZ-diabetic rat and alloxan diabetic rabbit results in restora- tion of nerve vascularity, blood flow and both large and small fibre dysfunction. Thus there is an intrinsic capacity to up-regulate VEGF but this appears insuffi- cient and may require exogenous delivery possibly via gene therapy. A phase I/II, double-blind, placebo controlled study to evaluate the safety and impact of phVEGF165 gene transfer in patients with diabetic neuropathy is currently underway and will involve 192 patients over a period of four years. SECTION II • DIABETIC NEUROPATHIES 90 IMMUNE MECHANISMS Studies suggest that sera from type 2 diabetic patients with neuropathy con- tain an autoimmune immunoglobulin that induces complement-independ- ent, calcium-dependent apoptosis in neuronal cells. The expression of these cytotoxic factors has been related to the severity of neuropathy and the type of neuronal cell killed. Thus it has been suggested that such toxic factors may contribute to diabetic neuropathy by acting in concert with hyperglycaemia to damage sensory/autonomic neurones. FURTHER READING Dyck PJ, Davies JL, Wilson DM, Service FJ, Melton LJ 3rd, O’Brien PC. Risk factors for severity of diabetic polyneuropathy: intensive longitudinal assessment of the Rochester Diabetic Neuropathy Study cohort. Diabetes Care 1999; 22: 1479–1486. Eichberg J. Protein kinase C changes in diabetes: is the concept relevant to neuropathy? Int Rev Neurobiol 2002; 50: 61–82. Gaede P, Vedel P, Larsen N, Jensen GV, Parving HH, Pedersen O. Multifactorial intervention and cardiovascular disease in patients with type 2 diabetes. N Engl J Med 2003; 348: 383–393. CURRENT ISSUES • Translational studies in diabetic patients continue to generate much debate and controversy over the cause(s) and treatment of human diabetic neuropathy. • Recent studies in patients with impaired glucose tolerance (IGT) suggest that even minor degrees of glucose dysmetabolism may lead to the development of neuropathy. • A meta-analysis of 19 randomized controlled trials of ARIs was disappointing as it demonstrated a small but statistically significant reduction in decline of motor NCV without benefit in sensory nerves. • Single-nucleotide polymorphisms of the genes for mitochondrial (SOD2) and extracellular (SOD3) superoxide dismutases may confer an increased risk for the development of neuropathy. • 1, 2-diacylglycerol (DAG) induced activation pf protein kinase C (PKC), in particular PKC-β has been proposed to play a major role in diabetic neuropathy. • Conventional risk factors for macrovascular disease such as deranged lipids and hypertension are important in the pathogenesis of diabetic neuropathy. • In cultured Schwann cells and the STZ-diabetic rat, IGF-1 prevents glucose- mediated neuronal and Schwann cell apoptosis. • VEGF was originally discovered as an endothelial-specific growth factor. CHAPTER 10 EPIDEMIOLOGY AND NATURAL HISTORY OF DPN Rayaz A. Malik MB.ChB, PhD, MRCP 91 INTRODUCTION Epidemiological studies of diabetic peripheral neuropathy (DPN) have pro- duced widely varying figures on incidence and prevalence. There are several reasons for this and include differing criteria to define neuropathy and the use of different populations i.e. clinic versus hospital patients. Furthermore, the clinical complexity of DPN due to the relative involvement of sensory motor and autonomic fibres has led to the use of multiple assessments to define DPN. These have commonly included varying combinations of positive and negative symptoms, neurological deficits derived from a clinical examination, quanti- tative sensory tests and electrophysiology. To attempt to draw some valid comparisons between different studies each of the end points utilized will be evaluated separately. The incidence or prevalence will be defined and the nat- ural history and risk factors for the end point considered. POSITIVE SENSORY SYMPTOMS (PAINFUL NEUROPATHY) Positive sensory symptoms arise spontaneously or as a response to stimuli and they may be divided into painful and non-painful categories. Table 10.1 presents the long list of positive sensory symptoms, which have been utilized in different studies and provides an insight into the difficulties of how differ- ent symptoms may be recorded and interpreted. Table 10.1 Descriptions of positive neuropathic sensory symptoms. J Neurolog Sci 2001; 189: 3–5. Descriptions of positive neuropathic sensory symptoms Non-painful Painful thick prickling stiff tingling asleep knife-like prickling electric shock-like tingling squeezing constricting hurting burning freezing throbbing allodynia* hyperalgaesia * Allodynia: the perception of pain from a non-noxious stimulus. Vascular Complications of Diabetes: Current Issues in Pathogenesis and Treatment, Second Edition Edited by Richard Donnelly, Edward Horton Copyright © 2005 by Blackwell Publishing Ltd SECTION II • DIABETIC NEUROPATHIES 92 A prevalence of 27% was demonstrated in the only population-based study to date and it employed questionnaires to define symptoms according to pain, tingling, numbness, and the inability to feel hot or cold. Other clinic or hospital based studies utilizing similar symptom profiles have reported a prevalence of painful symptoms which varies from 3–30%, although their focus was more heavily on pain. Data on the natural history of painful neu- ropathy are conflicting, with one study showing a decrease in the intensity of symptoms with worsening of quantitative measures of nerve function, whilst another study found an improvement of pain with improvement of sensory function in individuals treated with continuous subcutaneous insulin infu- sions (CSII). Of course CSII has been shown to have a direct effect on painful symptoms via decreasing glucose flux. The data on remission of symptoms over time are inconsistent, with one study showing an overall reduction in the severity of pain scores but without any full remissions, whilst the majority of other studies have observed remissions. Despite the large number of studies in patients with painful neuropathy the risk factors remain ill defined. Thus in a population based study the degree of hyperglycaemia, hypertension and diabetes duration were deemed important, but in a clinic based study, symptoms were related to diabetes duration but not HbA1 C . NEGATIVE SENSORY SYMPTOMS (HYPOAESTHETIC NEUROPATHY) Since it is well recognized that patients tend to underestimate their degree of insensitivity, there has been more reliance on quantitative sensory testing than on symptomatology in studies of hypoesthesia. The prevalence of hypoesthesia may vary greatly according to the criteria used to define abnor- mality. In a study that utilized three different quantitative sensory measure- ments in the same individuals, the prevalence of abnormalities varied from 8–34%, highlighting the impact of different tests on the outcome data gener- ated. The site chosen to measure the deficit is also an important factor as the presence of hypoesthesia increases substantially as measurements become more distal. Whilst vibration perception thresholds may be elevated in chil- dren with diabetes and also in adults at diagnosis of type 2 diabetes, age requires careful consideration since normal values increase markedly with age, especially after the 6 th decade. In patients with type 1 diabetes followed from diagnosis, abnormalities of thermal thresholds and electrophysiology preceded any abnormality of vibration threshold in the first five years after diagnosis. Early in the course of type 2 diabetes, small but statistically signif- icant increases of vibration and thermal thresholds have been observed over two years. A study that utilized the 10 g monofilament apparently showed no CHAPTER 10 • EPIDEMIOLOGY AND NATURAL HISTORY OF DPN 93 progression of neuropathy in the first few years of diabetes. This simply reflects that the 10 g monofilament is neither sensitive enough to detect early loss of sensation nor to detect a small change in progressive loss of sensation. This study highlights the inappropriate use of the 10g monofilament and shows that it is useful only to detect those at risk of foot ulceration i.e. those with severe neuropathy. Progression appears to be more rapid once decreased sensation appears and has been consistently related to diabetes duration, degree of hyperglycaemia, and height. COMBINED ASSESSMENTS Studies which have utilized various combinations of positive and negative symptoms, quantitative sensory testing, abnormalities of the neurological exam and electrophysiology have generally produced higher estimates of the preva- lence of neuropathy. In one of the largest and earliest epidemiological studies the prevalence of neuropathy was greater than 40% after 25 years of known dia- betes duration. The EURODIAB IDDM Complications Study provided an overall prevalence for neuropathy of 28%. However, among the 27 centres included in the study, the prevalence ranged from less than 20% in several cen- tres to over 50% in two centres. This highlights the variability in interpreting examination findings in different countries and also that the patients chosen for study in the different centres came from differing clinical populations. In the Diabetes Control and Complications Trial (DCCT) ‘clinically detectable’ neu- ropathy was found in 39% of the participants who had type 1 diabetes. In the Epidemiology of Diabetes Complications (EDC) Study, a prospective study of patients with type 1 diabetes, the overall prevalence of DPN at base-line was 37%. In the San Luis Valley Diabetes Study, a population based study of type 2 diabetic patients, there was an overall prevalence of 28%. The natural history of DPN is difficult to ascertain from such studies. Pirart’s study revealed strong univariate associations between neuropathy and the duration of diabetes as well as degree of hyperglycaemia. The EURODIAB IDDM study has employed mul- tivariate modeling to identify associations of neuropathy with age, duration of diabetes, HbA1 C and severe ketoacidosis. Furthermore, the prevalence of neu- ropathy was related to elevated diastolic blood pressure and triglycerides and decreased HDL-cholesterol. In the DCCT impairment of nerve conduction was found to be associated with age, diabetes duration HbA1 C , male gender and C- peptide deficiency. In the EDC Study DPN at base-line was related to age, dia- betes duration, HbA1, HDL-cholesterol, hypertension and cigarette smoking. The SLVDS found that DPN was related to age, diabetes duration, HbA1 C and insulin use. Several studies have also observed associations of DPN with retinopathy and nephropathy. In a case-control study, DPN was associated with lifetime cigarette smoking in type 1 but not type 2 diabetes. Table 10.2. Risk factors and their biological basis for DPN.Table 10.2. Risk factors and their biological basis for DPN. SECTION II • DIABETIC NEUROPATHIES 94 CONCLUSION Despite the seeming inconsistencies of findings in epidemiological studies of diabetic neuropathy, one thing is clear: DPN is very common. At least one man- ifestation of DPN is present in well over 20% of individuals with diabetes. However, the prevalence of painful neuropathy appears to be appreciably lower. Regarding natural history, some degree of abnormality occurs virtually from the diagnosis of Type 1 and particularly type 2 diabetes. Once DPN is detectable, especially with an abnormality in vibration sense, there is a tendency toward rapid progression. DPN is associated with the degree of hyperglycaemia, diabetes duration, height, conventional cardiovascular risk factors (including lipid and blood pressure indices), and other complications of diabetes. Other risk factors such as alcohol consumption and cigarette smoking have been less consistent in their associations with DPN. The definitive risk factors that have been identified have biological plausibility for their involvement in the pathogenesis of DPN (Table 10.2). Duration and degree of hyperglycaemia indicate the extent of over- all exposure to hyperglycaemia. Height, as a proxy for nerve length entertains the hypothesis that the longer nerves are more susceptible to the metabolic and vas- cular consequence of diabetes. Blood pressure and lipid indices would appear to strengthen the assertion that vascular abnormalities contribute to the develop- ment and progression of DPN and the relation with other complications sug- gests they have common (vascular) pathogenetic pathways. Thus epidemiologic studies provide important clues for future mechanistic research and also provide support for mechanisms already defined in the patho- genesis of DPN. Future epidemiological studies should pay particular attention to the endpoints and methodology utilized. Risk factors and their biological basis for DPN Risk factor Biological basis Age, duration of diabetes, HbA1 C Measures of total exposure to primary inducer severe ketoacidosis of multiple pathogenetic pathways. Height Proxy for longer nerves being more susceptible to metabolic/vascular mechanisms Dyslipidaemia Inducer of vascular dysfunction Hypertension Inducer of vascular dysfunction Nephropathy and Retinopathy Common vascular basis for all complications *Alcohol consumption May aggravate neuropathy *Cigarette smoking May aggravate vascular dysfunction * Less consistent associations with DPN. CHAPTER 10 • EPIDEMIOLOGY AND NATURAL HISTORY OF DPN 95 CURRENT ISSUES • Epidemiological studies of diabetic peripheral neuropathy (DPN) have produced widely varying figures on incidence and prevalence. • The prevalence of painful symptoms varies from 3–30%. • The 10 g monofilament is neither sensitive enough to detect early loss of sensation nor to detect a small change in progressive loss of sensation – and should only be used to detect those at risk of foot ulceration. • The EURODIAB IDDM study has identified associations between neuropathy and age, duration of diabetes, HbA1 C , diastolic blood pressure, triglycerides and decreased HDL-cholesterol. FURTHER READING EURODIAB IDDM Study Group. Prevalence of diabetic peripheral neuropathy and its relation to glycaemic control and potential risk factors: the EURODIAB IDDM com- pliction study. Diabetologia 1996; 39: 1377–1386. The EURODIAB Prospective Complications Study (PCS) Group. Cardiovascular risk factors predict diabetic peripheral neuropathy in type 1 subjects in Europe. Diabetologia 1999; 42: A50–181. Witte DR, Tesfaye S, Chaturvedi N, Eaton SE, Kempler P, Fuller JH, EURODIAB Prospective Complications Study Group. Risk factors for cardiac autonomic neurpathy in type 1 diabetes mellitus. Diabetologia 2005; 48: 164–171. CHAPTER 11 DETECTION/SCREENING/ASSESSMENT Rayaz A. Malik MB.ChB, PhD, MRCP 97 SYMPTOMS Patients often have difficulty in describing the symptoms of neuropathy. Therefore when physicians record symptoms in clinical practice they must avoid ‘interpreting’ or ‘translating’ what patients report, rather they should record the patient’s description verbatim. For simple screening, a number of questionnaires are available and include the Neuropathy Symptom Score (NSS), Michigan Neuropathy Screening Instrument (MNSI), and more recently the NSS +4. Where a response to treatment is being assessed, visual analogue or verbal descriptive scales are used, but few have been validated for diabetic neuropathy. Signs Composite scores which quantify the degree of neurological deficit in the lower limbs were pioneered by Dyck and co-workers who first described the Neuropathy Disability Score (NDS) and later the Neuropathy Impairment Score (NIS). A modified NDS has been used in several large studies (Table 11.1) and has been shown to predict foot ulceration in a large prospective community study. A clinical scoring system which documents and monitors neuropathy in the clinic has been validated by the Toronto group. CLINICAL SCREENING Whilst the simple hand held screening devices are less sensitive than the more sophisticated QST devices, they are cheap, portable and easy to use. Currently the most widely and sometimes perhaps inappropriately used device is the Semmes-Weinstein monofilament. The lack of perception of pressure to gentle pressure applied to the handle sufficient to buckle the nylon filament defines an abnormal response. Although filaments of many different sizes are available, the 10g or 5.07 (10 x log of the force generated to deform the filament) monofilament should be used. In identifying feet at risk of ulceration it has a sensitivity of 86–100%. The commonest algorithm recommends four sites per foot (hallux and metatarsal heads 1, 3, and 5). Although a recent study suggests that there is little advantage gained from multiple site assessment. Additionally a recent study has shown that fila- ments manufactured by a number of companies actually buckle at 8g rather than the validated 10g monofilament. The graduated Rydel-seiffer tuning fork is widely used in Europe, particularly in Germany. It allows the assessor to define a threshold on a 0–8 scale and has been shown to correlate well Vascular Complications of Diabetes: Current Issues in Pathogenesis and Treatment, Second Edition Edited by Richard Donnelly, Edward Horton Copyright © 2005 by Blackwell Publishing Ltd Table 11.1 Neuropathy Disability Score. SECTION II • DIABETIC NEUROPATHIES 98 with other QST measures. The recently reported Neuropen combines the assessment of pain using a neurotip with pressure using a 10g monofilament. QUANTITATIVE SENSORY TESTING (QST) QST may be defined as any procedure requiring a power source where the inten- sity and characteristics of the stimuli are well controlled and where the detection threshold is determined in parametric units that can be compared to established normal values. QST measures the three major modalities of vibration, thermal and pain thresholds and has been shown to detect sub-clinical neuropathy, track its progression and predict those patients at risk of foot ulceration. Despite the strengths and weaknesses of QST (Table 11.2) it has been used as a primary effi- cacy measure in a number of completed and on-going clinical trials of DPN. Vibration perception threshold (VPT) VPT reflects the activation of mechanoreceptors (i.e. Pacinian and Meissner corpuscles), conduction in peripheral large diameter myelinated axons, and transmission through the dorsal column spinal pathways. Vibratory thresholds Risk factors and their biological basis for DPN Right Left Vibration perception threshold 128 Hz tuning fork; apex of big toe: normal = can distinguish vibrating/ not vibrating Temperature perception on dorsum of the foot Use tuning fork with beaker of Normal = 0 Abnormal = 1 ice/warm water Pin-prick apply pin proximal to big toe nail just enough to deform the skin; trial pair = sharp, blunt ; normal = can distinguish sharp / not sharp Achilles reflex Present = 0 Present with reinforcement = 1 Absent = 2 NDS Total out of 10 CHAPTER 11 • DETECTION/SCREENING/ASSESSMENT have been shown to detect not only sub-clinical neuropathy in children and adolescents with type 1 diabetes but also provide a strong indication of risk for future ulceration. In a 4-year prospective study patients with base-line VPT >25 volts with the biothesiometer were seven times more likely to develop foot ulcers. Recently in 187 type 2 diabetic patients multivariate logistic regression has shown that an elevated VPT score was the strongest predictor of foot ulcer- ation (relative risk of 25.4). In a large prospective study of 1,035 type 1 and type 2 diabetic patients, each one unit increase in vibration threshold (voltage scale) at base-line increased the hazard of foot ulceration by 5.6% over one year. Thermal thresholds Separate cold and warm thermoreceptors conduct thermal energy in thinly myelinated Aδ or unmyelinated C fibres, respectively. Additionally ‘pain’ can be driven principally by high intensity stimulation of warm thermoreceptors. Thermal thresholds are abnormal in patients with sub-clinical neuropathy, increase with progression of neuropathy and also predict foot ulceration. Generally, there is a high correlation between elevated thermal and vibration thresholds, but when these measures are dissociated it suggests a predomi- nant small or large fibre neuropathy. Electrophysiology Multiple consensus panels have recommended electrophysiology in the eval- uation of diabetic peripheral neuropathy (DPN), and in its use as a primary measure of therapeutic efficacy in multicentre clinical intervention trials. The 99 Table 11.2 Strengths and limitations of QST. Strengths and limitations of QST Strengths • Accurate control of stimulus characteristics • Ability to assess multiple modalities • Use of well established psychophysical procedures to enhance sensitivity • Measures function over a wide range of intensity and hence neuropathic severity • Measures sensation at multiple anatomical sites • Data from large, age-matched normal groups available for comparison Limitations • Semi-objective measure affected by the subject’s motivation and cooperation • Affected by age, gender, body mass, history of smoking and alcohol consumption • Expectancy and subject bias • Affected by any change along the entire neuroaxis from nerve to cortex. • Chronic liver/renal disease [...]... Diabetes Care 1998; 21: 822–827 35 36 Study 3 (III-172) D’Hemercourt et al Wounds 1998; 10: 69– 75 36 44 Study 4 (III- 250 ) Smiell et al Wound Rep Regen 1999; 7: 3 35 346 32 36 Study 5 (IIIb-134) Embil et al Wound Rep Regen 2000; 8: 162–168 50 57 Table 12.2 Analysis of % complete wound closure at 20 weeks in PDGF studies CHAPTER 12 • FOOT ULCERATION AND CHARCOT ARTHROPATHY Fig 12.2 X-ray showing a mid foot Charcot... than the 4-week median (53 %) had a 12-week healing rate of 58 %, whereas those with a reduction in ulcer area less than the 4-week median had a healing rate of only 9% Thus the percent change in foot ulcer area after four weeks of observation appears to be a robust predictor of healing at 12 weeks Revascularization When associated with significant ischemia, diabetic foot ulcers require arterial revascularization... distal segment of the axon, including possible therapeutic benefits, may be poorly represented in F-wave measures The distribution of conduction velocities allows an assessment of the activity in small diameter axons The recent fusion of a collision technique with an analysis of the distribution of velocities has shown that it is highly sensitive for detecting sub-clinical neuropathy as 58 % of patients... difficult to manage as standard off-loading is made difficult due to the deformity (Fig 12.3) A study of randomly selected neuropathic patients reported radiological evidence of CN in 16% of patients, suggesting a key role of neuropathy in the pathogenesis of this condition Other factors which may predispose to CN include an alteration in bone architecture and strength In a study of 16 patients with acute... subcommittee of the American Academy of Neurology Quantitative sensory testing Neurology 2003; 602: 898–906 Vascular Complications of Diabetes: Current Issues in Pathogenesis and Treatment, Second Edition Edited by Richard Donnelly, Edward Horton Copyright © 20 05 by Blackwell Publishing Ltd Rayaz A Malik MB.ChB, PhD, MRCP CHAPTER 12 FOOT ULCERATION AND CHARCOT ARTHROPATHY The late sequelae of diabetic... Edmonds ME, Boulton AJ Bisphosphonates in the treatment of Charcot neuroarthropathy: a double-blind randomised controlled trial Diabetologia 2001; 44: 2032–2037 Koitka A, Abraham P, Bouhanick B, Sigaudo-Roussel D, Demiot C, Saumet JL Impaired pressure-induced vasodilation at the foot in young adults with type 1 diabetes Diabetes 2004; 53 : 721–7 25 Lavery LA, Higgins KR, Lanctot DR, Constantinides GP,... amplitude of either the sensory response (SNAP) or the compound muscle action potential (CMAP) reflects the number of responding Principle factors governing electrophysiological alterations in diabetic neuropathy • Integrity and degree of myelination of the largest diameter fibres • Mean cross-sectional diameter of the responding axons • Representative internodal distance in the segment under study • Micro-environment... and non-Charcot foot and a significantly lower foot, femoral neck and lumbar bone mineral density (BMD) in comparison with a control group However, it is important to remember that osteoporosis is more prevalent in Analysis of % complete wound closure at 20 weeks in PDGF studies Placebo 30 μg/g 100 μg/g Study 1 (II-118) Steed et al J Vasc Surg 19 95; 21: 71–78 25 48 Study 2 (III-382) Wieman et al Diabetes. .. decrease of approximately 150 myelinated fibres/mm2, while a loss of 200 fibres/mm2 is associated with an approximate 1.0 mV reduction in the mean amplitude of the CMAP from the ulnar, peroneal and tibial nerves The total area of the SNAP and CMAP may reflect the contribution of slower conducting fibres, but it is severely limited by variability F-waves reflect the antidromic conduction of the compound neural... Charcot joints, but the evidence of benefit is limited CHAPTER 12 • FOOT ULCERATION AND CHARCOT ARTHROPATHY FURTHER READING Abidia A, Laden G, Kuhan G, Johnson BF, Wilkinson AR, Renwick PM, Masson EA, McCollum PT The role of hyperbaric oxygen therapy in ischaemic diabetic lower extremity ulcers: a double-blind randomised-controlled trial Eur J Vasc Endovasc Surg 2003; 25: 51 3 51 8 Boulton AJ The diabetic . In the Diabetes Control and Complications Trial (DCCT) ‘clinically detectable’ neu- ropathy was found in 39% of the participants who had type 1 diabetes. In the Epidemiology of Diabetes Complications. VEGF expres- sion in controls. Intramuscular gene transfer of plasmid DNA encoding VEGF- 1 or VEGF-2 in the STZ-diabetic rat and alloxan diabetic rabbit results in restora- tion of nerve vascularity,. in Analysis of % complete wound closure at 20 weeks in PDGF studies Placebo 30 μg/g 100 μg/g Study 1 (II-118) 25 48 Steed et al. J Vasc Surg 19 95; 21: 71–78. Study 2 (III-382) 35 36 50 Wieman et al. Diabetes