Liposome-encapsulated mannose-1-phosphate therapy improves global N-glycosylation in different congenital disorders of glycosylation.

Phosphomannomutase 2 (PMM2) converts mannose-6-phospahate to mannose-1-phosphate; the substrate for GDP-mannose, a building block of the glycosylation biosynthetic pathway. Pathogenic variants in the PMM2 gene have been shown to be associated with protein hypoglycosylation causing PMM2-congenital disorder of glycosylation (PMM2-CDG). While mannose supplementation improves glycosylation in vitro, but not in vivo, we hypothesized that liposomal delivery of mannose-1-phosphate could increase the stability and delivery of the activated sugar to enter the targeted compartments of cells. Thus, we studied the effect of liposome-encapsulated mannose-1-P (GLM101) on global protein glycosylation and on the cellular proteome in skin fibroblasts from individuals with PMM2-CDG, as well as in individuals with two N-glycosylation defects early in the pathway, namely ALG2-CDG and ALG11-CDG. We leveraged multiplexed proteomics and N-glycoproteomics in fibroblasts derived from different individuals with various pathogenic variants in PMM2, ALG2 and ALG11 genes. Proteomics data revealed a moderate but significant change in the abundance of some of the proteins in all CDG fibroblasts upon GLM101 treatment. On the other hand, N-glycoproteomics revealed the GLM101 treatment enhanced the expression levels of several high-mannose and complex/hybrid glycopeptides from numerous cellular proteins in individuals with defects in PMM2 and ALG2 genes. Both PMM2-CDG and ALG2-CDG exhibited several-fold increase in glycopeptides bearing Man6 and higher glycans and a decrease in Man5 and smaller glycan moieties, suggesting that GLM101 helps in the formation of mature glycoforms. These changes in protein glycosylation were observed in all individuals irrespective of their genetic variants. ALG11-CDG fibroblasts also showed increase in high mannose glycopeptides upon treatment; however, the improvement was not as dramatic as the other two CDG. Overall, our findings suggest that treatment with GLM101 overcomes the genetic block in the glycosylation pathway and can be used as a potential therapy for CDG with enzymatic defects in early steps in protein N-glycosylation.

D-mannose as a new therapy for fucokinase deficiency-related congenital disorder of glycosylation (FCSK-CDG).

INTRODUCTION: Fucokinase deficiency-related congenital disorder of glycosylation (FCSK-CDG) is a rare autosomal recessive inborn error of metabolism characterized by a decreased flux through the salvage pathway of GDP-fucose biosynthesis due to a block in the recycling of L-fucose that exits the lysosome. FCSK-CDG has been described in 5 individuals to date in the medical literature, with a phenotype comprising global developmental delays/intellectual disability, hypotonia, abnormal myelination, posterior ocular disease, growth and feeding failure, immune deficiency, and chronic diarrhea, without clear therapeutic recommendations. PATIENT AND METHODS: In a so far unreported FCSK-CDG patient, we studied proteomics and glycoproteomics in vitro in patient-derived fibroblasts and also performed in vivo glycomics, before and after treatment with either D-Mannose or L-Fucose. RESULTS: We observed a marked increase in fucosylation after D-mannose supplementation in fibroblasts compared to treatment with L-Fucose. The patient was then treated with D-mannose at 850 mg/kg/d, with resolution of the chronic diarrhea, resolution of oral aversion, improved weight gain, and observed developmental gains. Serum N-glycan profiles showed an improvement in the abundance of fucosylated glycans after treatment. No treatment-attributed adverse effects were observed. CONCLUSION: D-mannose is a promising new treatment for FCSK-CDG.

Suppression of N-Glycosylation of Zinc Finger Protein 471 Affects Proliferation, Invasion, and Docetaxel Sensitivity of Tongue Squamous Cell Carcinoma via Regulation of c-Myc.

Zinc finger protein 471 (ZNF471) is a member of the Krüppel-related domain zinc finger protein family, and has recently attracted attention because of its anti-cancer effects. N-glycosylation regulates expression and functions of the protein. This study aimed to investigate the effects of ZNF471 N-glycosylation on the proliferation, invasion, and docetaxel sensitivity of tongue squamous cell carcinoma (TSCC). It analyzed the expression, function, and prognostic significance of ZNF471 in TSCC using bioinformatics techniques such as gene differential expression analysis, univariate Cox regression analysis, functional enrichment analysis, and gene set enrichment analysis. Using site-specific mutagenesis, this study generated three mutant sites for ZNF471 N-glycosylation to determine the effect of N-glycosylation on ZNF471 protein levels and function. Quantitative real-time PCR, Western blot analysis, and immunohistochemistry tests confirmed the down-regulation of ZNF471 expression in TSCC. Low expression of ZNF471 is associated with poor prognosis of patients with TSCC. Overexpression of ZNF471 in vitro retarded the proliferation of TSCC cells and suppressed cell invasion and migration ability. Asparagine 358 was identified as a N-glycosylation site of ZNF471. Suppressing N-glycosylation of ZNF471 enhanced the protein stability and promoted the translocation of protein to the cell nucleus. ZNF471 binding to c-Myc gene promoter suppressed oncogene c-Myc expression, thereby playing the anti-cancer effect and enhancing TSCC sensitivity to docetaxel. In all, N-glycosylation of ZNF471 affects the proliferation, invasion, and docetaxel sensitivity of TSCC via regulation of c-Myc.

Modulatory effects of rutin and vitamin A on hyperglycemia induced glycation, oxidative stress and inflammation in high-fat-fructose diet animal model.

In the current study we investigated the impact of combination of rutin and vitamin A on glycated products, the glyoxalase system, oxidative markers, and inflammation in animals fed a high-fat high-fructose (HFFD) diet. Thirty rats were randomly divided into six groups (n = 5). The treatments, metformin (120 mg/kg), rutin (100 mg/kg), vitamin A (43 IU/kg), and a combination of rutin (100 mg/kg) and vitamin A (43 IU/kg) were given to relevant groups of rats along with high-fructose high-fat diet for 42 days. HbA1c, D-lactate, Glyoxylase-1, Hexokinase 2, malondialdehyde (MDA), glutathione peroxidase (GPx), catalase (CAT), nuclear transcription factor-B (NF-κB), interleukin-6 (IL-6), interleukin-8 (IL-8) and histological examinations were performed after 42 days. The docking simulations were conducted using Auto Dock package. The combined effects of rutin and vitamin A in treated rats significantly (p < 0.001) reduced HbA1c, hexokinase 2, and D-lactate levels while preventing cellular damage. The combination dramatically (p < 0.001) decreased MDA, CAT, and GPx in treated rats and decreased the expression of inflammatory cytokines such as IL-6 andIL-8, as well as the transcription factor NF-κB. The molecular docking investigations revealed that rutin had a strong affinity for several important biomolecules, including as NF-κB, Catalase, MDA, IL-6, hexokinase 2, and GPx. The results propose beneficial impact of rutin and vitamin A as a convincing treatment strategy to treat AGE-related disorders, such as diabetes, autism, alzheimer's, atherosclerosis.

Glucosamine and Silibinin Alter Cartilage Homeostasis through Glycosylation and Cellular Stresses in Human Chondrocyte Cells.

Osteoarthritis is more prevalent than any other form of arthritis and is characterized by the progressive mechanical deterioration of joints. Glucosamine, an amino monosaccharide, has been used for over fifty years as a dietary supplement to alleviate osteoarthritis-related discomfort. Silibinin, extracted from milk thistle, modifies the degree of glycosylation of target proteins, making it an essential component in the treatment of various diseases. In this study, we aimed to investigate the functional roles of glucosamine and silibinin in cartilage homeostasis using the TC28a2 cell line. Western blots showed that glucosamine suppressed the N-glycosylation of the gp130, EGFR, and N-cadherin proteins. Furthermore, both glucosamine and silibinin differentially decreased and increased target proteins such as gp130, Snail, and KLF4 in TC28a2 cells. We observed that both compounds dose-dependently induced the proliferation of TC28a2 cells. Our MitoSOX and DCFH-DA dye data showed that 1 µM glucosamine suppressed mitochondrial reactive oxygen species (ROS) generation and induced cytosol ROS generation, whereas silibinin induced both mitochondrial and cytosol ROS generation in TC28a2 cells. Our JC-1 data showed that glucosamine increased red aggregates, resulting in an increase in the red/green fluorescence intensity ratio, while all the tested silibinin concentrations increased the green monomers, resulting in decreases in the red/green ratio. We observed increasing subG1 and S populations and decreasing G1 and G2/M populations with increasing amounts of glucosamine, while increasing amounts of silibinin led to increases in subG1, S, and G2/M populations and decreases in G1 populations in TC28a2 cells. MTT data showed that both glucosamine and silibinin induced cytotoxicity in TC28a2 cells in a dose-dependent manner. Regarding endoplasmic reticulum stress, both compounds induced the expression of CHOP and increased the level of p-eIF2α/eIF2α. With respect to O-GlcNAcylation status, glucosamine and silibinin both reduced the levels of O-GlcNAc transferase and hypoxia-inducible factor 1 alpha. Furthermore, we examined proteins and mRNAs related to these processes. In summary, our findings demonstrated that these compounds differentially modulated cellular proliferation, mitochondrial and cytosol ROS generation, the mitochondrial membrane potential, the cell cycle profile, and autophagy. Therefore, we conclude that glucosamine and silibinin not only mediate glycosylation modifications but also regulate cellular processes in human chondrocytes.

Potential inhibitory effect of highland barley protein hydrolysates on the formation of advanced glycation end-products (AGEs): A mechanism study.

Advanced glycation end products (AGEs) can be caused during a glycoxidation reaction. This reaction is associated with complications of diabetes and the consequences of health problems. Therefore, we are exploring the prohibitory effect of highland barley protein hydrolysates (HBPHs) on AGE formation. Herein, first extracted the protein from highland barley with various pH conditions and then hydrolyzed using four different proteolytic enzymes (flavourzyme, trypsin, papain, pepsin) under different degrees of hydrolysis. We assessed three degrees of hydrolysates (lowest, middle, highest) of enzymes used to characterize the antioxidant activity and physicochemical properties. Among all the hydrolysates, flavourzyme-treated hydrolysates F-1, F-2, and F-3 indicated the high ability to scavenge DPPH (IC50 values of 0.97 %, 0.63 %, and 0.90 %), structural and functional properties. Finally, the inhibitory effect of the most active hydrolysates F-1, F-2, and F-3 against the AGEs formation was evaluated in multiple glucose-glycated bovine serum albumin (BSA) systems. Additionally, in a BSA system, F-3 exhibited the strong antiglycation activity, effectively suppressed the non-fluorescent AGE (CML), and the fructosamine level. Moreover, it decreased carbonyl compounds while also preventing the loss of thiol groups. Our results would be beneficial in the application of the food industry as a potential antiglycation agent for several chronic diseases.

Protective effects of sugarcane polyphenol against UV-B-induced photoaging in Balb/c mouse skin: Antioxidant, anti-inflammatory, and anti-glycosylation Effects.

Although the benefits of sugarcane polyphenol (SP) are well documented, its function in preventing photoaging has not yet been investigated. This study aimed to investigate the protective effects of SP in preventing ultraviolet (UV)-B-induced skin photoaging in Balb/c mice, as well as the underlying mechanism. Chlorogenic acid was determined to be the primary component of SP by using high-performance liquid chromatography-mass spectrometry. SP and chlorogenic acid were orally administrated to mice for 56 days, and UV-B radiation exposure was administered 14 days after SP and chlorogenic acid administration and lasted 42 days to cause photoaging. SP and chlorogenic acid administrations significantly alleviated the UV-B-induced mouse skin photoaging, as indicated by the decrease in epidermal thickness, increase in the collagen (COL) volume fraction, and elevation in type 1 and type 3 COL contents. Notably, both SP and chlorogenic acid effectively reversed the overexpression of matrix metalloproteinase induced by UV-B exposure in the mouse skin. Furthermore, SP and chlorogenic acid reduced the expression of receptor for advanced glycosylation end products in the mice; amplified the activities of antioxidant enzymes superoxide dismutase and catalase; reduced malondialdehyde levels; and decreased inflammatory cytokines interleukin 1ß, interleukin 6, and tumor necrosis factor α levels. SP could be a prospective dietary supplement for anti-photoaging applications due to its antioxidant, anti-inflammatory, and anti-glycosylation attributes, and chlorogenic acid might play a major role in these effects. PRACTICAL APPLICATION: This study can provide a scientific basis for the practical application of sugarcane polyphenols. We expect that sugarcane polyphenols can be used in food and beverage products to provide flavor while combating skin aging.

Fatty acids characterisation by GC-MS, antiglycation effect at multiple stages and protection of erythrocytes cells from oxidative damage induced by glycation of albumin of Opuntia ficus-indica (L.) Mill seed oil cultivated in Eastern Morocco: Experimental and computational approaches.

ETHNOPHARMACOLOGICAL RELEVANCE: Opuntia ficus-indica (L.) Mill is frequently observed in the Moroccan traditional medicinal system, where these approaches are employed to mitigate the onset of diabetes and the subsequent complications it may entail. AIM OF THE STUDY: The aim of this research was to examine the effectiveness of Opuntia ficus-indica seed oil in preventing diabetic complications. Specifically, the study assessed its ability to counteract glycation at various stages, protected red blood cells from the harmful effects of glycated albumin, and inhibited pancreatic lipase digestive enzymes to understand its potential antihyperglycemic properties. Additionally, the study aimed to identify the chemical components responsible for these effects, evaluate antioxidant and anti-inflammatory properties, and conduct computational investigations such as molecular docking. MATERIALS AND METHODS: The assessement of Opuntia ficus-indica seed oil antiglycation properties involved co-incubating the extract oil with a bovine serum albumin-glucose glycation model. The study investigated various stages of glycation, incorporating fructosamine (inceptive stage), protein carbonyls (intermediate stage), and AGEs (late stage). Additionally, measurement of ß-amyloid aggregation of albumin was performed using Congo red, which is specific to amyloid structures. Additionally, the evaluation of oil's safeguarding effect on erythrocytes against toxicity induced by glycated albumin included the measurement of erythrocyte hemolysis, lipid peroxidation, reduced glutathione. The fatty acid of Opuntia ficus-indica seed oil were analyzed using Gas Chromatography-Mass Spectrometry (GC-MS). The in vitro evaluation of antihyperglycemic activity involved the use of pancreatic lipase enzyme, while the assessement of antioxidant capability was carried out through the utilization of the ABTS and FRAP methods. The in vitro assessement of the denaturation of albumin activity was also conducted. In conjunction with the experimental outcomes, computational investigations were undertaken, specifically employing ADMET (absorption, distribution, metabolism, excretion, and toxicity) analysis. Furthermore, molecular docking was utilized to predict antioxidant and antiglycation mechanisms based on protein targets. RESULTS: In vitro glycation assays, Opuntia ficus-indica seed oil displayed targeted inhibitory effects at multiple distinct stages. Within erythrocytes, in addition to mitigating hemolysis and lipid peroxidation induced by glycated albumin. GC-MS investigation revealed a richness of fatty acids and the most abundant compounds are Linoleic acid (36.59%), Palmitic acid (20.84%) and Oleic acid (19.33%) respectively. The findings of antioxidant ability showed a remarkable activity on FRAP and ABTS radicals. This oil showed a pronounced inhibitory impact (p < 0.001) on pancreatic lipase enzyme. It also exerted a notibale inhibition of albumin denaturation, in vitro. CONCLUSION: The identified results were supported by the abundant compounds of fatty acids unveiled through GC-MS analysis, along with the computational investigation and molecular docking.

Exploring the composition and properties of Centella asiatica metabolites and investigating their impact on BSA glycation, LDL oxidation and α-amylase inhibition.

Centella asiatica (L.) Urb. is a small herbaceous plant belonging to the Apiaceae family that is rich in triterpenes, such as asiaticoside and madecassoside. Centella asiatica finds broad application in promoting wound healing, addressing skin disorders, and boosting both memory and cognitive function. Given its extensive therapeutic potential, this study aimed not only to investigate the Centella asiatica ethanolic extract but also to analyze the biological properties of its organic fractions, such as antioxidant antiglycation capacity, which are little explored. We also identified the main bioactive compounds through spectrometry analysis. The ethanolic extract (EE) was obtained through a static maceration for seven days, while organic fractions (HF: hexane fraction; DF: dichloromethane fraction; EAF: ethyl acetate fraction; BF: n-butanol fraction and HMF: hydromethanolic fraction) were obtained via liquid-liquid fractionation. The concentration of phenolic compounds, flavonoids, and tannins in each sample was quantified. Additionally, the antiglycation (BSA/FRU, BSA/MGO, and ARG/MGO models) and antioxidant (FRAP, ORAC, and DPPH) properties, as well as the ability to inhibit LDL oxidation and hepatic tissue peroxidation were evaluated. The inhibition of enzyme activity was also analyzed (α-amylase, α-glycosidase, acetylcholinesterase, and butyrylcholinesterase). We also evaluated the antimicrobial and cytotoxicity against RAW 264.7 macrophages. The main compounds present in the most bioactive fractions were elucidated through ESI FT-ICR MS and HPLC-ESI-MS/MS analysis. In the assessment of antioxidant capacity (FRAP, ORAC, and DPPH), the EAF and BF fractions exhibited notable results, and as they are the phenolic compounds richest fractions, they also inhibited LDL oxidation, protected the hepatic tissue from peroxidation and inhibited α-amylase activity. Regarding glycation models, the EE, EAF, BF, and HMF fractions demonstrated substantial activity in the BSA/FRU model. However, BF was the only fraction that presented non-cytotoxic activity in RAW 264.7 macrophages at all tested concentrations. In conclusion, this study provides valuable insights into the antioxidant, antiglycation, and enzymatic inhibition capacities of the ethanolic extract and organic fractions of Centella asiatica. The findings suggest that further in vivo studies, particularly focusing on the butanol fraction (BF), may be promising routes for future research and potential therapeutic applications.

Elucidation of inhibitory effects of bioactive anthraquinones towards formation of DNA advanced glycation end products (DNA-AGEs).

DNA is essential in biological processes as it directs transcription and translation assisting in RNA and protein synthesis. Extended periods of elevated blood glucose levels cause non-enzymatic DNA glycation, which results in the formation of DNA-AGEs and the production of free radicals, causing structural perturbation of DNA. In this work, we have investigated the glycation of calf thymus (ct-DNA) DNA and examined its inhibition by two anthraquinone derivatives, purpurin and aloin. Ribose sugar served as the glycating agent inducing non-enzymatic glycation of DNA and subsequent DNA-AGEs formation. UV-vis and fluorescence spectroscopic methods were utilized to characterize DNA-AGE formation in vitro. Circular dichroism (CD) spectroscopy was used to observe the structural disruption of DNA caused by glycation. The changes in AGEs fluorescence intensity and melting temperature (Tm) were measured to assess the inhibition of glycation process by aloin and purpurin. These derivatives demonstrated inhibitory effects via binding to glycating sites of ct-DNA or by scavenging free radicals generated during glycation. The current study elucidates the inhibitory actions of aloin and purpurin on DNA glycation, suggesting their possible applications in mitigating the adverse consequences linked to increased ribose concentrations.

The antiglycation potential of H1 receptor antagonists - in vitro studies in bovine serum albumin model and in silico molecular docking analyses.

The H1 receptor belongs to the family of rhodopsin-like G-protein-coupled receptors activated by the biogenic amine histamine. H1 receptor antagonists are widely used in the treatment of allergies. However, these drugs could have a much broader spectrum of activity, including hypoglycemic effects, which can broaden the spectrum of their use. The aim of the study was to evaluate the antiglycation potential of twelve H1 receptor antagonists (diphenhydramine, antazoline, promethazine, ketotifen, clemastine, pheniramine, cetirizine, levocetirizine, bilastine, fexofenadine, desloratadine, and loratadine). Bovine serum albumin (BSA) was glycated with sugars (glucose, fructose, galactose, and ribose) and aldehydes (glyoxal and methylglyoxal) in the presence of H1 blockers. The tested substances did not induce a significant decrease in the content of albumin glycation end-products, and the inhibition rate of glycoxidation was not influenced by the chemical structure or generation of H1 blockers. None of the tested H1 receptor antagonists exhibited strong antiglycation activity. Antiglycemic potential of H1 blockers could be attributed to their antioxidant and anti-inflammatory activity, as well as their effects on carbohydrate metabolism/metabolic balance at the systemic level.

Emerging Glycation-Based Therapeutics-Glyoxalase 1 Inducers and Glyoxalase 1 Inhibitors.

The abnormal accumulation of methylglyoxal (MG) leading to increased glycation of protein and DNA has emerged as an important metabolic stress, dicarbonyl stress, linked to aging, and disease. Increased MG glycation produces inactivation and misfolding of proteins, cell dysfunction, activation of the unfolded protein response, and related low-grade inflammation. Glycation of DNA and the spliceosome contribute to an antiproliferative and apoptotic response of high, cytotoxic levels of MG. Glyoxalase 1 (Glo1) of the glyoxalase system has a major role in the metabolism of MG. Small molecule inducers of Glo1, Glo1 inducers, have been developed to alleviate dicarbonyl stress as a prospective treatment for the prevention and early-stage reversal of type 2 diabetes and prevention of vascular complications of diabetes. The first clinical trial with the Glo1 inducer, trans-resveratrol and hesperetin combination (tRES-HESP)-a randomized, double-blind, placebo-controlled crossover phase 2A study for correction of insulin resistance in overweight and obese subjects, was completed successfully. tRES-HESP corrected insulin resistance, improved dysglycemia, and low-grade inflammation. Cell permeable Glo1 inhibitor prodrugs have been developed to induce severe dicarbonyl stress as a prospective treatment for cancer-particularly for high Glo1 expressing-related multidrug-resistant tumors. The prototype Glo1 inhibitor is prodrug S-p-bromobenzylglutathione cyclopentyl diester (BBGD). It has antitumor activity in vitro and in tumor-bearing mice in vivo. In the National Cancer Institute human tumor cell line screen, BBGD was most active against the glioblastoma SNB-19 cell line. Recently, potent antitumor activity was found in glioblastoma multiforme tumor-bearing mice. High Glo1 expression is a negative survival factor in chemotherapy of breast cancer where adjunct therapy with a Glo1 inhibitor may improve treatment outcomes. BBGD has not yet been evaluated clinically. Glycation by MG now appears to be a pathogenic process that may be pharmacologically manipulated for therapeutic outcomes of potentially important clinical impact.

2-Deoxy-d-glucose induces deglycosylation of proinflammatory cytokine receptors and strongly reduces immunological responses in mouse models of inflammation.

Anti-proinflammatory cytokine therapies against interleukin (IL)-6, tumor necrosis factor (TNF)-α, and IL-1 are major advancements in treating inflammatory diseases, especially rheumatoid arthritis. Such therapies are mainly performed by injection of antibodies against cytokines or cytokine receptors. We initially found that the glycolytic inhibitor 2-deoxy-d-glucose (2-DG), a simple monosaccharide, attenuated cellular responses to IL-6 by inhibiting N-linked glycosylation of the IL-6 receptor gp130. Aglycoforms of gp130 did not bind to IL-6 or activate downstream intracellular signals that included Janus kinases. 2-DG completely inhibited dextran sodium sulfate-induced colitis, a mouse model for inflammatory bowel disease, and alleviated laminarin-induced arthritis in the SKG mouse, an experimental model for human rheumatoid arthritis. These diseases have been shown to be partially dependent on IL-6. We also found that 2-DG inhibited signals for other proinflammatory cytokines such as TNF-α, IL-1ß, and interferon -γ, and accordingly, prevented death by another inflammatory disease, lipopolysaccharide (LPS) shock. Furthermore, 2-DG prevented LPS shock, a model for a cytokine storm, and LPS-induced pulmonary inflammation, a model for acute respiratory distress syndrome of coronavirus disease 2019 (COVID-19). These results suggest that targeted therapies that inhibit cytokine receptor glycosylation are effective for treatment of various inflammatory diseases.

Host glycocalyx captures HIV proximal to the cell surface via oligomannose-GlcNAc glycan-glycan interactions to support viral entry.

Here, we present ultrastructural analyses showing that incoming HIV are captured near the lymphocyte surface in a virion-glycan-dependent manner. Biophysical analyses show that removal of either virion- or cell-associated N-glycans impairs virus-cell binding, and a similar glycan-dependent relationship is observed between purified HIV envelope (Env) and primary T cells. Trimming of N-glycans from either HIV or Env does not inhibit protein-protein interactions. Glycan arrays reveal HIV preferentially binds to N-acetylglucosamine and mannose. Interfering with these glycan-based interactions reduces HIV infectivity. These glycan interactions are distinct from previously reported glycan-lectin and non-specific electrostatic charge-based interactions. Specific glycan-glycan-mediated attachment occurs prior to virus entry and enhances efficiency of infection. Binding and fluorescent imaging data support glycan-glycan interactions as being responsible, at least in part, for initiating contact between HIV and the host cell, prior to viral Env-cellular CD4 engagement.

Human Sensory Neuron-like Cells and Glycated Collagen Matrix as a Model for the Screening of Analgesic Compounds.

Increased collagen-derived advanced glycation end-products (AGEs) are consistently related to painful diseases, including osteoarthritis, diabetic neuropathy, and neurodegenerative disorders. We have recently developed a model combining a two-dimensional glycated extracellular matrix (ECM-GC) and primary dorsal root ganglion (DRG) that mimicked a pro-nociceptive microenvironment. However, culturing primary cells is still a challenge for large-scale screening studies. Here, we characterized a new model using ECM-GC as a stimulus for human sensory-like neurons differentiated from SH-SY5Y cell lines to screen for analgesic compounds. First, we confirmed that the differentiation process induces the expression of neuron markers (MAP2, RBFOX3 (NeuN), and TUBB3 (ß-III tubulin), as well as sensory neuron markers critical for pain sensation (TRPV1, SCN9A (Nav1.7), SCN10A (Nav1.8), and SCN11A (Nav1.9). Next, we showed that ECM-GC increased c-Fos expression in human sensory-like neurons, which is suggestive of neuronal activation. In addition, ECM-GC upregulated the expression of critical genes involved in pain, including SCN9A and TACR1. Of interest, ECM-GC induced substance P release, a neuropeptide widely involved in neuroinflammation and pain. Finally, morphine, the prototype opiate, decreased ECM-GC-induced substance P release. Together, our results suggest that we established a functional model that can be useful as a platform for screening candidates for the management of painful conditions.

Targeting PGM3 as a Novel Therapeutic Strategy in KRAS/LKB1 Co-Mutant Lung Cancer.

In non-small-cell lung cancer (NSCLC), concurrent mutations in the oncogene KRAS and tumor suppressor STK11 (also known as LKB1) confer an aggressive malignant phenotype, an unfavourability towards immunotherapy, and overall poor prognoses in patients. In a previous study, we showed that murine KRAS/LKB1 co-mutant tumors and human co-mutant cancer cells have an enhanced dependence on glutamine-fructose-6-phosphate transaminase 2 (GFPT2), a rate-limiting enzyme in the hexosamine biosynthesis pathway (HBP), which could be targeted to reduce survival of KRAS/LKB1 co-mutants. Here, we found that KRAS/LKB1 co-mutant cells also exhibit an increased dependence on N-acetylglucosamine-phosphate mutase 3 (PGM3), an enzyme downstream of GFPT2. Genetic or pharmacologic suppression of PGM3 reduced KRAS/LKB1 co-mutant tumor growth in both in vitro and in vivo settings. Our results define an additional metabolic vulnerability in KRAS/LKB1 co-mutant tumors to the HBP and provide a rationale for targeting PGM3 in this aggressive subtype of NSCLC.

GALNT14 regulates ferroptosis and apoptosis of ovarian cancer through the EGFR/mTOR pathway.

Background: Chemoresistance usually occurs in ovarian cancer. We aimed to explore the mechanisms of chemoresistance. Methods: Western blotting assay was used to detect the expression of GALNT14. Further cell function experiments were performed to investigate the effect of GALNT14 in ovarian cancer. Results: GALNT14 is significantly upregulated in ovarian cancer. Downregulation of GALNT14 significantly inhibits both apoptosis and ferroptosis of ovarian cancer cells. A further mechanism assay illustrated that downregulation of GALNT14 suppresses the activity of the mTOR pathway through modifying O-glycosylation of EGFR. Finally, an additive effect promoting cell death occurs with a combination of an mTOR inhibitor and cisplatin. Conclusion: Our study might provide a promising method to overcome cisplatin resistance for patients with ovarian cancer.

Endoplasmic reticulum stress promotes HBV production by enhancing use of the autophagosome/multivesicular body axis.

BACKGROUND AND AIMS: HBV infection has been reported to trigger endoplasmic reticulum (ER) stress and initiate autophagy. However, how ER stress and autophagy influence HBV production remains elusive. Here, we studied the effect of tunicamycin (TM), an N-glycosylation inhibitor and ER stress inducer, on HBV replication and secretion and examined the underlying mechanisms. APPROACH AND RESULTS: Protein disulfide isomerase (an ER marker), microtubule-associated protein 1 light chain 3 beta (an autophagosome [AP] marker), and sequestosome-1 (a typical cargo for autophagic degradation) expression were tested in liver tissues of patients with chronic HBV infection and hepatoma cell lines. The role of TM treatment in HBV production and trafficking was examined in hepatoma cell lines. TM treatment that mimics HBV infection triggered ER stress and increased AP formation, resulting in enhanced HBV replication and secretion of subviral particles (SVPs) and naked capsids. Additionally, TM reduced the number of early endosomes and HBsAg localization in this compartment, causing HBsAg/SVPs to accumulate in the ER. Thus, TM-induced AP formation serves as an alternative pathway for HBsAg/SVP trafficking. Importantly, TM inhibited AP-lysosome fusion, accompanied by enhanced AP/late endosome (LE)/multivesicular body fusion, to release HBsAg/SVPs through, or along with, exosome release. Notably, TM treatment inhibited HBsAg glycosylation, resulting in impairment of HBV virions' envelopment and secretion, but it was not critical for HBsAg/SVP trafficking in our cell systems. CONCLUSIONS: TM-induced ER stress and autophagic flux promoted HBV replication and the release of SVPs and naked capsids through the AP-LE/MVB axis.

Inhibition of Advanced Glycation End Product Formation in Rat Tail Tendons by Polydatin and p-Coumaric acid: an In Vitro Study.

Advanced glycation end products (AGEs) formed through non-enzymatic glycosylation between a protein and sugar molecule are highly harmful to the human body. In hyperglycemic patients, AGE formation is more due to high glucose circulating in the blood, causing inter and intra molecular cross-linking of collagen leading to reduction of collagen elasticity. This cross-linked collagen develops resistance to matrix metalloproteinases leading to impaired collagen turnover. The aim of this work is to determine the anti-glycation effects of polydatin and p-coumaric acid in preventing collagen cross-linking by incubating rat tail tendons (RTTs) as collagen source in high glucose concentration (50 mM) for a week. The RTTs were then characterized for tensile strength, cross-linking efficiency, circular dichroism spectrometry, collagen, glucose, and aldehyde contents. Electrophoresis was carried out to evaluate the level of cross-linking in collagen and the results confirmed the ability of the drugs in preventing complex intermolecular cross-link formation induced by non-enzymatic glycosylation. CD data showed alteration in the secondary structure of collagen where AGE formation had occurred. More collagen was extracted by pepsin from RTTs treated with glucose alone (6.88 mg/10 mg tendon) when compared with drug-treated groups (4.25, 2.56 mg/10 mg tendon for polydatin and p-coumaric acid, respectively). Tensile strength (20.66% and 18.95%), cross-linking percentage (32.5% and 29.84%), and glucose content (2.3 and 1.8 mg/100 mg) of drug-treated groups were similar to the positive control (19.07%, 30.13%, and 2.61 mg/100 mg) thus proving the anti-glycation potential of the drugs. Hence, both polydatin and p-coumaric acid could play a pivotal role in preventing AGE formation.

A review on prevention of glycation of proteins: Potential therapeutic substances to mitigate the severity of diabetes complications.

Non-enzymatic reaction involving carbonyl of reducing sugars and amino groups in proteins produces advanced glycation end products (AGEs). AGE accumulation in vivo is a crucial factor in the progression of metabolic and pathophysiological mechanisms like obesity, diabetes, coronary artery disease, neurological disorders, and chronic renal failure. The body's own defense mechanism, synthetic inhibitors, and natural inhibitors can all help to prevent the glycation of proteins. Synthetic inhibitors have the potential to suppress the glycation of proteins through a variety of pathways. They could avoid Amadori product development by tampering with the addition of sugars to the proteins. Besides which, the free radical scavenging and blocking crosslink formation could be another mechanism behind their anti-glycation properties. In comparison with synthetic substances, naturally occurring plant products have been found to be comparatively non-toxic, cheap, and usable in an ingestible form. This review gives a brief introduction of the Maillard reaction; formation, characterization and pathology related to AGEs, potential therapeutic approaches against glycation, natural and synthetic inhibitors of glycation and their probable mechanism of action. The scientific community could get benefit from the combined knowledge about important molecules, which will further guide to the design and development of new pharmaceutical compounds.