Analytical Challenges in Diabetes Management: Towards Glycated Albumin Point-of-Care Detection

Diabetes mellitus is a worldwide-spread chronic metabolic disease that occurs when the pancreas fails to produce enough insulin levels or when the body fails to effectively use the secreted pancreatic insulin, eventually resulting in hyperglycemia. Systematic glycemic control is the only procedure at our disposal to prevent diabetes long-term complications such as cardiovascular disorders, kidney diseases, nephropathy, neuropathy, and retinopathy. Glycated albumin (GA) has recently gained more and more attention as a control biomarker thanks to its shorter lifespan and wider reliability compared to glycated hemoglobin (HbA1c), currently the “gold standard” for diabetes screening and monitoring in clinics. Various techniques such as ion exchange, liquid or affinity-based chromatography and immunoassay can be employed to accurately measure GA levels in serum samples; nevertheless, due to the cost of the lab equipment and complexity of the procedures, these methods are not commonly available at clinical sites and are not suitable to home monitoring. The present review describes the most up-to-date advances in the field of glycemic control biomarkers, exploring in particular the GA with a special focus on the recent experimental analysis techniques, using enzymatic and affinity methods. Finally, analysis steps and fundamental reading technologies are integrated into a processing pipeline, paving the way for future point-of-care testing (POCT). In this view, we highlight how this setup might be employed outside a laboratory environment to reduce the time from measurement to clinical decision, and to provide diabetic patients with a brand-new set of tools for glycemic self-monitoring

Evaluation of Glycated Albumin (GA) and GA/Hba1c Ratio for Diagnosis of Diabetes and Glycemic Control: A Comprehensive Review

Diabetes Mellitus (DM) is a group of metabolic diseases characterized by chronic high blood glucose concentrations (hyperglycemia). When it is left untreated or improperly managed, it can lead to acute complications including diabetic ketoacidosis and non-ketotic hyperosmolar coma. In addition, possible long-term complications include impotence, nerve damage, stroke, chronic kidney failure, cardiovascular disease, foot ulcers, and retinopathy. Historically, universal methods to measure glycemic control for the diagnosis of diabetes included fasting plasma glucose level (FPG), 2-h plasma glucose (2HP), and random plasma glucose. However, these measurements did not provide information about glycemic control over a long period of time. To address this problem, there has been a switch in the past decade to diagnosing diabetes and its severity through measurement of blood glycated proteins such as Hemoglobin A1c (HbA1c) and glycated albumin (GA). Diagnosis and evaluation of diabetes using glycated proteins has many advantages including high accuracy of glycemic control over a period of time. Currently, common laboratory methods used to measure glycated proteins are high-performance liquid chromatography (HPLC), immunoassay, and electrophoresis. HbA1c is one of the most important diagnostic factors for diabetes. However, some reports indicate that HbA1c is not a suitable marker to determine glycemic control in all diabetic patients. GA, which is not influenced by changes in the lifespan of erythrocytes, is thought to be a good alternative indicator of glycemic control in diabetic patients. Here, we review the literature that has investigated the suitability of HbA1c, GA and GA:HbA1c as indicators of long-term glycemic control and demonstrate the importance of selecting the appropriate glycated protein based on the patient’s health status in order to provide useful and modern point-of-care monitoring and treatment

Diabetes and baseline glucose are associated with inflammation, left ventricular function and short- and long-term outcome in acute coronary syndromes: role of the novel biomarker Cyr 61.

Hyperglycemia in the setting of an acute coronary syndrome (ACS) impacts short term outcomes, but little is known about longer term effects. We therefore designed this study to firstly determine the association between hyperglycemia and short term and longer term outcomes in patients presenting with ACS and secondly evaluate the prognostic role of diabetes, body mass index (BMI) and the novel biomarker Cyr61 on outcomes. The prospective Special Program University Medicine-Acute Coronary Syndrome (SPUM-ACS) cohort enrolled 2168 patients with ACS between December 2009 and October 2012, of which 2034 underwent PCI (93.8%). Patients were followed up for 12 months. Events were independently adjudicated by three experienced cardiologists. Participants were recruited from four tertiary hospitals in Switzerland: Zurich, Geneva, Lausanne and Bern. Participants presenting with acute coronary syndromes and who underwent coronary angiography were included in the analysis. Patients were grouped according to history of diabetes (or HbA1c greater than 6%), baseline blood sugar level (BSL; < 6, 6-11.1 and > 11.1 mmol/L) and body mass index (BMI). The primary outcome was major adverse cardiac events (MACE) which was a composite of myocardial infarction, stroke and all-cause death. Secondary outcomes included the individual components of the primary endpoint, revascularisations, bleeding events (BARC classification) and cerebrovascular events (ischaemic or haemorrhagic stroke or TIA). Patients with hyperglycemia, i.e. BSL ≥ 11.1 mmol/L, had higher levels of C-reactive protein (CRP), white blood cell count (WBC), creatinine kinase (CK), higher heart rates and lower left ventricular ejection fraction (LVEF) and increased N-terminal pro-brain natriuretic peptide. At 30 days and 12 months, those with BSL ≥ 11.1 mmol/L had more MACE and death compared to those with BSL < 6.0 mmol/L or 6.0-11.1 mmol/L (HR-ratio 4.78 and 6.6; p < 0.001). The novel biomarker Cyr61 strongly associated with high BSL and STEMI and was independently associated with 1 year outcomes (HR 2.22; 95% CI 1.33-3.72; Tertile 3 vs. Tertile 1). In this large, prospective, independently adjudicated cohort of in all comers ACS patients undergoing PCI, both a history of diabetes and elevated entry glucose was associated with inflammation and increased risk of MACE both at short and long-term. The mediators might involve increased sympathetic activation, inflammation and ischemia as reflected by elevated Cyr61 levels leading to larger levels of troponin and lower LVEF. Trial registration Clinical Trial Registration Number: NCT01000701. Registered October 23, 2009

Development of non-esterified fatty acid (NEFA) electrochemical biosensor for energy metabolism studies

PhD ThesisThere are many energy metabolism studies ongoing, including those for cardiovascular diseases and type-2-diabetes. With an increase in people being diagnosed with type-2-diabetes, there should be more ways to monitor not only the blood glucose levels but also the other biomarkers associated with type-2-diabetes. The metabolism biomarkers are essential in understanding the cause of diabetes early on. These biomarkers include: glucose, non-esterified fatty acid, lactate, urea, creatinine, glycosylated haemoglobin and cholesterol. Whilst glucose measurement has a clear role in type-2-diabetes management, the potential value of non-esterified fatty acid has not been explored or highlighted yet. The aim of this project is to develop an electrochemical biosensor for the non-esterified fatty acid in human blood, as non-esterified fatty acid can cause -cell loss in type-2-diabetes. Exploration of this biomarker would be a step forward in increasing research and patient understanding of the dynamic processes involved in establishing good metabolism control. The project uses the enzymes in commercial optical methods for non-esterified fatty acid detection. Oleic acid was used as the standard non-esterified fatty acid in this work. The electrochemical techniques employed are cyclic voltammetry, linear sweep voltammetry, chronoamperometry and electrochemical impedance spectroscopy. Enzyme electrodes were fabricated using the layer-by-layer immobilization of alternating polymer and enzyme combinations on carbon, cobalt phthalocyanine and single wall carbon nanotube screen printed electrodes. A chronoamperometric non-esterified fatty acid sensor was developed with the linear detection range of 0.10 mM to 0.90 mM oleic acid and with a sensitivity of 0.6562 A/mM oleic acid. This sensor was then further fabricated to detect non-esterified fatty acid concentrations in human plasma and serum samples. Commercial UV optical methods were used as method of validation of the blood sample concentrations. This work produced a platform for further non-esterified fatty acid detection studies.EPSRC

A Comparison of Screening Tools for the Early Detection of Peripheral Neuropathy in Adults With and Without Type 2 Diabetes

Objective. Examine the effectiveness of the 128 Hz tuning fork, two monofilaments, and Norfolk Quality of Life Diabetic Neuropathy (QOL-DN) questionnaire as tools for the early detection of diabetic peripheral neuropathy (DPN) in overweight, obese, and inactive (OOI) adults or those who have prediabetes (PD) or type 2 diabetes (T2D). Research Design and Methods. Thirty-four adults (mean age 58.4 years +/- 12.1) were divided by glycemia (10 OOI normoglycemic, 13 PD, and 11 T2D). Sural nerves were tested bilaterally with the NC-stat DPNCheck to determine sural nerve amplitude potential (SNAP) and sural nerve conduction velocity (SNCV). All other testing results were compared to SNAP and SNCV. Results. Total 1 g monofilament scores significantly correlated with SNAP values and yielded the highest sensitivity and specificity combinations of tested measures. Total QOL-DN scores negatively correlated with SNAP values, as did QOL-DN symptoms. QOL-DN activities of daily living correlated with the right SNAP, and the QOL-DN small fiber subscore correlated with SNCV. Conclusions. The 1 g monofilament and total QOL-DN are effective, low-cost tools for the early detection of DPN in OOI, PD, and T2D adults. The 128 Hz tuning fork and 10 g monofilament may assist DPN screening as a tandem, but not primary, early DPN detection screening tools

NO2Probe: Application of a new nitrite point-of-care test in biomedicine

Periodontal disease (PD) results from the inflammatory response to the presence of bacterial biofilms. The disease progression goes through two phases: gingivitis, characterized by bleeding and gingiva gums swelling, and periodontitis, an aggravated condition that can lead to bone, gingival, and teeth loss. For diagnosis, time-consuming periodontal charts and X-rays are evaluated. Thus, it is convenient to recognize clinical biomarkers measuring each individual's PD occurrence and extent. The nitrite (NO2-) level in saliva has been pointed out in this regard, but it is not yet clinically used as a PD biomarker. Thus, the study objectives were to evaluate the relationship between PD and nitrite concentration in non-stimulated saliva samples, and the implementation of a new specific and sensitive biosensor, the NO2Probe, for nitrite quantification during the medical appointment. The approach involved the screen-printed carbon electrodes (SPE) modification and optimization with the multihemic nitrite reductase enzyme (ccNiR), which catalyzes the direct reduction reaction of nitrite into ammonia. As an oxygen scavenging system, an ascorbate oxidase (AOx) layer was added and, finally, a polymer layer (PVA), to “delay” the ccNiR saturation. The amperometric biosensor (SPE/ccNiR/AOx/PVA) covers a linearity range between 5-300 μM, with a sensitivity of 0.015 μM-1. The obtained values were compared with the conventional Griess method. The biosensor was statistically validated. Initially, the centrifugation and freezing steps of the samples were evaluated, evaluating the two methods, and verifying that, with the biosensor, centrifugation is not necessary. In the Griess method, centrifugation is essential, as the nitrite concentration in non-centrifuged samples (199 ± 3 μM) decreases after centrifugation (68.8 ± 0.5 μM). When frozen, the NO2- concentration decreases, probably due to the NO2- instability. It was concluded that samples should be analyzed fresh as soon as possible. In the pilot study, carried out at the CiiEM Dental Clinic, it was not possible to establish a concrete relationship between the NO2- concentration and the disease stages and grades. Contrary to the expectations, the periodontally healthy group (n=11) had higher NO2- concentration (257 ± 30 μM) higher than those observed in the PD patients group (n=24).A Doença Periodontal (PD) decorre da resposta inflamatória à presença de biofilmes bacterianos. A progressão desta, passa por duas fases: a gengivite, caracterizada pelo sangramento e inchaço da gengiva, e a periodontite, condição agravada que pode conduzir à perda de tecidos ósseo, gengival e de dentes. Para diagnóstico, são avaliados morosos periodontogramas e os raios-X. Assim, é conveniente o reconhecimento de biomarcadores clínicos aferindo a ocorrência e extensão da PD em cada indivíduo. O nível de nitrito (NO2-) na saliva, tem sido apontado nesse sentido, mas ainda não é utilizado clinicamente como biomarcador da PD. Assim, os objetivos foram: avaliar a relação entre a PD e a concentração de nitrito, em amostras de saliva não estimulada, e a implementação de um novo biossensor, o NO2Probe, específico e sensível para quantificação de nitrito durante a consulta. A abordagem passou pela modificação e otimização de elétrodos impressos de carbono (SPE) com a enzima multihémica redutase do nitrito (ccNiR), que catalisa a reação de redução direta do nitrito a amónia. Como sistema de remoção de oxigénio, adicionou-se uma camada de Ascorbato oxidase (AOx) e, finalmente, uma camada de polímero (PVA), para “atrasar” a saturação da ccNiR. O biossensor amperometrico (SPE/ccNiR/AOx/PVA) mostrou abranger uma gama de linearidade entre 5-300 μM, com sensibilidade de 0.015 μM-1. Os valores obtidos foram comparados com o método convencional de Griess. Estatisticamente, o biossensor foi validado. Inicialmente, foram avaliados os passos centrifugação e congelamento das amostras avaliando os dois métodos, verificando-se que, com o biossensor, não é necessário centrifugar amostras. Já pelo método de Griess, a centrifugação é fundamental, pois a concentração de nitrito em amostras não centrifugadas (199 ± 3 μM) decresce após centrifugação (68.8 ± 0.5 μM). Quando congeladas, há diminuição da concentração de nitrito, provavelmente devido à instabilidade deste. Concluiu-se que as amostras devem ser analisadas de fresco, o mais rápido possível. No estudo piloto, conduzido na Clínica Dentária do CiiEM, não foi possível estabelecer uma relação concreta entre a concentração de nitrito e os estadios e graus da doença. Contrariamente ao esperado, o grupo composto por indivíduos periodontalmente saudáveis (n=11), apresentaram concentrações de nitrito (257 ± 30 μM) superiores às verificadas no grupo de doentes (n=24)

Emerging Theranostic Nanomaterials in Diabetes and Its Complications

Diabetes mellitus (DM) refers to a group of metabolic disorders that are characterized by hyperglycemia. Oral subcutaneously administered antidiabetic drugs such as insulin, glipalamide, and metformin can temporarily balance blood sugar levels, however, long-term administration of these therapies is associated with undesirable side effects on the kidney and liver. In addition, due to overproduction of reactive oxygen species and hyperglycemia-induced macrovascular system damage, diabetics have an increased risk of complications. Fortunately, recent advances in nanomaterials have provided new opportunities for diabetes therapy and diagnosis. This review provides a panoramic overview of the current nanomaterials for the detection of diabetic biomarkers and diabetes treatment. Apart from diabetic sensing mechanisms and antidiabetic activities, the applications of these bioengineered nanoparticles for preventing several diabetic complications are elucidated. This review provides an overall perspective in this field, including current challenges and future trends, which may be helpful in informing the development of novel nanomaterials with new functions and properties for diabetes diagnosis and therapy.Peer reviewe

Diagnostic circulating biomarkers to detect vision-threatening diabetic retinopathy: Potential screening tool of the future?

With the increasing prevalence of diabetes in developing and developed countries, the socio-economic burden of diabetic retinopathy (DR), the leading complication of diabetes, is growing. Diabetic retinopathy (DR) is currently one of the leading causes of blindness in working-age adults worldwide. Robust methodologies exist to detect and monitor DR; however, these rely on specialist imaging techniques and qualified practitioners. This makes detecting and monitoring DR expensive and time-consuming, which is particularly problematic in developing countries where many patients will be remote and have little contact with specialist medical centres. Diabetic retinopathy (DR) is largely asymptomatic until late in the pathology. Therefore, early identification and stratification of vision-threatening DR (VTDR) is highly desirable and will ameliorate the global impact of this disease. A simple, reliable and more cost-effective test would greatly assist in decreasing the burden of DR around the world. Here, we evaluate and review data on circulating protein biomarkers, which have been verified in the context of DR. We also discuss the challenges and developments necessary to translate these promising data into clinically useful assays, to detect VTDR, and their potential integration into simple point-of-care testing devices