Glycated Human Serum Albumin Research Articles

Advances in electrochemical biosensors employing carbon-based electrodes for detection of biomarkers in diabetes mellitus.

The increase in diabetes cases has become a major concern in the healthcare sector, necessitating the development of efficient and minimal diagnostic methods. This study aims to provide a comprehensive examination of electrochemical biosensors for detecting diabetes mellitus biomarkers, with a special focus on the utilization of carbon-based electrodes. A detailed analysis of electrochemical biosensors incorporating various carbon electrodes, including screen-printed carbon electrodes, glassy carbon electrodes, and carbon paste electrodes, is presented. The advantages of carbon-based electrodes in biosensor design are highlighted. The review covers the detection of several key diabetes biomarkers, such as glucose, glycated hemoglobin (HbA1c), glycated human serum albumin (GHSA), insulin, and novel biomarkers. Recent developments in electrochemical biosensor technology over the last decade are summarized, emphasizing their potential in clinical applications, particularly in point-of-care settings. The utilization of carbon-based electrodes in biosensors is shown to offer significant advantages, including enhanced sensitivity, selectivity, and cost-effectiveness. This review underscores the importance of carbon-based electrodes in the design of electrochemical biosensors and raises awareness for the detection of novel biomarkers for more specific and personalized diabetes mellitus cases. The advancements in this field highlight the potential of these biosensors in future clinical applications, especially in point-of-care diagnostics.

Aggregation of Apo/Glycated Human Serum Albumins and Aptamer-Saturated Graphene Quantum Dot: A Simulation Study.

Human serum albumin (HSA) is a protein carrier that transports a wide range of drugs and nutrients. The amount of glycated HSA (GHSA) is used as a diabetes biomarker. To quantify the GHSA amount, the fluorescent graphene-based aptasensor has been a successful method. In aptasensors, the key mechanism is the adsorption/desorption of albumin from the aptamer-graphene complex. Recently, the graphene quantum dot (GQD) has been reported to be an aptamer sorbent. Due to its comparable size to aptamers, it is attractive enough to explore the possibility of GQD as a part of an albumin aptasensor. Therefore, molecular dynamics (MD) simulations were performed here to reveal the binding mechanism of albumin to an aptamer-GQD complex in molecular detail. GQD saturated by albumin-selective aptamers (GQDA) is studied, and GHSA and HSA are studied in comparison to understand the effect of glycation. Fast and spontaneous albumin-GQDA binding was observed. While no specific GQDA-binding site on both albumins was found, the residues used for binding were confined to domains I and III for HSA and domains II and III for GHSA. Albumins were found to bind preferably to aptamers rather than to GQD. Lysines and arginines were the main contributors to binding. We also found the dissociation of GLC from all GHSA trajectories, which highlights the role of GQDA in interfering with the ligand binding affinity in Sudlow site I. The binding of GQDA appears to impair albumin structure and function. The insights obtained here will be useful for the future design of diabetes aptasensors.

Alleviation of albumin glycation-induced diabetic cardiomyopathy by L-Arginine: Insights into Nrf-2 signaling

In hyperglycemia, accelerated glycation and oxidative stress give rise to many diabetic complications, such as diabetic cardiomyopathy (DCM). Glycated human serum albumin (GHSA) has disturbed structural integrity and hampered functional capabilities. When GHSA accumulates around cardiac cells, Nrf-2 is dysregulated, aiding oxidative stress. L-Arginine (L-Arg) is prescribed to patients with diabetes and cardiovascular diseases. This research contributes to the mechanistic insights on antiglycation and antioxidant potential of L-Arg in alleviating DCM. HSA was glycated with methylglyoxal in the presence of L-Arg (20–640 mM). Structural and functional modifications of HSA were studied. L-Arg and HSA, GHSA interactions, and thermodynamics were determined by steady-state fluorescence. H9c2 cardiomyocytes were given treatments of GHSA-L-Arg along with the inhibitor of the receptor of AGEs. Cellular antioxidant levels, detoxification enzyme activities were measured. Gene, protein expressions, and immunofluorescence data examined the activation and nuclear translocation of Nrf-2 during glycation and oxidative stress. L-Arg protected HSA from glycation-induced structural and functional modifications. The binding affinity of L-Arg was more towards HSA (104 M−1). L-Arg, specifically at lower concentration (20 mM), upregulated Nrf-2 gene, protein expressions and facilitated its nuclear translocation by activating Nrf-2 signaling. The study concluded that L-Arg can be of therapeutic advantage in glycation-induced DCM and associated oxidative stress.

Binding of Apo and Glycated Human Serum Albumins to an Albumin-Selective Aptamer-Bound Graphene Quantum Dot Complex.

Diabetes mellitus is a chronic metabolic disease involving continued elevated blood glucose levels. It is a leading cause of mortality and reduced life expectancy. Glycated human serum albumin (GHSA) has been reported to be a potential diabetes biomarker. A nanomaterial-based aptasensor is one of the effective techniques to detect GHSA. Graphene quantum dots (GQDs) have been widely used in aptasensors as an aptamer fluorescence quencher due to their high biocompatibility and sensitivity. GHSA-selective fluorescent aptamers are first quenched upon binding to GQDs. The presence of albumin targets results in the release of aptamers to albumin and consequently fluorescence recovery. To date, the molecular details on how GQDs interact with GHSA-selective aptamers and albumin remain limited, especially the interactions of an aptamer-bound GQD (GQDA) with an albumin. Thus, in this work, molecular dynamics simulations were used to reveal the binding mechanism of human serum albumin (HSA) and GHSA to GQDA. The results show the rapid and spontaneous assembly of albumin and GQDA. Multiple sites of albumins can accommodate both aptamers and GQDs. This suggests that the saturation of aptamers on GQDs is required for accurate albumin detection. Guanine and thymine are keys for albumin-aptamer clustering. GHSA gets denatured more than HSA. The presence of bound GQDA on GHSA widens the entrance of drug site I, resulting in the release of open-chain glucose. The insight obtained here will serve as a base for accurate GQD-based aptasensor design and development.

Structural and Dynamic Alteration of Glycated Human Serum Albumin in Schiff Base and Amadori Adducts: A Molecular Simulation Study.

Human serum albumin (HSA) is a protein carrier in blood transporting metabolites and drugs. Glycated HSA (GHSA) acts as a potential biomarker for diabetes. Thus, many attempts have been made to detect GHSA. Glycation was reported to damage the structure and ligand binding capability, where no molecular detail is available. Recently, the crystal structure of GHSA has been solved, where two glucose isomers (pyranose/GLC and open-chain/GLO) are located at Sudlow's site I. GLO was found to covalently bind to K195, while GLC is trapped by noncontact interactions. GHSA exists in two forms (Schiff base (SCH) and Amadori (AMA) adducts), but how both disrupt albumin activity microscopically remains unknown. To this end, molecular dynamics simulations were performed here to explore the nature of SCH and AMA. Both forms are found to alter the main protein dynamics, resulting in (i) the widening of Sudlow's site I entrance, (ii) the size reduction of nine fatty acid-binding pockets, (iii) the enlargement of Sudlow's site I and the shrinking of Sudlow's site II, (iv) the enhancement of C34 reactivity, and (v) the change in the W214 microenvironment. These unique characteristics found here can be useful for understanding the effect of glycation on the albumin function in more detail and designing specific and selective GHSA detection strategies.

Flexible multielectrode arrays based electrochemical aptasensor for glycated human serum albumin detection

Assessment tests for glycemic control of patients with diabetes mellitus commonly use blood glucose and glycated hemoglobin A1C (HbA1 C) levels for estimating acute and long-term glycemia, respectively. Diabetes diet, as well as treatment, are adapted accordingly and also the risk of diabetic complications can be estimated. Glycated human serum albumin (GHSA) has been proposed as alternative to HbA1C since it possesses a four times shorter lifespan and its blood concentration is not affected by hemic diseases. Here, an electrochemical aptasensor is proposed for the simultaneous detection of human serum albumin (HSA) and GHSA utilizing flexible multielectrode arrays. The polymer chip carries two individual sets of gold electrodes, which are separately modified by the corresponding aptamers. The dual-target aptasensors possessed detection limits as low as 13 nM and 25 nM for HSA and GHSA, respectively, together with detection ranges lasting from 40 nM to 10 µM. Moreover, our aptasensor quantified selectively its target in diluted full-blood samples. This first PoC, low-cost plastic aptasensor facilitates the simultaneous and quantitative determination of HSA and GHSA ratios by analyzing the current signals from several redundant electrodes in the same sample, which permits a reliable and affordable estimation of long-term glycemia at point-of-care or home.

The Burden of Impaired Serum Albumin Antioxidant Properties and Glyco-Oxidation in Coronary Heart Disease Patients with and without Type 2 Diabetes Mellitus.

Human serum albumin (HSA) has an important antioxidant activity due to the presence of the reduced cysteine at position 34, which represents the most abundant free thiol in the plasma. In oxidative-based diseases, HSA undergoes S-thiolation (THIO-HSA) with changes in the antioxidant function of albumin that could contribute to the progression of the disease. The aim of this study was to verify, for the first time, the different burdens of THIO-HSA, glycated HSA (GLY-HSA), and advanced glycation end products (AGE) accumulation both in type 2 diabetes mellitus (T2DM) patients and in non-diabetic patients, with or without coronary heart disease (CHD). In this study, we assessed the presence of modified forms of HSA, THIO-HSA, and GLY-HSA by means of mass spectrometry in 33 patients with both T2DM and CHD, in 31 patients with T2DM and without CHD, in 30 patients without diabetes with a history of CHD, and 27 subjects without diabetes and CHD. All the patients’ anthropometric and clinical data were recorded including age, sex, duration of diabetes, body mass index (BMI), blood pressure, and history of CHD defined with anamnestic data. Metabolic parameters, such as fasting plasma glucose (FPG), glycated hemoglobin (HbA1c), lipids, pentosidine, AGE, receptor for advanced glycation end-products (RAGE) and its soluble form (sRAGE), were measured. AGE and pentosidine are significantly higher in T2DM patients with and without CHD with respect to non-diabetic patients with CHD and control subjects. RAGE levels are significantly higher in T2DM patients with respect to non-diabetic patients, and among T2DM patients, the group with CHD showed significantly higher RAGE levels than those without CHD (217 ± 171 pg/mL and 140 ± 61 pg/mL, respectively). Albumin isoforms discriminate between non-diabetic patients with CHD and T2DM patients with and without CHD and control subjects, with GLY-HSA levels higher in T2DM with and without CHD, and THIO-HSA higher in CHD patients without T2DM. Finally, we demonstrated that the oxidized forms of HSA can increase the expression of the inflammatory cytokine Tumor Necrosis Factor-alpha (TNFα) in monocytic cells. In patients with CHD, GLY-HSA and THIO-HSA have a different prevalent distribution, the first one prevailing in patients with T2DM and the second one in patients without T2DM. These findings suggest that albumin quality and homeostasis balance between glyco-oxidation and thiolation might have an impact on the antioxidant defense system in cardiovascular diseases.

Glycation end-products specific auto-antibodies in Systemic Lupus Erythematosus.

Systemic lupus erythematosus (SLE) is an autoimmune disease, which is highly inflammatory. Compared to a healthy control group, SLE patients exhibit a higher concentration of advanced glycation end products (AGEs) and a lower concentration of receptors for AGEs (RAGE) in serum, however, the exact aetiology is still unclear. In the present study, non-enzymatic glycation induced modification of human serum albumin (HSA) has been studied by biophysical techniques. Glycated HSA (G-HSA) was used as an antigen, and serum autoantibody levels were estimated in SLE and normal humans (NH) against it, using direct binding ELISA and competitive inhibition ELISA. Compared to N-HSA, remarkable structural modifications were observed in G-HSA. Modified HSA also showed increased pentosidine fluorescence (213.7 ± 13.4 AU). Glycation of HSA induced a conversion of α-helix and random coil to β-sheet and β-turns. Serum immuno assays results exhibited significantly (p < 0.001) higher binding of G-HSA with serum autoantibodies from SLE patients when compared with native HSA (N-HSA). Furthermore, competitive ELISA results showed significantly (p < 0.001) high percent inhibition of serum IgG from SLE patients with modified antigen. Chronic inflammation with excessive oxidative stress in SLE patients could be a possible reason for structural alterations in blood proteins, generating highly immunogenic unique new-epitopes. These in turn induce the generation of specific autoantibodies against G-HSA that may serve as a potential biomarker for SLE pathogenesis.

Understanding the adsorption of glycated human serum albumin on aptamer-modified graphene for diabetes aptasensor design

Glycated Human Serum Albumin (GHSA) (glucose-bound albumin) is found to act as a diabetes biomarker. Many attempts have been made to quantify GHSA level. Fluorescent aptasensor is one of potential techniques for detect GHSA. To detect GHSA, GHSA-selective aptamer is quenched by graphene surface. A presence of GHSA in a sample causes the desorption of aptamer to form the aptamer-GHSA complex in solution. The mechanism of GHSA-aptamer-GRA complex is important for aptasensor efficiency. This step is a key for aptasensor function.

The binding of apo and glucose-bound human serum albumins to a free graphene sheet in aqueous environment: Simulation studies

Human serum albumin (HSA) is a blood protein serving as a carrier for a wide range of drugs and nutrients. A level of glycated HSA (GHSA) is used as a diabetes biomarker. A graphene-based aptasensor is one of potential techniques to detect GHSA. Not only the interactions of albumin and aptamer, but the albumin-graphene (GRA) binding mechanism are also crucial for developing a diabetes aptasensor. In this work, Molecular Dynamics simulations (MD) were employed to explore the binding of GRA to both GHSA and HSA. The GRA binding from the back and front sides of an albumin are fast and spontaneous. The multiple GRA binding sites are identified. GRA causes more denaturation of helical characteristics in GHSA (∼12% reduction of helical structure). Both back and front GRA adhesions generate comparable degrees of helical unfolding. Importantly, the presence of bound GRA induces the release of glucose from drug sites implying the loss of ligand-binding affinity. This loss of drug site activity is independent on the GRA binding positions because all bound positions lead to the exit of sugars. The GRA binding deconstructs not only secondary structure, but also albumin function. Apparently, GRA is a non-biocompatible material for albumin. To construct a potential graphene-based aptasensor to detect GHSA, it is necessary to be certain that no free GRA surface is available because a bare GRA can bind and denature both HSA and GHSA which can cause misleading data.

Resolving interactions of miglitol with normal and glycated human serum albumin by multivariate methods

This article reports results of one of our projects related to the investigation of interactions of miglitol (MIG) with normal human serum albumin (HSA) and glycated HSA (GHSA) with the help of recording spectroscopic and electrochemical data. The experimental data were analyzed by conventional and chemometric methods to extract useful information for comprehensive justifications of the interactions of the MIG with HSA and GHSA. Hard- and soft-modeling chemometric methods were used to extract quantitative and qualitative information. Then, molecular docking techniques were used to further investigation of the binding of the MIG with HSA and GHSA and the extracted results were compatible with those obtained by experimental methods. Finally, according to the binding of the BV with HSA and GHSA, second-order differential pulse voltammetric data were recorded and calibrated with three-way calibration methods for exploiting second-order advantage for determination of the GHSA in the presence of the HSA to develop a novel chemometrics assisted-electroanalytical method for diagnostic and monitoring of diabetic.

Chemometrics-assisted multi-instrumental techniques for investigation of interactions of dapagliflozin with normal and glycated human serum albumin: Application to exploiting second-order advantage for determination of glycated human serum albumin as a biomarker for controlling diabetes

In this paper, we have collected useful information about interactions of dapagliflozin (DPA) with normal human serum albumin (HSA) and glycated HSA (GHSA) by using spectroscopic and electrochemical methods. Instrumental data were analyzed by graphical and conventional methods and then, the data were mathematically modeled and simultaneously analyzed by multivariate curve resolution-alternating least squares (MCR-ALS). Hard-modeling algorithm of the data helped us to obtain qualitative and quantitative information to cast a deep look at the interactions of the DPA with HSA and GHSA. The results of MCR-BANDS showed that the MCR-ALS gave us unambiguous and reliable results. Afterwards, molecular docking techniques were used to investigate binding of the DPA with HSA and GHSA to obtain new information and to complete the previous findings. The DPA was bound with HSA and GHSA with different binding strengths. Fortunately, the results were compatible and confirmed each other. Finally, according to the results of interaction and binding of the DPA with HSA and GHSA, a second-order calibration model was developed with aim of exploiting second-order advantage for determination of the GHSA in the presence of HSA as uncalibrated interference. The developed method had a good performance for determination of the GHSA in both of synthetic and real samples.

Immobilization-Free Electrochemical Sensor Coupled with a Graphene-Oxide-Based Aptasensor for Glycated Albumin Detection.

An immobilization-free electrochemical sensor coupled with a graphene oxide (GO)-based aptasensor was developed for glycated human serum albumin (GHSA) detection. The concentration of GHSA was monitored by measuring the electrochemical response of free GO and aptamer-bound GO in the presence of glycated albumin; their currents served as the analytical signals. The electrochemical aptasensor exhibited good performance with a base-10 logarithmic scale. The calibration curve was achieved in the range of 0.01–50 µg/mL. The limit of detection (LOD) was 8.70 ng/mL. The developed method was considered a one-drop measurement process because a fabrication step and the probe-immobilization process were not required. This simple sensor offers a cost-effective, rapid, and sensitive detection method, and could be an alternative approach for determination of GHSA levels.

Exploring the Binding Affinities of Fructose and Galactose to Human Serum Albumin

Human serum albumin (HSA) is the most abundant transport protein found in human blood. It is known to bind a wide range of drugs, metabolites, and nutrients. The glycation of HSA causes abnormal biological effects. Importantly, glycated HSA (GHSA) is of interest as a potential diabetes biomarker. Many biosensor studies successfully used GHSA as a diabetes biomarker. Recently, a crystal structure of glycated HSA has been reported where a glucose dimer were found in a drug-binding site. HSA can also bind fructose and galactose, but no in-depth binding mechanism is available. To explore binding mechanisms of these sugars, we therefore employed molecular dynamics (MD) simulations and residues interaction energy (INT) calculations to obtain microscopic details in comparison to glucose-bound system form a previous work. Our results show that glucose is the most preferrable sugar for HSA. However, comparing between fructose and galactose, HSA prefers fructose to galactose. Being dimer is a key for tight binding. The movements of top domains are also crucial for sugar binding affinities.Furthermore, key interactions that promote the sugar binding are also extracted here. An insight obtained here will play a role in design and development of diabetes biosensor.

Dual sensitive and rapid detection of glycated human serum albumin using a versatile lead/graphene nanocomposite probe as a fluorescence-electrochemical aptasensor.

Monitoring of glycated human serum albumin (GHSA) as a glycemic marker for screening and monitoring of diabetes mellitus is widely practiced for patients with conditions that affect red blood cells. In this study, a complex comprising Pb ions adsorbed on graphene oxide (GO-Pb) was fabricated and utilized as a versatile probe in a fluorescence-electrochemical aptasensor for GHSA quantification. To simplify the aptasensor, the GO-Pb complex probe was prepared via an ion adsorption process. After modification with a fluorophore-labeled aptamer, the GO-Pb complex served as an excellent energy acceptor in fluorescence-based analysis, as well as generating a high current in the electrochemical transducer. Additionally, the proposed platform can detect GHSA via the dual technique from a single sample, allowing for precise and accurate results. Under optimal conditions, the fluorescence-electrochemical aptasensor exhibited a linear relationship with GHSA concentrations from 0.001 to 80 μg mL-1 and from 0.005 to 10 μg mL-1 for fluorescence and electrochemical detection, respectively. The corresponding detection limits were 8.80 ng mL-1 and 0.77 ng mL-1, respectively. The proposed aptasensor additionally displayed good selectivity and excellent stability. Moreover, its successful application in the analysis of clinical samples further demonstrated its utility. Therefore, the proposed platform has significant potential as a novel, facile, highly responsive, and low-cost monitoring method for the development of diabetes mellitus diagnostic devices intended for a clinical setting.

Two- and three-way chemometric analyses for investigation of interactions of acarbose with normal and glycated human serum albumin: Developing a novel biosensing system

In this project, our research group has performed an interesting work in which interactions of acarbose (ACB) with normal human serum albumin (HSA) and glycated HSA (GHSA) have been investigated by chemometrics assisted-electrochemical and spectroscopic techniques. To have a deep insight to the interactions of ACB with HSA and GHSA, different electrochemical and spectroscopic techniques were used to monitor ACB-HSA and ACB-GHSA interactions and analyzed by multivariate curve resolution alternating least squares (MCR-ALS), MCR-BANNDS and EQUISPEC as efficient chemometric algorithms. Then, molecular docking techniques were used to extract more information which helped us to better justify binding of ACB with HSA and GHSA. After obtaining qualitative and quantitative information and justification of the ACB-HSA and ACB-GHSA interactions, a novel electrochemical technique was developed for exploiting second-order advantage from differential pulse voltammetric data for determination of GHSA as a potential biomarker in diabetes in the presence of HSA. Calibration and validation of the developed system showed us that our system was successful in determination of GHSA in synthetic and real samples.

Development of a Novel ssDNA Sequence for a Glycated Human Serum Albumin and Construction of a Simple Aptasensor System Based on Reduced Graphene Oxide (rGO).

Diabetes is one of the top 10 global causes of death. About one in 11 global adults have diabetes. As the disease progresses, the mortality rate increases, and complications can develop. Thus, early detection and effective management of diabetes are especially important. Herein, we present a novel glycated human serum albumin (GHSA) aptamer, i.e., GABAS-01, which has high affinity and specificity. The aptamer was selected by reduced graphene oxide-based systematic evolution of ligands by exponential enrichement (rGO-based SELEX) against GHSA. After five rounds of selection through gradually harsher conditions, GABAS-01 with high affinity and specificity for the target was obtained. GABAS-01 was labeled by FAM at the 5′-end and characterized by measuring the recovery of a fluorescence signal that is the result of fluorescence quenching effect of rGO. As a result, GABAS-01 had low-nanomolar Kd values of 1.748 ± 0.227 nM and showed a low limit of detection of 16.40 μg/mL against GHSA. This result shows the potential application of GABAS-01 as an effective on-site detection probe of GHSA. In addition, these properties of GABAS-01 are expected to contribute to detection of GHSA in diagnostic fields.

Involvement of glycated albumin in adipose-derived-stem cell-mediated interleukin 17 secreting T helper cell activation

BACKGROUNDAdvanced glycation end products (AGE) are a marker of various diseases including diabetes, in which they participate to vascular damages such as retinopathy, nephropathy and coronaropathy. Besides those vascular complications, AGE are involved in altered metabolism in many tissues, including adipose tissue (AT) where they contribute to reduced glucose uptake and attenuation of insulin sensitivity. AGE are known to contribute to type 1 diabetes (T1D) through promotion of interleukin (IL)-17 secreting T helper (Th17) cells.AIMTo investigate whether lean adipose-derived stem cells (ASC) could be able to induce IL-17A secretion, with the help of AGE.METHODSAs we have recently demonstrated that ASC are involved in Th17 cell promotion when they are harvested from obese AT, we used the same co-culture model to measure the impact of glycated human serum albumin (G-HSA) on human lean ASC interacting with blood mononuclear cells. IL-17A and pro-inflammatory cytokine secretion were measured by ELISA. Receptor of AGE (RAGE) together with intercellular adhesion molecule 1 (ICAM-1), human leukocyte Antigen (HLA)-DR, cluster of differentiation (CD) 41, and CD62P surface expressions were measured by cytofluorometry. Anti-RAGE specific monoclonal antibody was added to co-cultures in order to evaluate the role of RAGE in IL-17A production.RESULTSResults showed that whereas 1% G-HSA only weakly potentiated the production of IL-17A by T cells interacting with ASC harvested from obese subjects, it markedly increased IL-17A, but also interferon gamma and tumor necrosis factor alpha production in the presence of ASC harvested from lean individuals. This was associated with increased expression of RAGE and HLA-DR molecule by co-cultured cells. Moreover, RAGE blockade experiments demonstrated RAGE specific involvement in lean ASC-mediated Th-17 cell activation. Finally, platelet aggregation and ICAM-1, which are known to be induced by AGE, were not involved in these processes.CONCLUSIONThus, our results demonstrated that G-HSA potentiated lean ASC-mediated IL-17A production in AT, suggesting a new mechanism by which AGE could contribute to T1D pathophysiology.

Vitamin D attenuates human gingival fibroblast inflammatory cytokine production following advanced glycation end product interaction with receptors for AGE

Vitamin D [1,25(OH)2 D3 or 1,25D3] is critical in musculoskeletal health, inflammation, immune response, and glucose metabolism. Patients with vitamin D deficiency may be at higher risk of diabetes and periodontitis. Diabetic patients exhibit exacerbated inflammation and more periodontal destruction. Advanced glycation end products (AGEs), formed during diabetic hyperglycemia, activate inflammatory pathways in periodontitis. Human gingival fibroblasts (HGFs) express receptors for AGEs (RAGEs) and can contribute to inflammation. Determine whether glycated human serum albumin (G-HSA) augments HGF IL-6 and IL-8 production, and whether treatment with 1,25D3 attenuates cytokine production following stimulation with G-HSA+IL-1β and/or IL-17. HGFs were incubated ±G-HSA or normal human serum albumin (HSA), ±IL-1β and/or IL-17, ±1,25D3. Cytokines were measured by ELISA. Neutralizing anti-RAGE was used to assess AGE-RAGE interaction. Endotoxin was measured using the ToxinSensor™ System. Data were expressed as mean±standard deviation and analyzed using a one-way analysis of variance (ANOVA) and Scheffe's F procedure for post hoc comparisons. G-HSA or IL-1β, but not HSA, significantly stimulated IL-6 and IL-8 production. G-HSA or HSA when combined with IL-1β or IL-1β+IL-17 synergistically stimulated IL-6 and IL-8. Neutralizing anti-RAGE inhibited IL-6 and IL-8 produced by cells stimulated with IL-1β+G-HSA but not (+HSA). Synergism caused by HSA did not appear to be mediated by endotoxin since its levels in G-HSA and HSA were not sufficient to stimulate fibroblasts. Vitamin D inhibited IL-6 and IL-8 production stimulated by G-HSA or HSA + IL-1β or IL-1β + IL-17. Results suggest that the "perioprotective" effects of vitamin D are related to its ability to regulate inflammatory cytokine production by HGFs following AGE-RAGE interaction.

Glycation alter serum albumin binding of valsartan and nateglinide when studied contemporarily

ABSTRACTIn this research work, an attempt was made to study alteration in glycated serum albumin binding of valsartan and nateglinide using validated HPLC-UV method and ultrafiltration as in vitro protein binding study model. The chromatographic conditions involved stationary phase Kromasil-100 C18 (100 × 4.6 mm, 3.5 µm) with mobile phase of 10 mM phosphate buffer, acetonitrile, isopropyl alcohol in the ratio of 30:65:5 as isocratic mode at a flow rate of 0.8 mL/min; and the eluent was monitored at 218 nm. Protein precipitation technique was used to extract the drugs from human plasma. The calibration curve was found linear in the range from 50 to 5000 ng/mL. Glycation of human serum albumin was achieved at different concentration levels using D-(+)-glucose and glycated human serum albumin (Gly-HSA) were prepared. Valsartan and nateglinide were not affected the plasma protein binding of each other when studied using HSA. The unbound fraction of valsartan and nateglinide was increased to 10–20 times when spiked with Gly-HSA. About 20% increase in unbound fraction of valsartan was observed when spiked with 10 µg/mL of nateglinide. Furthermore, the unbound fraction of nateglinide was increased nearly to 10% more when incubated with Gly-HSA as compare to recombinant human serum albumin.