Label-free visualization of unfolding and crosslinking mediated protein aggregation in nonenzymatically glycated proteins.

Nonenzymatic glycation (NEG) unfolds and crosslinks proteins, resulting in aggregation. Label-free evaluation of such structural changes, without disturbing molecular integrity, would be beneficial for understanding the fundamental mechanisms of protein aggregation. The current study demonstrates the assessment of NEG-induced protein aggregation by combining autofluorescence (AF) spectroscopy and imaging. The methylglyoxal (MG) induced protein unfolding and the formation of cross-linking advanced glycation end-products (AGEs) leading to aggregation were evaluated using deep-UV-induced-autofluorescence (dUV-AF) spectroscopy in proteins with distinct structural characteristics. Since the AGEs formed on proteins are fluorescent, the study demonstrated the possibility of autofluorescence imaging of NEG-induced protein aggregates. Autofluorescence spectroscopy can potentially reveal molecular alterations such as protein unfolding and cross-linking. In contrast, AGE-based autofluorescence imaging offers a means to visually explore the structural arrangement of aggregates, regardless of whether they are amyloid or non-amyloid in nature.

Diagnostic Accuracy of Serum Glycosylated Fibronectin in Prediction of Preeclampsia: A Nested Case-Control Study.

BACKGROUND: Preeclampsia is a life-threatening complication of pregnancy that occurs in approximately 7% of all pregnancies. In India, the incidence of preeclampsia is 8%-10% and the prevalence is 5.4%, whereas the prevalence of hypertensive disorders of pregnancy is 7.8%. AIM AND OBJECTIVES: This study was aimed at evaluating the diagnostic accuracy of serum glycosylated fibronectin (S. GlyFn) in the prediction of preeclampsia. METHODS: A nested case-control study was carried out for 16 months in the department of obstetrics and gynecology. A total of 240 women were recruited and followed after written consent and ethical clearance. Six were lost to follow-up, 15 had second-trimester abortions (excluded from the study), and 32 women developed hypertensive disorders of pregnancy (cases), out of which 1 woman developed antepartum eclampsia, 10 women developed preeclampsia with severe features, and 21 women developed preeclampsia without severe features. One hundred and eighty-seven women remained normotensive throughout the pregnancy until 6 weeks postpartum. After randomization, out of these samples, 54 were analyzed and considered controls. Levels of S. GlyFn were estimated using an ELISA kit using the ELISA technique. RESULTS: The mean S. GlyFn level was significantly higher at the time of enrollment among those women who later developed preeclampsia (127.59 ± 27.68 ng/m) as compared to controls (107.79-53.51 ng/mL). GlyFn at a cutoff value of 126.70 ng/mL significantly (P = 0.034) discriminates cases of preeclampsia with severe features from healthy controls with a sensitivity of 90.00%, a specificity of 63.00%, a 31.03% positive predictive value, and 97.14% negative predictive value. CONCLUSION: S. GlyFn, at a cutoff value of 126.70 ng/mL, had good sensitivity to discriminate PE from normotensive and was also a good prognostic marker.

Résumé Contexte:La prééclampsie est une complication potentiellement mortelle de la grossesse qui survient dans environ 7 % de toutes les grossesses. En Inde, l'incidence de la prééclampsie est de 8 % à 10 % et la prévalence est de 5,4 %, alors que la prévalence des troubles hypertensifs de la grossesse est 7,8 %. But et objectifs : Cette étude visait à évaluer la précision diagnostique de la fibronectine sérique glycosylée (S. GlyFn) chez la prédiction de la prééclampsie.Méthodes:Une étude cas-témoin nichée a été menée pendant 16 mois dans le service d'obstétrique et gynécologie. Au total, 240 femmes ont été recrutées et suivies après consentement écrit et autorisation éthique. Six ont été perdus de vue, 15 avaient avortements au deuxième trimestre (exclus de l'étude), et 32 femmes ont développé des troubles hypertensifs de la grossesse (cas), dont 1 femme a développé une éclampsie antepartum, 10 femmes ont développé une prééclampsie avec des caractéristiques sévères et 21 femmes ont développé une prééclampsie sans traits sévères. Cent quatre-vingt sept femmes sont restées normotendues tout au long de la grossesse jusqu'à 6 semaines après l'accouchement. Après randomisation, sur ces échantillons, 54 ont été analysés et considérés comme témoins. Les niveaux de S. GlyFn ont été estimés à l'aide d'un kit ELISA en utilisant la technique ELISA.Résultats:Le niveau moyen de S. GlyFn était significativement plus élevé au moment de l'inscription chez les femmes qui ont développé plus tard une prééclampsie (127,59 ± 27,68 ng/m) par rapport aux témoins (107,79­53,51 ng/mL). GlyFn à une valeur seuil de 126,70 ng/mL de manière significative (P = 0,034) discrimine les cas de prééclampsie avec des caractéristiques sévères des témoins sains avec une sensibilité de 90,00 %, un spécificité de 63,00 %, une valeur prédictive positive de 31,03 % et une valeur prédictive négative de 97,14 %.Conclusion:S. GlyFn, à une valeur seuil de 126,70 ng/mL, avait une bonne sensibilité pour distinguer l'EP du normotendu et était également un bon marqueur pronostique.

Advanced glycation end product-modified low-density lipoprotein promotes pro-osteogenic reprogramming via RAGE/NF-κB pathway and exaggerates aortic valve calcification in hamsters.

BACKGROUND: Advanced glycation end product-modified low-density lipoprotein (AGE-LDL) is related to inflammation and the development of atherosclerosis. Additionally, it has been demonstrated that receptor for advanced glycation end products (RAGE) has a role in the condition known as calcific aortic valve disease (CAVD). Here, we hypothesized that the AGE-LDL/RAGE axis could also be involved in the pathophysiological mechanism of CAVD. METHODS: Human aortic valve interstitial cells (HAVICs) were stimulated with AGE-LDL following pre-treatment with or without interleukin 37 (IL-37). Low-density lipoprotein receptor deletion (Ldlr-/-) hamsters were randomly allocated to chow diet (CD) group and high carbohydrate and high fat diet (HCHFD) group. RESULTS: AGE-LDL levels were significantly elevated in patients with CAVD and in a hamster model of aortic valve calcification. Our in vitro data further demonstrated that AGE-LDL augmented the expression of intercellular cell adhesion molecule-1 (ICAM-1), interleukin-6 (IL-6) and alkaline phosphatase (ALP) in a dose-dependent manner through NF-κB activation, which was attenuated by nuclear factor kappa-B (NF-κB) inhibitor Bay11-7082. The expression of RAGE was augmented in calcified aortic valves, and knockdown of RAGE in HAVICs attenuated the AGE-LDL-induced inflammatory and osteogenic responses as well as NF-κB activation. IL-37 suppressed inflammatory and osteogenic responses and NF-κB activation in HAVICs. The vivo experiment also demonstrate that supplementation with IL-37 inhibited valvular inflammatory response and thereby suppressed valvular osteogenic activities. CONCLUSIONS: AGE-LDL promoted inflammatory responses and osteogenic differentiation through RAGE/NF-κB pathway in vitro and aortic valve lesions in vivo. IL-37 suppressed the AGE-LDL-induced inflammatory and osteogenic responses in vitro and attenuated aortic valve lesions in a hamster model of CAVD.

Glycated LDL generates reactive species that damage cell components, oxidize hemoglobin and alter surface morphology in human erythrocytes.

Low-density lipoprotein (LDL) transports cholesterol to various tissues via the blood. Glycation of LDL occurs during hyperglycemic condition which is characterised by persistently high blood glucose level. Circulating erythrocytes can come in direct contact with glycated LDL (G-LDL). The objective of this study was to investigate the effect of G-LDL on human erythrocytes, specifically on hemoglobin, intracellular generation of reactive species and the antioxidant defence system. Isolated erythrocytes were incubated with G-LDL (3 and 6 mg/ml) and native LDL (6 mg/ml) at 37 °C for 24 h. Native LDL and G-LDL untreated erythrocytes were similarly incubated at 37 °C and served as control. G-LDL treatment increased hemolysis compared to control and native LDL-treated erythrocytes. Incubation of erythrocytes with G-LDL led to an increase in protein oxidation and lipid peroxidation while greatly decreasing the total sulfhydryl content. It also significantly enhanced hemoglobin oxidation, heme degradation, and the release of free iron moiety. Treatment with G-LDL led to an appreciable increase in the production of reactive oxygen and nitrogen species. The antioxidant power and activities of major antioxidant enzymes were drastically reduced, while critical membrane-bound enzymes were inhibited. The surface morphology of G-LDL-treated erythrocytes was altered leading to the formation of echinocytes. Importantly, treatment of erythrocytes with native LDL did not significantly affect the above-mentioned parameters and values were similar to the corresponding controls. Thus, G-LDL is cytotoxic to human erythrocytes and causes oxidative damage to cell components. This can reduce the oxygen-transporting ability of blood and also result in red cell senescence and anemia.

The inhibitory effects of endophytic metabolites on glycated proteins under non-communicable disease conditions: A review.

Protein glycation in human body is closely linked to the onset/progression of diabetes associated complications. These glycated proteins are commonly known as advanced glycation end products (AGEs). Recent literature has also highlighted the involvement of AGEs in other non-communicable diseases (NCDs) such as cardiovascular, cancer, and Alzheimer's diseases and explored the impact of plant metabolites on AGEs formation. However, the significance of endophytic metabolites against AGEs has recently garnered attention but has not been thoroughly summarized thus far. Therefore, the objective of this review is to provide a comprehensive overview of the importance of endophytic metabolites in combating AGEs under NCDs conditions. Additionally, this review aims to elucidate the processes of AGEs formation, absorption, metabolism, and their harmful effects. Collectively, endophytic metabolites play a crucial role in modulating signaling pathways and enhancing the digestibility properties of gut microbiota (GM) by targeting on AGEs/RAGE (receptor for AGEs) axis. Furthermore, these metabolites exhibit anti-AGEs activities similar to those derived from host plants, but at a lower cost and higher production rate. The use of endophytes as a source of such metabolites offers a risk-free and sustainable approach that holds substantial potential for the treatment and management of NCDs.