Advanced Glycation End Products Effects on Adipocyte Niche Stiffness and Cell Signaling.

Adipose tissue metabolism under hyperglycemia results in Type II diabetes (T2D). To better understand how the adipocytes function, we used a cell culture that was exposed to glycation by adding intermediate carbonyl products, which caused chemical cross-linking and led to the formation of advanced glycation end products (AGEs). The AGEs increased the cells and their niche stiffness and altered the rheological viscoelastic properties of the cultured cells leading to altered cell signaling. The AGEs formed concomitant with changes in protein structure, quantified by spectroscopy using the 8-ANS and Nile red probes. The AGE effects on adipocyte differentiation were viewed by imaging and evidenced in a reduction in cellular motility and membrane dynamics. Importantly, the alteration led to reduced adipogenesis, that is also measured by qPCR for expression of adipogenic genes and cell signaling. The evidence of alteration in the plasma membrane (PM) dynamics (measured by CTxB binding and NP endocytosis), also led to the impairment of signal transduction and a decrease in AKT phosphorylation, which hindered downstream insulin signaling. The study, therefore, presents a new interpretation of how AGEs affect the cell niche, PM stiffness, and cell signaling leading to an impairment of insulin signaling.

Nutrition Alters the Stiffness of Adipose Tissue and Cell Signaling.

Adipose tissue is a complex organ composed of various cell types and an extracellular matrix (ECM). The visceral adipose tissue (VAT) is dynamically altered in response to nutritional regimens that lead to local cues affecting the cells and ECM. The adipocytes are in conjunction with the surrounding ECM that maintains the tissue's niche, provides a scaffold for cells and modulates their signaling. In this study, we provide a better understanding of the crosstalk between nutritional regimens and the ECM's stiffness. Histological analyses showed that the adipocytes in mice fed a high-fat diet (HFD) were increased in size, while the ECM was also altered with changes in mass and composition. HFD-fed mice exhibited a decrease in elastin and an increase in collagenous proteins. Rheometer measurements revealed a stiffer ECM in whole tissue (nECM) and decellularized (deECM) in HFD-fed animals. These alterations in the ECM regulate cellular activity and influence their metabolic function. HFD-fed mice expressed high levels of the receptor for advanced-glycation-end-products (RAGE), indicating that AGEs might play a role in these processes. The cells also exhibited an increase in phosphoserine332 of IRS-1, a decrease in the GLUT4 transporter levels at the cells' membrane, and a consequent reduction in insulin sensitivity. These results show how alterations in the stiffness of ECM proteins can affect the mechanical cues transferred to adipocytes and, thereby, influence the adipocytes' functionality, leading to metabolic disorders.

Revealing Advanced Glycation End Products Associated Structural Changes in Serum Albumin.

Structural alterations in proteins have a significant impact on their function and body physiology. Glycation via nonenzymatic forms of cross-linking leads to proteins' conformational changes, the macromolecule being recognized as a stable fibrillary structure, oligomerization, and becoming advanced glycation end products (AGEs). Protein that undergoes glycation-related modifications, namely, ß-sheet enriched structural changes, are recognized as amyloid. In the current study, we characterized a single protein modified in vitro under physiological conditions to represent a protein glycation model. The glycation altered the helical conformation of serum albumin (SA) and promoted the formation of a ß-sheet enriched with amyloid fibrils detected at multidimensional levels. The nanoscale resolution by spectroscopy in the presence of thioflavin-T (ThT) and 8-anilinonaphthalene-1-sulfonic acid (8-ANS) showed binding of the fibrils formed in the presence of glucose (GLU) and the carbonyl metabolites methylglyoxal (MGO) and glycolaldehyde (GAD). In the presence of MGO and GAD, the SA becomes insoluble aggregates, demonstrated by TEM microscopy and dynamic light scattering (DLS). The protein oligomerization was visualized when separated via SDS gel electrophoresis and mass photometry (MP) assays. Following the glycation, eventually, the material polymerized and became stiffer. The level of stiffness was analyzed by a rheometer that revealed a quick alteration under MGO and GAD. This is the first study to combine multiple spectroscopy assays, imaging, and rheology measurements of SA and to demonstrate a resolution on a nanoscale structural toward better resolution of the conformational changes of glycated SA, oligomerization, and protein aggregations under physiological conditions.

Exploring the Cell Stemness and the Complexity of the Adipose Tissue Niche.

Adipose tissue is a complex organ composed of different cellular populations, including mesenchymal stem and progenitor cells, adipocytes, and immune cells such as macrophages and lymphocytes. These cellular populations alter dynamically during aging or as a response to pathophysiology such as obesity. Changes in the various inflammatory cells are associated with metabolic complications and the development of insulin resistance, indicating that immune cells crosstalk with the adipocytes. Therefore, a study of the cell populations in the adipose tissue and the extracellular matrix maintaining the tissue niche is important for the knowledge on the regulatory state of the organ. We used a combination of methods to study various parameters to identify the composition of the resident cells in the adipose tissue and evaluate their profile. We analyzed the tissue structure and cells based on histology, immune fluorescence staining, and flow cytometry of cells present in the tissue in vivo and these markers' expression in vitro. Any shift in cells' composition influences self-renewal of the mesenchymal progenitors, and other cells affect the functionality of adipogenesis.