Association of transketolase polymorphisms with diabetic polyneuropathy in the general population: The KORA F4 study.

AIMS: We recently reported that genetic variability in the TKT gene encoding transketolase, a key enzyme in the pentose phosphate pathway, is associated with measures of diabetic sensorimotor polyneuropathy (DSPN) in recent-onset diabetes. Here, we aimed to substantiate these findings in a population-based KORA F4 study. MATERIALS AND METHODS: In this cross-sectional study, we assessed seven single nucleotide polymorphisms (SNPs) in the transketolase gene in 952 participants from the KORA F4 study with normal glucose tolerance (NGT; n = 394), prediabetes (n = 411), and type 2 diabetes (n = 147). DSPN was defined by the examination part of the Michigan Neuropathy Screening Instrument (MNSI) using the original MNSI > 2 cut-off and two alternative versions extended by touch/pressure perception (TPP) (MNSI > 3) and by TPP plus cold perception (MNSI > 4). RESULTS: After adjustment for sex, age, BMI, and HbA1c, in type 2 diabetes participants, four out of seven transketolase SNPs were associated with DSPN for all three MNSI versions (all p ≤ 0.004). The odds ratios of these associations increased with extending the MNSI score, for example, OR (95% CI) for SNP rs62255988 with MNSI > 2: 1.99 (1.16-3.41), MNSI > 3: 2.27 (1.26-4.09), and MNSI > 4: 4.78 (2.22-10.26); SNP rs9284890 with MNSI > 2: 2.43 (1.42-4.16), MNSI > 3: 3.46 (1.82-6.59), and MNSI > 4: 4.75 (2.15-10.51). In contrast, no associations were found between transketolase SNPs and the three MNSI versions in the NGT and prediabetes groups. CONCLUSIONS: The link of genetic variation in transketolase enzyme to diabetic polyneuropathy corroborated at the population level strengthens the concept suggesting an important role of pathways metabolising glycolytic intermediates in the evolution of diabetic polyneuropathy.

Associations between multiple neurological biomarkers and distal sensorimotor polyneuropathy: KORA F4/FF4 study.

AIMS: The aim of this study was to assess associations between neurological biomarkers and distal sensorimotor polyneuropathy (DSPN). MATERIALS AND METHODS: Cross-sectional analyses were based on 1032 participants aged 61-82 years from the population-based KORA F4 survey, 177 of whom had DSPN at baseline. The prevalence of type 2 diabetes was 20%. Prospective analyses used data from 505 participants without DSPN at baseline, of whom 125 had developed DSPN until the KORA FF4 survey. DSPN was defined based on the examination part of the Michigan Neuropathy Screening Instrument. Serum levels of neurological biomarkers were measured using proximity extension assay technology. Associations between 88 biomarkers and prevalent or incident DSPN were estimated using Poisson regression with robust error variance and are expressed as risk ratios (RR) and 95% CI per 1-SD increase. Results were adjusted for multiple confounders and multiple testing using the Benjamini-Hochberg procedure. RESULTS: Higher serum levels of CTSC (cathepsin C; RR [95% CI] 1.23 (1.08; 1.39), pB-H = 0.044) and PDGFRα (platelet-derived growth factor receptor A; RR [95% CI] 1.21 (1.08; 1.35), pB-H = 0.044) were associated with prevalent DSPN in the total study sample. CDH3, JAM-B, LAYN, RGMA and SCARA5 were positively associated with DSPN in the diabetes subgroup, whereas GCP5 was positively associated with DSPN in people without diabetes (all pB-H for interaction <0.05). None of the biomarkers showed an association with incident DSPN (all pB-H>0.05). CONCLUSIONS: This study identified multiple novel associations between neurological biomarkers and prevalent DSPN, which may be attributable to functions of these proteins in neuroinflammation, neural development and myelination.

Association of dietary patterns with diabetes-related comorbidities varies among diabetes endotypes.

BACKGROUND AND AIMS: Differences of dietary pattern adherence across the novel diabetes endotypes are unknown. This study assessed adherence to pre-specified dietary patterns and their associations with cardiovascular risk factors, kidney function, and neuropathy among diabetes endotypes. METHODS AND RESULTS: The cross-sectional analysis included 765 individuals with recent-onset (67 %) and prevalent diabetes (33 %) from the German Diabetes Study (GDS) allocated into severe autoimmune diabetes (SAID, 35 %), severe insulin-deficient diabetes (SIDD, 3 %), severe insulin-resistant diabetes (SIRD, 5 %), mild obesity-related diabetes (MOD, 28 %), and mild age-related diabetes (MARD, 29 %). Adherence to a Mediterranean diet score (MDS), Dietary Approaches to Stop Hypertension (DASH) score, overall plant-based diet (PDI), healthful (hPDI) and unhealthful plant-based diet index (uPDI) was derived from a food frequency questionnaire and associated with cardiovascular risk factors, kidney function, and neuropathy using multivariable linear regression analysis. Differences in dietary pattern adherence between endotypes were assessed using generalized mixed models. People with MARD showed the highest, those with SIDD and MOD the lowest adherence to the hPDI. Adherence to the MDS, DASH, overall PDI, and hPDI was inversely associated with high-sensitivity C-reactive protein (hsCRP) among people with MARD (ß (95%CI): -9.18 % (-15.61; -2.26); -13.61 % (-24.17; -1.58); -19.15 % (-34.28; -0.53); -16.10 % (-28.81; -1.12), respectively). Adherence to the PDIs was associated with LDL cholesterol among people with SAID, SIRD, and MOD. CONCLUSIONS: Minor differences in dietary pattern adherence (in particular for hPDI) and associations with markers of diabetes-related complications (e.g. hsCRP) were observed between endotypes. So far, evidence is insufficient to derive endotype-specific dietary recommendations. TRIAL REGISTRATION: Clinicaltrials.gov: NCT01055093.

Analysis of type 2 diabetes heterogeneity with a tree-like representation: insights from the prospective German Diabetes Study and the LURIC cohort.

BACKGROUND: Heterogeneity in type 2 diabetes can be represented by a tree-like graph structure by use of reversed graph-embedded dimensionality reduction. We aimed to examine whether this approach can be used to stratify key pathophysiological components and diabetes-related complications during longitudinal follow-up of individuals with recent-onset type 2 diabetes. METHODS: For this cohort analysis, 927 participants aged 18-69 years from the German Diabetes Study (GDS) with recent-onset type 2 diabetes were mapped onto a previously developed two-dimensional tree based on nine simple clinical and laboratory variables, residualised for age and sex. Insulin sensitivity was assessed by a hyperinsulinaemic-euglycaemic clamp, insulin secretion was assessed by intravenous glucose tolerance test, hepatic lipid content was assessed by 1 H magnetic resonance spectroscopy, serum interleukin (IL)-6 and IL-18 were assessed by ELISA, and peripheral and autonomic neuropathy were assessed by functional and clinical measures. Participants were followed up for up to 16 years. We also investigated heart failure and all-cause mortality in 794 individuals with type 2 diabetes undergoing invasive coronary diagnostics from the Ludwigshafen Risk and Cardiovascular Health (LURIC) cohort. FINDINGS: There were gradients of clamp-measured insulin sensitivity (both dimensions: p<0·0001) and insulin secretion (pdim1<0·0001, pdim2=0·00097) across the tree. Individuals in the region with the lowest insulin sensitivity had the highest hepatic lipid content (n=205, pdim1<0·0001, pdim2=0·037), pro-inflammatory biomarkers (IL-6: n=348, pdim1<0·0001, pdim2=0·013; IL-18: n=350, pdim1<0·0001, pdim2=0·38), and elevated cardiovascular risk (nevents=143, pdim1=0·14, pdim2<0·00081), whereas individuals positioned in the branch with the lowest insulin secretion were more prone to require insulin therapy (nevents=85, pdim1=0·032, pdim2=0·12) and had the highest risk of diabetic sensorimotor polyneuropathy (nevents=184, pdim1=0·012, pdim2=0·044) and cardiac autonomic neuropathy (nevents=118, pdim1=0·0094, pdim2=0·06). In the LURIC cohort, all-cause mortality was highest in the tree branch showing insulin resistance (nevents=488, pdim1=0·12, pdim2=0·0032). Significant gradients differentiated individuals having heart failure with preserved ejection fraction from those who had heart failure with reduced ejection fraction. INTERPRETATION: These data define the pathophysiological underpinnings of the tree structure, which has the potential to stratify diabetes-related complications on the basis of routinely available variables and thereby expand the toolbox of precision diabetes diagnosis. FUNDING: German Diabetes Center, German Federal Ministry of Health, Ministry of Culture and Science of the state of North Rhine-Westphalia, German Federal Ministry of Education and Research, German Diabetes Association, German Center for Diabetes Research, European Community, German Research Foundation, and Schmutzler Stiftung.

Intramyocellular Triglyceride Content During the Early Course of Type 1 and Type 2 Diabetes.

Intramyocellular lipid content (IMCL) is elevated in insulin-resistant humans, but it changes over time, and relationships with comorbidities remain unclear. We examined IMCL during the initial course of diabetes and its associations with complications. Participants of the German Diabetes Study (GDS) with recent-onset type 1 (n = 132) or type 2 diabetes (n = 139) and glucose-tolerant control subjects (n = 128) underwent 1H-MRS to measure IMCL and muscle volume, whole-body insulin sensitivity (hyperinsulinemic-euglycemic clamps; M-value), and cycling spiroergometry (VO2max). Subgroups underwent the same measurements after 5 years. At baseline, IMCL was ∼30% higher in type 2 diabetes than in other groups independently of age, sex, BMI, and muscle volume. In type 2 diabetes, the M-value was ∼36% and ∼62% lower compared with type 1 diabetes and control subjects, respectively. After 5 years, the M-value decreased by ∼29% in type 1 and ∼13% in type 2 diabetes, whereas IMCL remained unchanged. The correlation between IMCL and M-value in type 2 diabetes at baseline was modulated by VO2max. IMCL also associated with microalbuminuria, the Framingham risk score for cardiovascular disease, and cardiac autonomic neuropathy. Changes in IMCL within 5 years after diagnosis do not mirror the progression of insulin resistance in type 2 diabetes but associate with early diabetes-related complications. ARTICLE HIGHLIGHTS: Intramyocellular lipid content (IMCL) can be elevated in insulin-resistant humans, but its dynamics and association with comorbidities remain unclear. Independently of age, sex, body mass, and skeletal muscle volume, IMCL is higher in recent-onset type 2, but not type 1 diabetes, and remains unchanged within 5 years, despite worsening insulin resistance. A degree of physical fitness modulates the association between IMCL and insulin sensitivity in type 2 diabetes. Whereas higher IMCL associates with lower insulin sensitivity in people with lower physical fitness, there is no association between IMCL and insulin sensitivity in those with higher degree of physical fitness. IMCL associates with progression of microalbuminuria, cardiovascular disease risk, and cardiac autonomic neuropathy.

Association between diabetes and thiamine status - A systematic review and meta-analysis.

BACKGROUND: Thiamine (vitamin B1) is an essential cofactor in glucose metabolism, but it remains unclear whether thiamine status is lower in individuals with diabetes compared to individuals with normal glucose metabolism. AIMS: We conducted a systematic review and meta-analysis to study whether the circulating concentrations of various thiamine analytes differ between people with and those without diabetes. METHODS: PubMed and the Cochrane Central Register of Controlled Trials were searched according to the study protocol. The standardized mean difference (SMD) and 95 % confidence intervals (CI) of thiamine markers between individuals with and without diabetes were used as effect size (random effects model). Subgroup analysis considered albuminuria as an additional variable. RESULTS: Out of the 459 articles identified, 24 full-texts were eligible for the study, 20 of which qualified for the data analysis and four were evaluated for coherence. Compared to controls, individuals with diabetes showed lower concentrations of thiamine (pooled estimate SMD [95 % CI]: -0.97 [-1.89, -0.06]), thiamine monophosphate (-1.16 [-1.82, -0.50]), and total thiamine compounds (-1.01 [-1.48, -0.54]). Thiamine diphosphate (-0.72 [-1.54, 0.11] and erythrocyte transketolase activity (-0.42 [-0.90, 0.05]) tended to be lower in persons with diabetes than in controls without reaching statistical significance. Subgroup analysis showed that individuals with diabetes and albuminuria had lower thiamine levels than the controls (-2.68 [-5.34, -0.02]). CONCLUSIONS: Diabetes is associated with lower levels of various thiamine markers, suggesting that individuals with diabetes may have higher thiamine requirements than those without diabetes, but well-designed studies are required to confirm these findings.

Serum neurofilament light chain: a novel biomarker for early diabetic sensorimotor polyneuropathy.

AIMS/HYPOTHESIS: No established blood-based biomarker exists to monitor diabetic sensorimotor polyneuropathy (DSPN) and evaluate treatment response. The neurofilament light chain (NFL), a blood biomarker of neuroaxonal damage in several neurodegenerative diseases, represents a potential biomarker for DSPN. We hypothesised that higher serum NFL levels are associated with prevalent DSPN and nerve dysfunction in individuals recently diagnosed with diabetes. METHODS: This cross-sectional study included 423 adults with type 1 and type 2 diabetes and known diabetes duration of less than 1 year from the prospective observational German Diabetes Study cohort. NFL was measured in serum samples of fasting participants in a multiplex approach using proximity extension assay technology. DSPN was assessed by neurological examination, nerve conduction studies and quantitative sensory testing. Associations of serum NFL with DSPN (defined according to the Toronto Consensus criteria) were estimated using Poisson regression, while multivariable linear and quantile regression models were used to assess associations with nerve function measures. In exploratory analyses, other biomarkers in the multiplex panel were also analysed similarly to NFL. RESULTS: DSPN was found in 16% of the study sample. Serum NFL levels increased with age. After adjustment for age, sex, waist circumference, height, HbA1c, known diabetes duration, diabetes type, cholesterol, eGFR, hypertension, CVD, use of lipid-lowering drugs and use of non-steroidal anti-inflammatory drugs, higher serum NFL levels were associated with DSPN (RR [95% CI] per 1-normalised protein expression increase, 1.92 [1.50, 2.45], p<0.0001), slower motor (all p<0.0001) and sensory (all p≤0.03) nerve conduction velocities, lower sural sensory nerve action potential (p=0.0004) and higher thermal detection threshold to warm stimuli (p=0.023 and p=0.004 for hand and foot, respectively). There was no evidence for associations between other neurological biomarkers and DSPN or nerve function measures. CONCLUSIONS/INTERPRETATION: Our findings in individuals recently diagnosed with diabetes provide new evidence associating higher serum NFL levels with DSPN and peripheral nerve dysfunction. The present study advocates NFL as a potential biomarker for DSPN.

Dynamic Long-Range Interactions Influence Substrate Binding and Catalysis by Human Histidine Triad Nucleotide-Binding Proteins (HINTs), Key Regulators of Multiple Cellular Processes and Activators of Antiviral ProTides.

Human histidine triad nucleotide-binding (hHINT) proteins catalyze nucleotide phosphoramidase and acyl-phosphatase reactions that are essential for the activation of antiviral proTides, such as Sofosbuvir and Remdesivir. hHINT1 and hHINT2 are highly homologous but exhibit disparate roles as regulators of opioid tolerance (hHINT1) and mitochondrial activity (hHINT2). NMR studies of hHINT1 reveal a pair of dynamic surface residues (Q62, E100), which gate a conserved water channel leading to the active site 13 Å away. hHINT2 crystal structures identify analogous residues (R99, D137) and water channel. hHINT1 Q62 variants significantly alter the steady-state kcat and Km for turnover of the fluorescent substrate (TpAd), while stopped-flow kinetics indicate that KD also changes. hHINT2, like hHINT1, exhibits a burst phase of adenylation, monitored by fluorescent tryptamine release, prior to rate-limiting hydrolysis and nucleotide release. hHINT2 exhibits a much smaller burst-phase amplitude than hHINT1, which is further diminished in hHINT2 R99Q. Kinetic simulations suggest that amplitude variations can be accounted for by a variable fluorescent yield of the E·S complex from changes in the environment of bound TpAd. Isothermal titration calorimetry measurements of inhibitor binding show that these hHINT variants also alter the thermodynamic binding profile. We propose that these altered surface residues engender long-range dynamic changes that affect the orientation of bound ligands, altering the thermodynamic and kinetic characteristics of hHINT active site function. Thus, studies of the cellular roles and proTide activation potential by hHINTs should consider the importance of long-range interactions and possible protein binding surfaces far from the active site.

Screening and diagnosis of diabetic polyneuropathy in clinical practice: A survey among German physicians (PROTECT Study Survey).

AIMS: We sought to obtain detailed information on the procedures and appraisal of screening for and diagnosing diabetic sensorimotor polyneuropathy (DSPN) in clinical practice. METHODS: This cross-sectional survey included 574 physicians from 13 federal states across Germany who responded to a tripartite questionnaire. RESULTS: The vast majority of the respondents reported to screen for DSPN at least once a year (87 %), while 65 % reported to examine the feet of DSPN patients at least twice a year. However, only 28 % and 20 % of the respondents used questionnaires and scores to assess the severity of neuropathic symptoms and signs, respectively. The rates of participants reporting that they do not use a standardized testing procedure were 58 % for pressure sensation, 62 % for pain sensation, and 54 % for thermal sensation. The rates of respondents reporting that they do not deploy a standardized assessment were 41 % for vibration sensation, 73 % for pressure sensation, 77 % for pain sensation, and 66 % for thermal sensation. Half of the physicians oriented themselves towards clinical guidelines when diagnosing DSPN. CONCLUSIONS: Despite relatively high screening rates, the willingness to implement both standardized testing procedures and assessment and to follow guidelines is low among physicians when screening for and clinically diagnosing DSPN.

High-intensity interval training for 12 weeks improves cardiovascular autonomic function but not somatosensory nerve function and structure in overweight men with type 2 diabetes.

AIMS/HYPOTHESIS: It remains unclear whether and which modality of exercise training as a component of lifestyle intervention may exert favourable effects on somatosensory and autonomic nerve tests in people with type 2 diabetes. METHODS: Cardiovascular autonomic and somatosensory nerve function as well as intraepidermal nerve fibre density (IENFD) were assessed in overweight men with type 2 diabetes (type 2 diabetes, n = 20) and male glucose-tolerant individuals (normal glucose tolerance [NGT], n = 23), comparable in age and BMI and serving as a control group, before and after a supervised high-intensity interval training (HIIT) intervention programme over 12 weeks. Study endpoints included clinical scores, nerve conduction studies, quantitative sensory testing, IENFD, heart rate variability, postural change in systolic blood pressure and spontaneous baroreflex sensitivity (BRS). RESULTS: After 12 weeks of HIIT, resting heart rate decreased in both groups ([mean ± SD] baseline/12 weeks: NGT: 65.1 ± 8.2/60.2 ± 9.0 beats per min; type 2 diabetes: 68.8 ± 10.1/63.4 ± 7.8 beats per min), while three BRS indices increased (sequence analysis BRS: 8.82 ± 4.89/14.6 ± 11.7 ms2/mmHg; positive sequences BRS: 7.19 ± 5.43/15.4 ± 15.9 ms2/mmHg; negative sequences BRS: 12.8 ± 5.4/14.6 ± 8.7 ms2/mmHg) and postural change in systolic blood pressure decreased (-13.9 ± 11.6/-9.35 ± 9.76 mmHg) in participants with type 2 diabetes, and two heart rate variability indices increased in the NGT group (standard deviation of R-R intervals: 36.1 ± 11.8/55.3 ± 41.3 ms; coefficient of R-R interval variation: 3.84 ± 1.21/5.17 ± 3.28) (all p<0.05). In contrast, BMI, clinical scores, nerve conduction studies, quantitative sensory testing, IENFD and the prevalence rates of diabetic sensorimotor polyneuropathy and cardiovascular autonomic neuropathy remained unchanged in both groups. In the entire cohort, correlations between the changes in two BRS indices and changes in [Formula: see text] over 12 weeks of HIIT (e.g. sequence analysis BRS: r = 0.528, p=0.017) were observed. CONCLUSIONS/INTERPRETATION: In male overweight individuals with type 2 diabetes, BRS, resting heart rate and orthostatic blood pressure regulation improved in the absence of weight loss after 12 weeks of supervised HIIT. Since no favourable effects on somatic nerve function and structure were observed, cardiovascular autonomic function appears to be more amenable to this short-term intervention, possibly due to improved cardiorespiratory fitness.

BOND study: a randomised double-blind, placebo-controlled trial over 12 months to assess the effects of benfotiamine on morphometric, neurophysiological and clinical measures in patients with type 2 diabetes with symptomatic polyneuropathy.

INTRODUCTION: Diabetic sensorimotor polyneuropathy (DSPN) affects approximately 30% of people with diabetes, while around half of cases are symptomatic. Currently, there are only few pathogenetically oriented pharmacotherapies for DSPN, one of which is benfotiamine, a prodrug of thiamine with a high bioavailability and favourable safety profile. While benfotiamine has shown positive effects in preclinical and short-term clinical studies, no long-term clinical trials are available to demonstrate disease-modifying effects on DSPN using a comprehensive set of disease-related endpoints. METHODS AND ANALYSIS: The benfotiamine on morphometric, neurophysiological and clinical measures in patients with type 2 diabetes trial is a randomised double-blind, placebo-controlled parallel group monocentric phase II clinical trial to assess the effects of treatment with benfotiamine compared with placebo in participants with type 2 diabetes and mild to moderate symptomatic DSPN. Sixty participants will be 1:1 randomised to treatment with benfotiamine 300 mg or placebo two times a day over 12 months. The primary endpoint will be the change in corneal nerve fibre length assessed by corneal confocal microscopy (CCM) after 12 months of benfotiamine treatment compared with placebo. Secondary endpoints will include other CCM measures, skin biopsy and function indices, variables from somatic and autonomic nerve function tests, clinical examination and questionnaires, general health, health-related quality of life, cost, safety and blood tests. ETHICS AND DISSEMINATION: The trial was approved by the competent authority and the local independent ethics committee. Trial results will be published in peer-reviewed journals, conference abstracts, and via online and print media. TRIAL REGISTRATION NUMBER: DRKS00014832.

Neuron-specific biomarkers predict hypo- and hyperalgesia in individuals with diabetic peripheral neuropathy.

AIMS/HYPOTHESIS: The individual risk of progression of diabetic peripheral neuropathy is difficult to predict for each individual. Mutations in proteins that are responsible for the process of myelination are known to cause neurodegeneration and display alteration in experimental models of diabetic neuropathy. In a prospective observational human pilot study, we investigated myelin-specific circulating mRNA targets, which have been identified in vitro, for their capacity in the diagnosis and prediction of diabetic neuropathy. The most promising candidate was tested against the recently established biomarker of neural damage, neurofilament light chain protein. METHODS: Schwann cells were cultured under high-glucose conditions and mRNAs of various myelin-specific genes were screened intra- and extracellularly. Ninety-two participants with type 2 diabetes and 30 control participants were enrolled and evaluated for peripheral neuropathy using neuropathy deficit scores, neuropathy symptom scores and nerve conduction studies as well as quantitative sensory testing at baseline and after 12/24 months of a follow-up period. Magnetic resonance neurography of the sciatic nerve was performed in 37 individuals. Neurofilament light chain protein and four myelin-specific mRNA transcripts derived from in vitro screenings were measured in the serum of all participants. The results were tested for associations with specific neuropathic deficits, fractional anisotropy and the progression of neuropathic deficits at baseline and after 12 and 24 months. RESULTS: In neuronal Schwann cells and human nerve sections, myelin protein zero was identified as the strongest candidate for a biomarker study. Circulating mRNA of myelin protein zero was decreased significantly in participants with diabetic neuropathy (p < 0.001), whereas neurofilament light chain protein showed increased levels in participants with diabetic neuropathy (p < 0.05). Both variables were linked to altered electrophysiology, fractional anisotropy and quantitative sensory testing. In a receiver-operating characteristic curve analysis myelin protein zero improved the diagnostic performance significantly in combination with a standard model (diabetes duration, age, BMI, HbA1c) from an AUC of 0.681 to 0.836 for the detection of diabetic peripheral neuropathy. A follow-up study revealed that increased neurofilament light chain was associated with the development of a hyperalgesic phenotype (p < 0.05), whereas decreased myelin protein zero predicted hypoalgesia (p < 0.001) and progressive loss of nerve function 24 months in advance (HR of 6.519). CONCLUSIONS/INTERPRETATION: This study introduces a dynamic and non-invasive assessment strategy for the underlying pathogenesis of diabetic peripheral neuropathy. The diagnosis of axonal degeneration, associated with hyperalgesia, and demyelination, linked to hypoalgesia, could benefit from the usage of neurofilament light chain protein and circulating mRNA of myelin protein zero as potential biomarkers.

Progression and regression of nerve fibre pathology and dysfunction early in diabetes over 5 years.

It has traditionally been suggested that the early development of diabetic sensorimotor polyneuropathy (DSPN) is characterized by predominant and progressive injury to small nerve fibres followed by large fibre impairment. We alternatively hypothesized that small and large fibre damage due to DSPN in type 1 and type 2 diabetes could develop in parallel and may not only be progressive but also reversible. Participants from the German Diabetes Study baseline cohort with recent-onset type 1/type 2 diabetes (n = 350/570) and age-matched glucose-tolerant control individuals (Control 1/Control 2: n = 114/190) were assessed using nerve conduction studies, thermal detection thresholds, vibration perception thresholds, neuropathy symptom scores, neuropathy disability scores and intraepidermal nerve fibre density (IENFD) in skin biopsies (type 1/type 2 diabetes: n = 102/226; Control 1/Control 2: n = 109/208). Subsets of participants with type 1/type 2 diabetes were followed for 5 years (n = 184/307; IENFD subset: n = 18/69). DSPN was defined by the Toronto Consensus criteria. At baseline, DSPN was present in 8.1% and 13.3% of the type 1 and type 2 diabetes groups, respectively. The most frequently abnormal tests in the lower limbs below or above the 2.5th and 97.5th centiles of the controls were the IENFD (13.7%) and individual nerve conduction studies (up to 9.4%) in type 1 diabetes participants and IENFD (21.8%), malleolar vibration perception thresholds (17.5%), and individual nerve conduction studies (up to 11.8%) in those with type 2 diabetes, whereas thermal detection threshold abnormalities did not differ between the control and diabetes groups. After 5 years, the highest progression rates from the normal to the abnormal range in type 2 diabetes participants were found for IENFD (18.8%) by -4.1 ± 2.8 fibres/mm, malleolar vibration perception threshold (18.6%) by 9.1 ± 20.2 µm and nerve conduction studies (15.0%) by 3.7 ± 1.5 points, while vice versa the highest regression rates were observed for neuropathy disability scores (11.2%) by -3.1 ± 1.3 points, sural nerve amplitudes (9.1%) by 4.7 ± 3.0 µV, IENFD (8.7%) by 1.4 ± 1.3 fibres/mm, and neuropathy symptom scores (8.2%) by -5.8 ± 1.6 points. In type 1 diabetes participants, no major progression was seen after 5 years, but subclinical DSPN regressed in 10.3%. These findings point to early parallel damage to both small and large nerve fibres in well-controlled recent-onset type 2 and, to a lesser extent, type 1 diabetes. After 5 years, peripheral nerve morphology and function and clinical measures progress to the abnormal range in type 2 diabetes, but initial nerve alterations are also reversible to a meaningful degree.

"Switching On" Enzyme Substrate Specificity Analysis with a Fluorescent Competitive Inhibitor.

Enzymatically driven change to the spectroscopic properties of a chemical substrate or product has been a linchpin in the development of continuous enzyme kinetics assays. These assays inherently necessitate substrates or products that naturally comply with the constraints of the spectroscopic technique being used, or they require structural changes to the molecules involved to make them observable. Here we demonstrate a new analytical kinetics approach with enzyme histidine triad nucleotide binding protein 1 (HINT1) that allows us to extract both useful kcat values and a rank-ordered list of substrate specificities without the need to track substrates or products directly. Instead, this is accomplished indirectly using a "switch on" competitive inhibitor that fluoresces maximally only when bound to the HINT1 enzyme active site. Kinetic information is extracted from the duration of the diminished fluorescence when the monitorable inhibitor-bound enzyme is challenged with saturating concentrations of a nonfluorescent substrate. We refer to the loss of fluorescence, while the substrate competes for the fluorescent probe in the active site, as the substrate's residence transit time (RTT). The ability to assess kcat values and substrate specificity by monitoring the RTTs for a set of substrates with a competitive "switch on" inhibitor should be broadly applicable to other enzymatic reactions in which the "switch on" inhibitor has sufficient binding affinity over the enzymatic product.

Association of cardiac autonomic dysfunction with higher levels of plasma lipid metabolites in recent-onset type 2 diabetes.

AIMS/HYPOTHESIS: Emerging evidence suggests that in addition to hyperglycaemia, dyslipidaemia could represent a contributing pathogenetic factor to diabetic neuropathy, while obesity and insulin resistance play a role in the development of diabetic cardiac autonomic neuropathy (CAN) characterised by reduced heart rate variability (HRV), particularly in type 2 diabetes. We hypothesised that distinct lipid metabolites are associated with diminished HRV in recent-onset type 2 diabetes rather than type 1 diabetes. METHODS: We analysed 127 plasma lipid metabolites (11 acylcarnitines, 39 NEFA, 12 sphingomyelins (SMs), 56 phosphatidylcholines and nine lysophosphatidylcholines) using MS in participants from the German Diabetes Study baseline cohort recently diagnosed with type 1 (n = 100) and type 2 diabetes (n = 206). Four time-domain HRV indices (number of normal-to-normal (NN) intervals >50 ms divided by the number of all NN intervals [pNN50]; root mean square of successive differences [RMSSD]; SD of NN intervals [SDNN]; and SD of differences between adjacent NN intervals) and three frequency-domain HRV indices (very-low-frequency [VLF], low-frequency [LF] and high-frequency [HF] power spectrum) were computed from NN intervals recorded during a 3 h hyperinsulinaemic-euglycaemic clamp at baseline and in subsets of participants with type 1 (n = 60) and type 2 diabetes (n = 95) after 5 years. RESULTS: In participants with type 2 diabetes, after Bonferroni correction and rigorous adjustment, SDNN was inversely associated with higher levels of diacyl-phosphatidylcholine (PCaa) C32:0, PCaa C34:1, acyl-alkyl-phosphatidylcholine (PCae) C36:0, SM C16:0 and SM C16:1. SD of differences between NN intervals was inversely associated with PCaa C32:0, PCaa C34:1, PCaa C34:2, PCae C36:0 and SM C16:1, and RMSSD with PCae C36:0. For VLF power, inverse associations were found with PCaa C30:0, PCaa C32:0, PCaa C32:1, PCaa C34:2 and SM C16:1, and for LF power inverse associations were found with PCaa C32:0 and SM C16:1 (r = -0.242 to r = -0.349; p ≤ 0.0005 for all correlations). In contrast, no associations of lipid metabolites with measures of cardiac autonomic function were noted in participants recently diagnosed with type 1 diabetes. After 5 years, HRV declined due to ageing rather than diabetes, whereby prediction analyses for lipid metabolites were hampered. CONCLUSIONS/INTERPRETATION: Higher plasma levels of specific lipid metabolites are closely linked to cardiac autonomic dysfunction in recent-onset type 2 diabetes but not type 1 diabetes, suggesting a role for perturbed lipid metabolism in the early development of CAN in type 2 diabetes. Graphical abstract.

Interaction between magnesium and methylglyoxal in diabetic polyneuropathy and neuronal models.

OBJECTIVE: The lack of effective treatments against diabetic sensorimotor polyneuropathy demands the search for new strategies to combat or prevent the condition. Because reduced magnesium and increased methylglyoxal levels have been implicated in the development of both type 2 diabetes and neuropathic pain, we aimed to assess the putative interplay of both molecules with diabetic sensorimotor polyneuropathy. METHODS: In a cross-sectional study, serum magnesium and plasma methylglyoxal levels were measured in recently diagnosed type 2 diabetes patients with (n = 51) and without (n = 184) diabetic sensorimotor polyneuropathy from the German Diabetes Study baseline cohort. Peripheral nerve function was assessed using nerve conduction velocity and quantitative sensory testing. Human neuroblastoma cells (SH-SY5Y) and mouse dorsal root ganglia cells were used to characterize the neurotoxic effect of methylglyoxal and/or neuroprotective effect of magnesium. RESULTS: Here, we demonstrate that serum magnesium concentration was reduced in recently diagnosed type 2 diabetes patients with diabetic sensorimotor polyneuropathy and inversely associated with plasma methylglyoxal concentration. Magnesium, methylglyoxal, and, importantly, their interaction were strongly interrelated with methylglyoxal-dependent nerve dysfunction and were predictive of changes in nerve function. Magnesium supplementation prevented methylglyoxal neurotoxicity in differentiated SH-SY5Y neuron-like cells due to reduction of intracellular methylglyoxal formation, while supplementation with the divalent cations zinc and manganese had no effect on methylglyoxal neurotoxicity. Furthermore, the downregulation of mitochondrial activity in mouse dorsal root ganglia cells and consequently the enrichment of triosephosphates, the primary source of methylglyoxal, resulted in neurite degeneration, which was completely prevented through magnesium supplementation. CONCLUSIONS: These multifaceted findings reveal a novel putative pathophysiological pathway of hypomagnesemia-induced carbonyl stress leading to neuronal damage and merit further investigations not only for diabetic sensorimotor polyneuropathy but also other neurodegenerative diseases associated with magnesium deficiency and impaired energy metabolism.

Increased spontaneous CCL2 (MCP-1) and CCL5 (RANTES) secretion in vitro in LADA compared to type 1 diabetes and type 2 diabetes: Action LADA 14.

AIMS: Immune-mediated type 1 diabetes (T1D) in adulthood and latent autoimmune diabetes in adults (LADA) share similar pathological mechanisms but differ clinically in disease progression. The aim of this study was to acquire insights into spontaneous and stimulated chemokine secretion of immune cells in different diabetes types. MATERIALS AND METHODS: We investigated in vitro spontaneous, mitogen (PI) and antigen (HSP60, p277, pGAD, pIA2) stimulated chemokine secretion of leucocytes from patients with T1D (n = 32), LADA (n = 22), type 2 diabetes (T2D; n = 49), and glucose-tolerant individuals (n = 13). Chemokine concentration in supernatants was measured for CCL2 (MCP-1), CXCL10 (IP10) and CCL5 (RANTES) using a multiplex bead array assay. RESULTS: Spontaneous secretion of CCL2 and CCL5 were higher in LADA compared to T1D and T2D (all p < 0.05) while CXCL10 was similar in the groups. Mitogen-stimulated secretion of CCL2 in LADA was lower compared to T1D and T2D (all p < 0.05) while CXCL10 and CCL5 were similar in all groups. Upon stimulation with pIA2 the secretion of CCL2 in LADA was lower compared to T2D (p < 0.05). Spontaneous CXCL10 secretion in LADA was positively associated with body mass index (r2  = 0.35; p = 0.0035) and C-peptide (r2  = 0.30; p = 0.009). CONCLUSIONS: Chemokine secretion is altered between different diabetes types. Increased spontaneous secretion of CCL2 and CCL5 and decreased secretion of CCL2, upon stimulation with PI and pIA2, in LADA compared to T1D and T2D could reflect altered immune responsiveness in LADA patients in association with their slower clinical progression compared to insulin dependence.

Polyneuropathy is inadequately treated despite increasing symptom intensity in individuals with and without diabetes (PROTECT follow-up study).

AIMS/INTRODUCTION: Despite its major clinical impact, distal symmetric polyneuropathy remains frequently undiagnosed and undertreated in clinical practice. We previously reported in the PROTECT Study that 70% of type 2 diabetes patients with distal symmetric polyneuropathy were unaware of having the latter condition. MATERIALS AND METHODS: In the present follow up after 2.5 ± 0.7 years, 122 and 85 participants with and without type 2 diabetes, respectively, completed questionnaires to obtain information about the further course of disease and its management. RESULTS: At follow up, 49 and 48% of the respondents with type 2 diabetes and without diabetes, respectively, reported that the intensity of paresthesia or numbness in the feet increased, whereas for burning and pain in the feet the corresponding percentages were 56 and 61%. However, 33 and 40% of the respondents with type 2 diabetes and without diabetes, respectively, reporting neuropathic symptoms at follow up did not receive any pharmacotherapy. Pharmacotherapy of neuropathic symptoms at follow up among participants with type 2 diabetes and without diabetes included mainly World Health Organization Step 1 analgesics (17% each; excluding acetylsalicylic acid), pregabalin/gabapentin (20 and 12%), vitamin B complex (13 and 22%), benfotiamine (13 and 2%), opioids (7 and 12%), antidepressants (4 and 5%) and α-lipoic acid (4 and 2%). CONCLUSIONS: These findings point to insufficient care, inadequate treatment adherence or limited efficacy of treatments in patients with polyneuropathy, suggesting that effective measures should be implemented to correct these healthcare deficits.