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Acidic glycans are essential for the biology of multicellular eukaryotes. To utilize them, microbial life including symbionts and pathogens has evolved polysaccharide lyases (PL) that cleave their 1,4 glycosidic linkages via a β-elimination mechanism. PL family 33 (PL33) enzymes have the unusual ability to target a diverse range of glycosaminoglycans (GAGs), as well as the bacterial polymer, gellan gum. In order to gain more detailed insight into PL33 activities we recombinantly expressed 10 PL33 members derived from all major environments and further elucidated the detailed biochemical and biophysical properties of five, showing that their substrate specificity is conferred by variations in tunnel length and topography. The key amino acids involved in catalysis and substrate interactions were identified, and employing a combination of complementary biochemical, structural, and modeling approaches, we show that the tunnel topography is induced by substrate binding to the glycan. Structural and bioinformatic analyses revealed that these features are conserved across several lyase families as well as in mammalian GAG epimerases. [ABSTRACT FROM AUTHOR]

Quantile regression is an extension of linear regression which estimates a conditional quantile of interest. In this paper, we propose an empirical likelihood-based non-parametric procedure to detect structural changes in the quantile regression models. Further, we have modified the proposed smoothed empirical likelihood-based method using adjusted smoothed empirical likelihood and transformed smoothed empirical likelihood techniques. We have shown that under the null hypothesis, the limiting distribution of the smoothed empirical likelihood ratio test statistic is identical to that of the classical parametric likelihood. Simulations are conducted to investigate the finite sample properties of the proposed methods. Finally, to demonstrate the effectiveness of the proposed method, it is applied to urinary Glycosaminoglycans (GAGs) data to detect structural changes. [ABSTRACT FROM AUTHOR]

Chemical modification of naturally derived glycosaminoglycans (GAGs) expands their potential utility for applications in soft tissue repair and regenerative medicine. Here we report the preparation of a novel crosslinked chondroitin sulfate (~200 to 2000 kilodaltons) that is both soluble in aqueous solution and microfilterable. We refer to these materials as "SuperGAGs." One can further conjugate these materials with diverse capture agents to further modify polymer properties and add new capabilities. A representative material (GLX-100) demonstrated durable restoration of bladder impermeability in a gold standard animal model of Interstitial Cystitis/Bladder Pain Syndrome (IC/BPS). Histologic examination of the animal bladders treated with a GLX-100 SuperGAG conjugated to biotin as a reporter demonstrated that the residence time of GLX-100 is superior to chondroitin sulfate (a product that is currently used for clinical treatment of patients with IC/BPS). As expected, this novel crosslinked GAG biopolymer was restricted to the luminal surface of the bladder wall. In this communication we describe a simple and versatile synthesis of a crosslinked glycosaminoglycan (GAG) biopolymer for soft tissue repair. Chondroitin sulfate (~12 kD) was crosslinked to form a water soluble and microfilterable polymer with approximately 200 to 2000 kD molecular weight. The synthesis presented here allows for control of molecular weight while avoiding formation of an extended block gel. Moreover, the procedure enables further chemical modification of the SuperGAG through the selection of a capture agent. A set of agents have been used, demonstrating the preparation of a family of SuperGAGs with diverse capabilities. We can optimize polymer properties, adjust adherence to various tissues, add reporters, and engage the biochemistry of surrounding tissues with peptides and other bioactives. [ABSTRACT FROM AUTHOR]

Lung cancer still represents a global health problem, being the main type of tumor responsible for cancer deaths. In this context, the tumor microenvironment, and the extracellular matrix (ECM) pose as extremely relevant. Thus, this study aimed to explore the prognostic value of epithelial-to-mesenchymal transition (EMT), Wnt signaling, and ECM proteins expression in patients with non–small-cell lung carcinoma (NSCLC) with clinical stages I-IIIA. For that, we used 120 tissue sections from patients and evaluated the immunohistochemical, immunofluorescence, and transmission electron microscopy (TEM) to each of these markers. We also used in silico analysis to validate our data. We found a strong expression of E-cadherin and β-catenin, which reflects the differential ECM invasion process. Therefore, we also noticed a strong expression of chondroitin sulfate (CS) and collagens III and V. This suggests that, after EMT, the basal membrane (BM) enhanced the motility of invasive cells. EMT proteins were directly associated with WNT5A, and collagens III and V, which suggests that the WNT pathway drives them. On the other hand, heparan sulfate (HS) was associated with WNT3A and SPARC, while WNT1 was associated with CS. Interestingly, the association between WNT1 and Col IV suggested negative feedback of WNT1 along the BM. In our cohort, WNT3A, WNT5A, heparan sulfate and SPARC played an important role in the Cox regression model, influencing the overall survival (OS) of patients, be it directly or indirectly, with the SPARC expression stratifying the OS into two groups: 97 months for high expression; and 65 for low expression. In conclusion, the present study identified a set of proteins that may play a significant role in predicting the prognosis of NSCLC patients with clinical stages I-IIIA. [ABSTRACT FROM AUTHOR]

Autism Spectrum Disorder (ASD) is a complex condition with a multifactorial aetiology including both genetic and epigenetic factors. MicroRNAs (miRNAs) play a role in ASD and may influence metabolic pathways. Glycosylation (the glycoconjugate synthesis pathway) is a necessary process for the optimal development of the central nervous system (CNS). Congenital Disorders of Glycosylation (CDGs) (CDGs) are linked to over 180 genes and are predominantly associated with neurodevelopmental disorders (NDDs) including ASD. From a literature search, we considered 64 miRNAs consistently deregulated in ASD patients (ASD-miRNAs). Computational tools, including DIANA-miRPath v3.0 and TarBase v8, were employed to investigate the potential involvement of ASD-miRNAs in glycosylation pathways. A regulatory network constructed through miRNet 2.0 revealed the involvement of these miRNAs in targeting genes linked to glycosylation. Protein functions were further validated through the Human Protein Atlas. A total of twenty-five ASD-miRNAs were identified, including nine miRNAs that were differentially expressed in cells or brain tissue in ASD patients and associated with glycosylation pathways, specifically protein N- and O-glycosylation and glycosaminoglycan biosynthesis (heparan sulfate). A number of CDG genes and/or ASD-risk genes, including DOLK, GALNT2, and EXT1, were identified as targets, along with validated interactions involving four key miRNAs (hsa-miR-423-5p, hsa-miR-30c-5p, hsa-miR-195-5p, and hsa-miR-132-5p). B4GALT1, an ASD susceptibility gene, emerged as a central regulatory hub, reinforcing the link between glycosylation and ASD. In sum, the evidence presented here supports the hypothesis that ASD-miRNAs mediate the epigenetic regulation of glycosylation, thus unveiling possible novel patho-mechanisms underlying ASD. [ABSTRACT FROM AUTHOR]

Cell-penetrating peptides (CPPs) are a diverse group of peptides, typically composed of 4 to 40 amino acids, known for their unique ability to transport a wide range of substances—such as small molecules, plasmid DNA, small interfering RNA, proteins, viruses, and nanoparticles—across cellular membranes while preserving the integrity of the cargo. CPPs exhibit passive and non-selective behavior, often requiring functionalization or chemical modification to enhance their specificity and efficacy. The precise mechanisms governing the cellular uptake of CPPs remain ambiguous; however, electrostatic interactions between positively charged amino acids and negatively charged glycosaminoglycans on the membrane, particularly heparan sulfate proteoglycans, are considered the initial crucial step for CPP uptake. Clinical trials have highlighted the potential of CPPs in diagnosing and treating various diseases, including cancer, central nervous system disorders, eye disorders, and diabetes. This review provides a comprehensive overview of CPP classifications, potential applications, transduction mechanisms, and the most relevant algorithms to improve the accuracy and reliability of predictions in CPP development. [ABSTRACT FROM AUTHOR]

Simple Summary: This review explores how radiotherapy disrupts critical metabolic pathways in cancer cells. It highlights vulnerabilities in glucose metabolism, lipid synthesis, and amino acid pathways, which radiotherapy exploits to enhance tumour control. Advancements such as nanoparticle-enhanced and FLASH radiotherapy are discussed for their ability to improve precision and reduce damage to healthy tissues. Tools like FDG-PET scans are emphasised for detecting metabolic changes and guiding therapy combinations, such as precision radiotherapy with metabolic inhibitors. This review also outlines future prospects, including personalised metabolic imaging, integration with systemic therapies, targeting tumour-specific metabolic dependencies, and innovations like immune modulation and ferroptosis induction. These approaches aim to transform radiotherapy from a localised treatment to a systemic strategy that targets cancer's metabolic complexities, improving both efficacy and patient outcomes. Radiotherapy has traditionally been viewed as a localised treatment modality, primarily inducing DNA damage to eradicate cancer cells. However, emerging research reveals that radiation also profoundly affects cellular metabolism, particularly in reprogramming tumour cells' energy production and survival pathways. This article explores the role of metabolic reprogramming in the response of cancer cells to radiotherapy, highlighting how these biological processes can be leveraged to enhance treatment efficacy and overcome resistance. Radiotherapy is also discussed, focusing on how metabolic and molecular profiling of tumours can help personalise radiation treatments to improve outcomes. [ABSTRACT FROM AUTHOR]

Context: Heparan sulfate (HS) linear polysaccharide glycosaminoglycan compound is linked to components from the cell surface and the extracellular matrix. HS mediates SARS-CoV-2 infection through spike protein binding to cell surface receptors and is required to bind ACE2, prompting the need for electronic structure and molecular docking evaluation of this core system to exploit this attachment in developing new derivatives. Therefore, we have studied five molecules based on HS using molecular docking and electronic structure analysis. Non-covalent interaction analysis shows hydrogen bonding and van der Waals interactions in the binding to RBD-ACE2 interface and 3CLpro. SDM3 and SDM1 molecules present the lowest gap, including solvent effect under 154.6 kcal/mol, and exhibit the most reactivity behavior in this group, potentially leading to enhanced interaction in docking studies. Methods: Heparan sulfate and four derivatives were optimized using B3LYP functional with two basis sets 6–31 + G(d,p) and def2SVP. Electronic structure was used to explore the main interactions and the reactivity of these molecules, and these optimized structures were used in the molecular docking study against 3CLpro, RBD, and ACE2. [ABSTRACT FROM AUTHOR]

Introduction: Excess weight during pregnancy is a condition that can affect both mother and fetus, through the maternal-fetal interface, which is constituted by the placenta and umbilical cord. The umbilical vein is responsible for transporting oxygen and nutrients to the fetus, and its proper functioning depends on the integrity of its structure. The remodeling of the umbilical vein represents one of the causes of inadequate transport of nutrients to the fetus, being potentially harmful. This study aims to evaluate whether maternal overweight alters the structural characteristics of the umbilical vein. Methods: Umbilical cords were collected from eutrophic and overweight pregnant women and were processed according to histological routine. We analyzed morphometry parameters, collagen and elastin fibers deposition, glycosaminoglycan level, and cell proliferation. Results: Veins from overweight pregnant women were found to have greater total area, wall area, wall thickness, and diameter. There was higher collagen labeling in the perivascular region of the overweight group and a higher amount of type III collagen in the vascular smooth muscle. The proliferation of muscle and perivascular cells was higher in overweight pregnant women. A positive, although weak, correlation was observed between BMI and vessel thickness and with type III collagen deposition in vascular smooth muscle. Discussion: With this study, we show that being overweight can structurally alter the umbilical vein, causing vascular remodeling of the vessel, through hypertrophy and hyperplasia. [ABSTRACT FROM AUTHOR]

Background. The COVID-19 epidemic and the war in Ukraine have led to a significant increase in the number of children suffering from metabolic dysfunction-associated steatotic liver disease (MASLD). One of the unresolved problems associated with MASLD is the identification of individuals at risk of rapid disease progression and development of irreversible liver changes. The search for alternative noninvasive markers suitable for the early detection of liver fibrosis in children remains extremely relevant. The aim of the study was to determine the diagnostic value of serum fibrosis markers and their relationship with sonographic and body composition parameters in children with MASLD. Materials and methods. The case-control study included 80 children aged 6 to 17 years (mean of (12.15 ± 2.51) years). The presence of steatosis and liver fibrosis was determined by transient elastography (Fibro­Scan® 502 touch F60156, Echosens, France). All subjects underwent anthropometric studies to determine body mass index. If it was within one-two Z-score, overweight was diagnosed. If the body mass index exceeded two Z-score, obesity was diagnosed. According to transient elastography and body mass index, all children were divided into four groups: group I — 27 children with MASLD and fibrosis ≥ F1, group II — 35 children with MASLD without fibrosis, group III — 18 obese or overweight children without MASLD and without fibrosis. The control group IV consisted of 14 children with normal weight without MASLD and without fibrosis. The groups had no significant differences in age and gender distribution. The study of body composition was performed by bioimpedance analysis using a TANITA MC-780MA analyzer (manufactured by Maeno-cho, Itabashi-ku, Tokyo, Japan). Quantitative determination of the serum concentration of vascular endothelial growth factor (VEGF) was performed by enzyme-linked immunosorbent assay (ELISA) using test systems from Wuhan Fine Biotech Co., Ltd (China) according to the manufacturer’s recommendations. The level of serum cytokeratin 18 (CK-18) was evaluated with IDL Biotech AB kits (Sweden) for ELISA. Serum content of transforming growth factor beta 1 (TGF-β1) was studied using an ELISA test system from IBL International (Germany). Fibrogenesis processes were evaluated by the serum content of free hydroxyproline (HPf), protein-bound hydroxyproline (HPp/b) and glycosaminoglycans (GAG). Results. The study revealed a significant increase in the level of CK-18 and TGF-β1 in children with MASLD-associated liver fibrosis. In children with liver fibrosis, an increase in the ratio of HPf/HPp/b and the level of GAG in the blood serum was observed compared to patients with MASLD without fibrosis and with overweight and obese children. The threshold value of CK-18 for liver fibrosis diagnosis was 90.3 U/l (sensitivity 81.3 %, specificity 76.9 %, AUC 0.843, p

Background: Tendinopathy alters the compositional properties of the Achilles tendon by increasing fluid and glycosaminoglycan content. It has been speculated that these changes may affect intratendinous pressure, but the extent of this relationship remains unclear. Therefore, we aimed to investigate the impact of elevated fluid and glycosaminoglycan content on Achilles tendon intratendinous pressure and to determine whether hyaluronidase (HYAL) therapy can intervene in this potential relationship. Methods: Twenty paired fresh-frozen cadaveric Achilles tendons were mounted in a tensile-testing machine and loaded up to 5% strain. Intratendinous resting (at 0% strain) and dynamic pressure (at 5% strain) were assessed using the microcapillary infusion technique. First, intratendinous pressure was measured under native conditions before and after infusion of 2 mL physiological saline. Next, 80 mg of glycosaminoglycans were administered bilaterally to the paired tendons. The right tendons were additionally treated with 1500 units of HYAL. Finally, both groups were retested, and the glycosaminoglycan content was analyzed. Results: It was found that both elevated fluid and glycosaminoglycan content resulted in higher intratendinous resting and dynamic pressures (p

Abstract Glycosaminoglycan (GAG) and sialic acid (total and free) assays are used as first‐line screening tests for the diagnosis of mucopolysaccharidoses and glycoproteinoses, respectively. There is a pronounced age‐dependent variation in the urinary concentrations of these metabolites in the normal population, and the stratification of the reference values into discrete age ranges may lead to an undesirably high number of false‐positive or false‐negative results. The aim of this study was to design a method for calculating continuous reference intervals as a function of age and its application to the analysis of GAGs and sialic acid (total, free, and conjugated) in urine. In the postpubertal period, concentrations of urinary GAGs and sialic acid have reached a plateau, so a traditional calculation of the reference range in this specific age group was considered appropriate. In the prepubertal period, a nonlinear regression performed with the Excel add‐in Solver was used to fit the logarithmized concentrations of the controls to a curve that represents the mean values as a function of age. A uniform distribution of the residuals was obtained, which allowed the calculation of the reference intervals by adding the values of their 2.5 and 97.5 percentiles to the independent variable of the regression curve to calculate the upper and lower reference curves. The main advantages of the developed method are (1) a reduction in the number of control samples needed to obtain adequate reference intervals and (2) an improvement in the reliability of diagnostic screening by reducing the uncertainty generated by the gaps in the traditional age‐stratified method.

Background and objective: Screening for bladder cancer (BCa) could reduce mortality via early detection of early-stage high-grade (Ta/T1 N0 M0 grade 2–3) disease. Noninvasive biomarkers could aid in screening, but current markers lack the specificity required. The urinary free glycosaminoglycan profile (GAGome) is a promising biomarker for early detection of BCa metabolism. Methods: In a prospective case-control development study, we included patients with BCa or no evidence of disease (NED) and measured the urinary GAGome. We then developed a score to predict the probability of BCa using GAGome features that correlated with BCa versus NED according to Bayesian regression. Next, in a retrospective, population-based, case-control study, we included adults from the Lifelines Cohort Study who were presumed healthy at baseline. All cases with BCa confirmed in the cancer registry by the 2-yr or 6-yr study visit were matched to randomly selected control subjects. We developed a reference logistic regression model using age and sex to predict BCa at 7 yr after baseline. We then added the GAGome score to the model and assessed model improvement using the likelihood ratio test. We dichotomized outputs for the reference model and saturated model (reference + GAGome score) into high-risk versus low-risk categories using a 99% specificity cutoff and estimated the sensitivity for association with BCa at 7 yr. Key findings and limitations: We prospectively included 51 individuals with BCa and 38 with NED and observed alterations in three GAGome features compatible with BCa. We developed a score that discriminated BCa with an area under the receiver operating characteristic curve of 0.77 (95% confidence interval [CI] 0.67–0.87). We retrospectively selected a cohort of 1088 presumed healthy adults (median age 48 yr, 56% females), of whom 48 had developed BCa by 7 yr after baseline (median time to diagnosis 1.4 yr). The GAGome score was an independent predictor of BCa at 7 yr when added to the reference model (p

Abstract Mucopolysaccharidosis type II (MPS II) is a rare X-linked recessive inherited lysosomal storage disease. With pathogenic variants of the IDS gene, the activity of iduronate-2-sulfatase (IDS) is reduced or lost, causing the inability to degrade glycosaminoglycans (GAGs) in cells and influencing cell function, eventually resulting in multisystemic manifestations, such as a coarse face, dysostosis multiplex, recurrent respiratory tract infections, and hernias. Diagnosing MPS II requires a combination of clinical manifestations, imaging examinations, urinary GAGs screening, enzyme activity, and genetic testing. Currently, symptomatic treatment is the main therapeutic approach. Owing to economic and drug availability issues, only a minority of patients opt for enzyme replacement therapy or hematopoietic stem cell transplantation. The limited awareness of the disease, the lack of widespread detection technology, and uneven economic development contribute to the high rates of misdiagnosis and missed diagnosis in China.

The article explores the heterogeneity of cancer-associated fibroblasts (CAFs) in gastric cancer, categorizing them into three subtypes: myofibroblastic, immune-regulatory, and inflammatory. The study reveals that high expression of these CAF subtypes is associated with poor prognosis in gastric cancer patients. Additionally, the research suggests that targeting CAFs could offer new personalized treatment options for gastric cancer. The study was funded by the National Research Foundation of Korea and approved by the Institutional Review Board of Yonsei Cancer Hospital. [Extracted from the article]

The study of biomolecules and their interactions in their natural environment requires increasingly sophisticated technological and methodological developments. The complexity of these developments is due, among other things, to the nature of these molecules and the small quantities available depending on their origin. In this context, this study focuses on the conditions for improving the detection of glycosaminoglycans on a miniaturized scale by mass spectrometry. These multicharged anionic linear polysaccharides are in fact difficult to study by mass spectrometry and can present, for a given molecule, a large number of signals linked to different charge states, to the loss of one or more sulfate groups and to the presence of different adducts, which reduces sensitivity and complicates the interpretation of the spectra. In order to reduce this complexity, we have investigated different sample preparation methods applicable to small sample volumes. The development of home-made capillary ion-exchange columns, for example, makes it possible to control the adducts formed in nano-ESI coupling. However, their use on a miniaturized scale for detection by MALDI-TOF-MS does not allow for performances as high as those obtained with treatment with a commercial DOWEXTM resin. However, experimental results allowed us to demonstrate that the presence of DOWEXTM resin colloid residues in the aqueous phase greatly improves the quality of the spectra obtained by MALDI-TOFMS on a Fondaparinux model glycosaminoglycan. [ABSTRACT FROM AUTHOR]

The complex collagen network of the native meniscus and the gradient of the density and alignment of this network through the meniscal enthesis is essential for the proper mechanical function of these tissues. This architecture is difficult to recapitulate in tissue-engineered replacement strategies. Prenatally, the organization of the collagen fiber network is established and aggrecan content is minimal. In vitro, fibrochondrocytes (FCCs) produce proteoglycans and associated glycosaminoglycan (GAG) chains early in culture, which can inhibit collagen fiber formation during the maturation of tissue-engineered menisci. Thus, it would be beneficial to both specifically and temporarily block deposition of proteoglycans early in culture. In this study, we transiently inhibited aggrecan production by meniscal fibrochondrocytes using siRNA in collagen gel-based tissue-engineered constructs. We evaluated the effect of siRNA treatment on the formation of collagen fibrils and bulk and microscale tensile properties. Specific inhibition of aggrecan production by fibrochondrocytes via siRNA was successful both in 2D monolayer cell culture and 3D tissue culture. This inhibition during early maturation of these in vitro constructs increased collagen fibril diameter by more than 2-fold. This increase in fibril diameter allowed these tissues to distribute strains more effectively at the local level, particularly at the interface of the bone and soft tissue. These data show that siRNA can be used to modulate the ECM to improve collagen fiber formation and mechanical properties in tissue-engineered constructs, and that a transient decrease in aggrecan promotes the formation of a more robust fiber network. [ABSTRACT FROM AUTHOR]

Graves’ disease (GD) is an autoimmune disease that can affect other tissues in addition to the thyroid gland. The clinical manifestations are a result of the impact of TSH receptor antibodies. Depending on the intensity of the immune response, in addition to hyperthyroidism, orbitopathy, dermatopathy and acropachy can also occur. Extrathyroidal manifestations of GD are most often the result of a more pronounced immune response. Dermatopathy is a rare extrathyroidal manifestation, with a prevalence of 0.5-4.3%, and when it occurs, it almost always accompanies orbitopathy (96%) and hyperthyroidism, while it is associated with a severe form of orbitopathy in 13-15% of cases. It is characteristic of long-standing disease and an intense autoimmune response. We present a case series of five patients with dysthyroid dermopathy. Our patients had different thyroid function disorders (hypo/hyperthyroidism), different forms of dermopathy, and varying times of onset during the disease. Association with orbitopathy and high TRAb concentrations were also present in all our patients. The effects of therapy applied for orbitopathy were monitored, which showed a favorable response on dermopathy, especially with the use of tocilizumab in some of our patients. [ABSTRACT FROM AUTHOR]

Type I collagen is the most abundant structural protein in the body and, with other fibrillar collagens, forms the fibrous network of the extracellular matrix. Another group of extracellular matrix polymers, the glycosaminoglycans, and glycosaminoglycan-modified proteoglycans, play important roles in regulating collagen behaviors and contribute to the compositional, structural, and mechanical complexity of the extracellular matrix. While the binding between collagen and small leucine-rich proteoglycans has been studied in detail, the interactions between collagen and the large bottlebrush proteoglycan versican are not well understood. Here, we report that versican binds collagen directly and regulates collagen structure and mechanics. Versican colocalizes with collagen fibers in vivo and binds to collagen via its C-terminal G3 domain (a non-GAG-modified domain present in all known versican isoforms) in vitro; it promotes the deposition of a highly aligned collagen-rich matrix by fibroblasts. Versican also shows an unexpected effect on the rheology of collagen gels in vitro, causing decreased stiffness and attenuated shear strain stiffening, and the cleavage of versican in the liver results in reduced tissue compression stiffening. Thus, versican is an important collagen-binding partner and plays a role in modulating collagen organization and mechanics. [ABSTRACT FROM AUTHOR]

The majority of naturally occurring mutations of the human gene XDH, are associated with reduced or completely absent xanthine oxidoreductase (XOR) activity, leading to a disease known as classical xanthinuria, which is due to the accumulation and excretion of xanthine in urine. Three types of classical xanthinuria have been identified: type I, characterised by XOR deficiency, type II, caused by XOR and aldehyde oxidase (AO) deficiency, and type III due to XOR, AO, and sulphite oxidase (SO) deficiency. Type I and II are considered rare autosomal recessive disorders, a condition where two copies of the mutated gene must be present to develop the disease or trait. In most cases, xanthinuria type I and II result to be asymptomatic, and only occasionally lead to renal failure due to urolithiasis caused by xanthine deposition. However, in the last 10–15 years, new observations have been made about the link between naturally occurring mutations and pathological phenotypes particularly pertinent to cardiovascular diseases (CVD). These links have been attributed to a genetically driven increase of XOR expression and activity that is responsible for what is thought to be damaging uric acid (UA) and reactive oxygen species (ROS) accumulation, nitric oxide (·NO) depletion and endothelial dysfunction. In this review, we discuss the importance of genetics for interindividual variability of XOR expression and activity while focusing mainly on those variants thought to be relevant for CVD. In addition, we discuss the potential exploitation of the genetically driven increase of XOR activity to deliver more beneficial bioavailable ·NO. Finally, we examine the effect that non-synonymous mutations have on the tertiary structure of the protein and consequently on its capacity to interact with glycosaminoglycans (GAGs) localised on the outer surface of endothelial cells. [ABSTRACT FROM AUTHOR]

Glycosaminoglycans (GAGs), including hyaluronic acid (HA), chondroitin sulfate (CS)/dermatan sulfate (DS), heparan sulfate (HS)/heparin (HP), and keratan sulfate (KS), play pivotal roles in living organisms. Generally, GAGs are analyzed after enzymatic digestion into unsaturated or saturated disaccharides. Due to high structural similarity between disaccharides, however, separation during analysis is challenging. Additionally, little is known about the structures of GAGs and their functional relationships. Elucidating the function of GAGs requires highly sensitive quantitative analytical methods. We developed a method for the simultaneous analysis of 18 types of disaccharides derived from HA (1 type), CS/DS (7 types), HS/HP (8 types), and KS (2 types) potentially detectable in analyses of human urine. The simple method involves HPLC separation with fluorescence detection following derivatization of GAG-derived disaccharides using 4-aminobenzoic acid ethyl ester (ABEE) as a pre-labeling agent and 2-picoline borane as a reductant. The ABEE derivatization reaction can be performed under aqueous conditions, and excess derivatization reagents can be easily, rapidly, and safely removed. This method enables highly sensitive simultaneous analysis of the 18 abovementioned types of GAG-derived disaccharides using HPLC with fluorescence detection in small amounts of urine (1 mL) in a single run. The versatile method described here could be applied to the analysis of GAGs in other biological samples. [ABSTRACT FROM AUTHOR]

Hurler–Scheie syndrome (MPS IH/S), also known as mucopolysaccharidosis type I-H/S (MPS IH/S), is a lysosomal storage disorder caused by deficiency of the enzyme alpha-L-iduronidase (IDUA) leading to the accumulation of glycosaminoglycans (GAGs) in various tissues, resulting in a wide range of symptoms affecting different organ systems. Postgenomic omics technologies offer the promise to understand the changes in proteome, phosphoproteome, and phosphorylation-based signaling in MPS IH/S. Accordingly, we report here a large dataset and the proteomic and phosphoproteomic analyses of fibroblasts derived from patients with MPS IH/S (n = 8) and healthy individuals (n = 8). We found that protein levels of key lysosomal enzymes such as cathepsin D, prosaposin, arylsulfatases (arylsulfatase A and arylsulfatase B), and IDUA were downregulated. We identified 16,693 unique phosphopeptides, corresponding to 4,605 proteins, in patients with MPS IH/S. We found that proteins related to the cell cycle, mitotic spindle assembly, apoptosis, and cytoskeletal organization were differentially phosphorylated in MPS IH/S. We identified 12 kinases that were differentially phosphorylated, including hyperphosphorylation of cyclin-dependent kinases 1 and 2, hypophosphorylation of myosin light chain kinase, and calcium/calmodulin-dependent protein kinases. Taken together, the findings of the present study indicate significant alterations in proteins involved in cytoskeletal changes, cellular dysfunction, and apoptosis. These new observations significantly contribute to the current understanding of the pathophysiology of MPS IH/S specifically, and the molecular mechanisms involved in the storage of GAGs in MPS more generally. Further translational clinical omics studies are called for to pave the way for diagnostics and therapeutics innovation for patients with MPS IH/S. [ABSTRACT FROM AUTHOR]

Aortic dissection is a life-threatening condition caused by layer separation. Despite extensive research, the relationship between the aortic wall's structural integrity and dissection risk remains unclear. Glycosaminoglycan (GAG) accumulation and elastin loss are suspected to play significant roles. We investigated how age-related changes in aortic structure affect dissection susceptibility. Peeling tests were performed on longitudinal and circumferential thoracic (TA) and abdominal aortic (AA) strips from 35 donors aged 13–76 years (mean 38 ± 15 years, 34 % female). GAG, elastin, collagen, and smooth muscle cell (SMC) contents were assessed using bidirectional histology. Young TAs resisted longitudinal peeling better than circumferential, with delamination strengths of 65.4 mN/mm and 44.2 mN/mm, respectively. Delamination strength decreased with age in both directions, more rapidly longitudinally, equalizing at ∼20–25 mN/mm in older TAs. Delamination strength in AAs was 22 % higher than in TAs. No sex differences were observed. GAG density increased, while elastin density decreased by 2.5 % and 4 % per decade, respectively. Collagen density did not change with age, while SMC density decreased circumferentially. GAGs partially mediated the reduction in longitudinal delamination strength due to aging, while circumferential strength reduction was not mediated by changes in either GAG or elastin densities. This study explains why aortic dissections are more common in TAs, especially in older individuals, and why they typically propagate spirally. TAs exhibit lower delamination strength compared to AAs and experience strength reduction with age, a phenomenon linked to increased GAG accumulation and elastin loss. These findings enhance our understanding of the pathophysiological mechanisms behind aortic dissection. This work explores the age-dependent relationships between delamination strength in human aortas and wall structural content. We investigated 35 human aortas from donors aged 13 to 76 years, providing new insights into the biomechanical and histological factors that influence aortic dissection risk. Our findings elucidate how variations in elastin, glycosaminoglycan, collagen, and smooth muscle cell densities impact the structural integrity of the aorta, contributing significantly to the understanding of aortic dissection mechanisms. [Display omitted] [ABSTRACT FROM AUTHOR]

• Intratendinous pressure of the Achilles tendon increases during tensile loading. It is unknown whether tendinopathic changes, namely fluid and glycosaminoglycan accumulation, may impact this. • Intratendinous pressure of 20 Achilles tendons was measured at rest and during loading under natural conditions and after administration of fluid and glycosaminoglycans. Additionally, half of the tendons were treated with hyaluronidase. • Elevated fluid and glycosaminoglycan content resulted in higher intratendinous pressures. Hyaluronidase reduced glycosaminoglycan content and lowered intratendinous pressures. • These findings show that fluid and glycosaminoglycan accumulation lead to increased intratendinous pressures and that hyaluronidase may serve as a potential treatment by restoring intratendinous pressures. Tendinopathy alters the compositional properties of the Achilles tendon by increasing fluid and glycosaminoglycan content. It has been speculated that these changes may affect intratendinous pressure, but the extent of this relationship remains unclear. Therefore, we aimed to investigate the impact of elevated fluid and glycosaminoglycan content on Achilles tendon intratendinous pressure and to determine whether hyaluronidase (HYAL) therapy can intervene in this potential relationship. Twenty paired fresh-frozen cadaveric Achilles tendons were mounted in a tensile-testing machine and loaded up to 5% strain. Intratendinous resting (at 0% strain) and dynamic pressure (at 5% strain) were assessed using the microcapillary infusion technique. First, intratendinous pressure was measured under native conditions before and after infusion of 2 mL physiological saline. Next, 80 mg of glycosaminoglycans were administered bilaterally to the paired tendons. The right tendons were additionally treated with 1500 units of HYAL. Finally, both groups were retested, and the glycosaminoglycan content was analyzed. It was found that both elevated fluid and glycosaminoglycan content resulted in higher intratendinous resting and dynamic pressures (p < 0.001). HYAL treatment induced a 2.3-fold reduction in glycosaminoglycan content (p = 0.002) and restored intratendinous pressures. The results of this study demonstrated that elevated fluid and glycosaminoglycan content in Achilles tendinopathy contribute to increased intratendinous resting and dynamic pressures, which can be explained by the associated increased volume and reduced permeability of the tendon matrix, respectively. HYAL degrades glycosaminoglycans sufficiently to lower intratendinous pressures and may, therefore, serve as a promising treatment. [Display omitted] [ABSTRACT FROM AUTHOR]

Repair of the damaged meniscus is a scientific challenge owing to the poor self-healing potential of the white area of the meniscus. Tissue engineering provides a new method for the repair of meniscus injuries. In this study, we explored the superiority of 2% hyaluronic acid chitin hydrogel in temperature sensitivity, in vitro degradation, biocompatibility, cell adhesion, and other biological characteristics, and investigated the advantages of hyaluronic acid (HA) and Transforming Growth Factor β1 (TGF-β1) in promoting cell proliferation and a matrix formation phenotype. The hydrogel loaded with HA and TGF-β1 promoted cell proliferation. The HA + TGF-β1 mixed group showed the highest glycosaminoglycan (GAG) content and promoted cell migration. Hydroxypropyl chitin (HPCH), HA, and TGF-β1 were combined to form a composite hydrogel with a concentration of 2% after physical cross-linking, and this was injected into a rabbit model of a meniscus full-thickness tear. After 12 weeks of implantation, the TGF-β1 + HA/HPCH composite hydrogel was significantly better than HPCH, HA/HPCH, TGF-β1 + HPCH, and the control group in promoting meniscus repair. In addition, the new meniscus tissue of the TGF-β1 + HA/HPCH composite hydrogel had a tissue structure and biochemical content similar to that of the normal meniscus tissue. [ABSTRACT FROM AUTHOR]

Background: The use of biological scaffolds in regenerative endodontics has gained much attention in recent years. The search for a new biomimetic scaffold that contains tissue-specific cell homing factors could lead to more predictable tissue regeneration. The aim of this study was to prepare and characterize decellularized bovine dental pulp-derived extracellular matrix (P-ECM) hydrogels for regenerative endodontic applications. Methods: Freshly extracted bovine molar teeth were collected. Bovine dental pulp tissues were harvested, and stored at -40º C. For decellularization, a 5-day protocol was implemented incorporating trypsin/EDTA, deionized water and DNase treatment. Decellularization was evaluated by DNA quantification and histological examination to assess collagen and glycosaminoglycans (GAGs) content. This was followed by the preparation of P-ECM hydrogel alone or combined with hyaluronic acid gel (P-ECM + HA). The fabricated scaffolds were then characterized using protein quantification, hydrogel topology and porosity, biodegradability, and growth factor content using Enzyme-linked immunosorbent assay (ELISA): transforming growth factor beta-1(TGF-β1), basic fibroblast growth factor (bFGF), bone morphogenetic protein 2 (BMP-2) and vascular endothelial growth factor (VEGF). Results: Decellularization was histologically confirmed, and DNA content was below (50 ng/mg tissue). P-ECM hydrogel was prepared with a final ECM concentration of 3.00 mg/ml while P-ECM + HA hydrogel was prepared with a final ECM concentration of 1.5 mg/ml. Total protein content in P-ECM hydrogel was found to be (439.0 ± 123.4 µg/µl). P-ECM + HA showed sustained protein release while the P-ECM group showed gradual decreasing release. Degradation was higher in P-ECM + HA which had a significantly larger fiber diameter, while P-ECM had a larger pore area percentage. ELISA confirmed the retention and release of growth factors where P-ECM hydrogel had higher BMP-2 release, while P-ECM + HA had higher release of TGF-β1, bFGF, and VEGF. Conclusions: Both P-ECM and P-ECM + HA retained their bioactive properties demonstrating a potential role as functionalized scaffolds for regenerative endodontic procedures. [ABSTRACT FROM AUTHOR]

Background: Scheie syndrome, an attenuated subtype of mucopolysaccharidosis type I, is a rare storage disease that causes progressive glycosaminoglycans (GAGs) accumulation. Cardiovascular disorders determine the prognosis, and cardiac valve abnormalities are the most common cause. The patients are usually young so mechanical valve replacement is often performed, but because of the features of this disease, the surgical treatment is very risky. Recently, transcatheter aortic valve implantation (TAVI) has been reported as an alternative choice for aortic stenosis, but optimal choice is still unclear. Here, we introduce a patient that underwent TAVI and refer to the histological finding of a biological valve extracted in relation to GAGs accumulation. Case presentation: A 54-year-old woman with Scheie syndrome underwent valve surgeries three times throughout her whole life. At age 41, she received a mitral valve replacement with a mechanical valve for mitral stenosis. She promptly developed severe diastolic dysfunction and low output syndrome after the release of aortic clamping, thus requiring temporary mechanical circulatory support4. At age 51, she suffered from heart failure due to severe aortic stenosis and underwent TAVI because conventional aortic valve replacement (AVR) was deemed too risky. Three years later, her heart failure relapsed, and an echocardiogram unexpectedly revealed thickened bioprosthetic valve leaflets and a significant pressure gradient across the valve, consistent with early structural valve deterioration. AVR was performed via median sternotomy with a mechanical valve. As with the first operation, she presented refractory heart failure requiring mechanical circulatory support and was meticulously managed. However, she steadily showed worsening of multiple organ systems and died 9 days after the operation. Pathological autopsy and histological examination revealed accumulation of tissue with GAGs on the leaflets of the bioprosthetic valve of TAVI, which may have been the cause of early structural valve deterioration. Conclusion: For patients with Scheie syndrome, a biological valve can be compromised by the accumulation of GAGs, thereby causing early SVD. These findings may support valve selection for these high-risk patients. [ABSTRACT FROM AUTHOR]

Age is a prominent risk factor for cardiometabolic disease, often leading to heart structural and functional changes. However, precise molecular mechanisms underlying cardiac remodeling and dysfunction exclusively resulting from physiological aging remain elusive. Previous research demonstrated age‐related functional alterations in baboons, analogous to humans. The goal of this study is to identify early cardiac molecular alterations preceding functional adaptations, shedding light on the regulation of age‐associated changes. Unbiased transcriptomics of left ventricle samples are performed from female baboons aged 7.5–22.1 years (human equivalent ≈30–88 years). Weighted‐gene correlation network and pathway enrichment analyses are performed, with histological validation. Modules of transcripts negatively correlated with age implicated declined metabolism‐oxidative phosphorylation, tricarboxylic acid cycle, glycolysis, and fatty‐acid β‐oxidation. Transcripts positively correlated with age suggested a metabolic shift toward glucose‐dependent anabolic pathways, including hexosamine biosynthetic pathway (HBP). This shift is associated with increased glycosaminoglycan synthesis, modification, precursor synthesis via HBP, and extracellular matrix accumulation, verified histologically. Upregulated extracellular matrix‐induced signaling coincided with glycosaminoglycan accumulation, followed by cardiac hypertrophy‐related pathways. Overall, these findings revealed a transcriptional shift in metabolism favoring glycosaminoglycan accumulation through HBP before cardiac hypertrophy. Unveiling this metabolic shift provides potential targets for age‐related cardiac diseases, offering novel insights into early age‐related mechanisms. [ABSTRACT FROM AUTHOR]

Cellular senescence is associated with various age‐related disorders and is assumed to play a major role in the pathogenesis of osteoarthritis (OA). Based on this, we tested a senolytic combination therapy using Dasatinib (D) and Quercetin (Q) on aged isolated human articular chondrocytes (hACs), as well as in OA‐affected cartilage tissue (OARSI grade 1–2). Stimulation with D + Q selectively eliminated senescent cells in both, cartilage explants and isolated hAC. Furthermore, the therapy significantly promoted chondroanabolism, as demonstrated by increased gene expression levels of COL2A1, ACAN, and SOX9, as well as elevated collagen type II and glycosaminoglycan biosynthesis. Additionally, D + Q treatment significantly reduced the release of SASP factors (IL6, CXCL1). RNA sequencing analysis revealed an upregulation of the anabolic factors, inter alia, FGF18, IGF1, and TGFB2, as well as inhibitory effects on cytokines and the YAP‐1 signaling pathway, explaining the underlying mechanism of the chondroanabolic promotion upon senolytic treatment. Accordingly, stimulation of untreated hAC with conditioned medium of D + Q‐treated cells similarly induced the expression of chondrogenic markers. Detailed analyses demonstrated that chondroanabolic effects could be mainly attributed to Dasatinib, while monotherapeutical application of Quercetin or Navitoclax did not promote the chondroanabolism. Overall, D + Q therapy restored the chondrogenic phenotype in OA hAC most likely by creating a pro‐chondroanabolic environment through the reduction of SASP factors and upregulation of growth factors. This senolytic approach could therefore be a promising candidate for further testing as a disease‐modifying osteoarthritis drug. [ABSTRACT FROM AUTHOR]

Mucopolysaccharidosis type II (MPS II) is a rare X-linked recessive inherited lysosomal storage disease. With pathogenic variants of the IDS gene, the activity of iduronate-2-sulfatase (IDS) is reduced or lost, causing the inability to degrade glycosaminoglycans (GAGs) in cells and influencing cell function, eventually resulting in multisystemic manifestations, such as a coarse face, dysostosis multiplex, recurrent respiratory tract infections, and hernias. Diagnosing MPS II requires a combination of clinical manifestations, imaging examinations, urinary GAGs screening, enzyme activity, and genetic testing. Currently, symptomatic treatment is the main therapeutic approach. Owing to economic and drug availability issues, only a minority of patients opt for enzyme replacement therapy or hematopoietic stem cell transplantation. The limited awareness of the disease, the lack of widespread detection technology, and uneven economic development contribute to the high rates of misdiagnosis and missed diagnosis in China. [ABSTRACT FROM AUTHOR]

Tendons enable movement through their highly aligned extracellular matrix (ECM), predominantly composed of collagen I. Tendinopathies disrupt the structural integrity of tendons by causing fragmentation of collagen fibers, disorganization of fiber bundles, and an increase in glycosaminoglycans and microvasculature, thereby driving the apparent biomechanical and regenerative capacity in patients. Moreover, the complex cellular communication within the tendon microenvironment ultimately dictates the fate between healthy and diseased tendon, wherein extracellular vesicles (EVs) may facilitate the tendon's fate by transporting biomolecules within the tissue. In this study, we aimed to elucidate how the EV functionality is altered in the context of tendon microenvironments by using polycaprolactone (PCL) electrospun scaffolds mimicking healthy and pathological tendon matrices. Scaffolds were characterized for fiber alignment, mechanical properties, and cellular activity. EVs were isolated and analyzed for concentration, heterogeneity, and protein content. Our results show that our mimicked healthy tendon led to an increase in EV secretion and baseline metabolic activity over the mimicked diseased tendon, where reduced EV secretion and a significant increase in metabolic activity over 5 days were observed. These findings suggest that scaffold mechanics may influence EV functionality, offering insights into tendon homeostasis. Future research should further investigate how EV cargo affects the tendon's microenvironment. [ABSTRACT FROM AUTHOR]

Background: Non-invasive detection of renal cell carcinoma (RCC) recurrence is a major challenge that could radically affect patient survival. To date, there are no approved biomarkers for inclusion in the monitoring and follow-up of RCC; therefore, assessment of treatment response is lacking. Materials and Methods: A Cross-sectional study was carried out on biopsy-proven renal cell carcinoma patients scheduled for nephrectomy at Hayatabad Medical Complex (HMC) Hospital, Peshawar, Pakistan, between September 2022 and January 2024. Results: A total of 160 samples were included in the study. Eighty samples were obtained from biopsy-proven non-metastatic renal cell carcinoma patients, of which 40 were collected pre-nephrectomy, 40 were collected post-nephrectomy from the same patients, and 80 samples were collected from age and gender-matched healthy individuals. Total plasma glycosaminoglycans (GAGs) levels were analyzed through a manual enzyme-linked immunosorbent assay using a non-competitive sandwich technique. Quantitative variables were summarized as means and standard deviation, while qualitative variables were summarized as frequency and percentage. A paired t-test was performed to check whether the difference between the mean plasma GAG levels in prenephrectomy and post-nephrectomy groups was significant. Conclusion: The results revealed that post-nephrectomy GAG levels have significantly decreased compared to pre-nephrectomy levels (P<0.001). Plasma glycosaminoglycan levels could be promising markers for monitoring renal cell carcinoma patients post-nephrectomy. [ABSTRACT FROM AUTHOR]

During HIV-1 virus assembly, the genomic RNA (vRNA) is selected for packaging by the viral protein Gag because it contains a specific packaging signal, Psi. While there have been numerous studies of Gag–Psi interactions, there is almost no information on the kinetic aspects of this interaction. We investigated the kinetics of Gag binding to different RNAs using switchSENSE DRX2 technology. We measured the association rate of Gag binding to monomeric Psi, to a "Multiple Binding Site Mutant" Psi that is inactive for genome packaging in vivo, and to a scrambled Psi. We discovered that Gag binds more rapidly to Psi RNA than to the mutant or scrambled RNAs. Furthermore, rapid Gag association kinetics are retained within sub-regions of Psi: Gag associates more rapidly with RNA containing only the 3′ two of the three Psi stem-loops than with monomeric RNA containing the 5′ two stem-loops or a scrambled RNA. No differences were detectable with individual Psi stem-loops. Interestingly, the rate of binding of Gag molecules to Psi increases with increasing Gag concentration, suggesting cooperativity in binding. The results are consistent with the hypothesis that selectivity in packaging derives from kinetic differences in initiation of particle assembly. [ABSTRACT FROM AUTHOR]

Current strategies for autologous chondrocyte implantation explore the use of gel systems that have structural similarity to an extracellular matrix of cartilage tissue and can act as chondrocyte carriers. However, the major requirement is to address the challenge of maintaining the phenotype and bio-functionality of chondrocytes which often undergo dedifferentiation on expansion, and the appropriate mechanical properties of the matrix that can influence cell attachment and growth. Injectable gel systems are developed in this study by blending different ratios of methacrylated chitosan and polyethylene glycol diacrylate, and photocross-linking the system to attain a stable gel. We hypothesize a multiscale approach of evaluating various parameters like the physical and structural properties of the gel, its wettability and surface free energy and mechano-rheological properties along with the biological characterization (viability assay, collagen, and glycosaminoglycan estimation by immunostaining, biochemical analysis, and RT-PCR assay) of the gel in ascertaining its ideal matrix system. The study provides a correlation between the different parameters to arrive at the most optimum characteristic of an ideal gel system which is dependent not only on the elastic modulus and surface free energy properties of the matrix but also on the composition of the matrix and the phenotype and functionality of the encapsulated chondrocytes. [ABSTRACT FROM AUTHOR]

Background. The COVID-19 epidemic and the war in Ukraine have led to a significant increase in the number of children suffering from metabolic dysfunction-associated steatotic liver disease (MASLD). One of the unresolved problems associated with MASLD is the identification of individuals at risk of rapid disease progression and development of irreversible liver changes. The search for alternative noninvasive markers suitable for the early detection of liver fibrosis in children remains extremely relevant. The aim of the study was to determine the diagnostic value of serum fibrosis markers and their relationship with sonographic and body composition parameters in children with MASLD. Materials and methods. The case-control study included 80 children aged 6 to 17 years (mean of (12.15 ± 2.51) years). The presence of steatosis and liver fibrosis was determined by transient elastography (FibroScan® 502 touch F60156, Echosens, France). All subjects underwent anthropometric studies to determine body mass index. If it was within one-two Z-score, overweight was diagnosed. If the body mass index exceeded two Z-score, obesity was diagnosed. According to transient elastography and body mass index, all children were divided into four groups: group I — 27 children with MASLD and fibrosis ≥ F1, group II — 35 children with MASLD without fibrosis, group III — 18 obese or overweight children without MASLD and without fibrosis. The control group IV consisted of 14 children with normal weight without MASLD and without fibrosis. The groups had no significant differences in age and gender distribution. The study of body composition was performed by bioimpedance analysis using a TANITA MC-780MA analyzer (manufactured by Maeno-cho, Itabashi-ku, Tokyo, Japan). Quantitative determination of the serum concentration of vascular endothelial growth factor (VEGF) was performed by enzyme-linked immunosorbent assay (ELISA) using test systems from Wuhan Fine Biotech Co., Ltd (China) according to the manufacturer’s recommendations. The level of serum cytokeratin 18 (CK-18) was evaluated with IDL Biotech AB kits (Sweden) for ELISA. Serum content of transforming growth factor beta 1 (TGF-β1) was studied using an ELISA test system from IBL International (Germany). Fibrogenesis processes were evaluated by the serum content of free hydroxyproline (HPf), protein-bound hydroxyproline (HPp/b) and glycosaminoglycans (GAG). Results. The study revealed a significant increase in the level of CK-18 and TGF-β1 in children with MASLD-associated liver fibrosis. In children with liver fibrosis, an increase in the ratio of HPf/HPp/b and the level of GAG in the blood serum was observed compared to patients with MASLD without fibrosis and with overweight and obese children. The threshold value of CK-18 for liver fibrosis diagnosis was 90.3 U/l (sensitivity 81.3 %, specificity 76.9 %, AUC 0.843, p < 0.001). The sensitivity of the threshold value of serum TGF-β1 (96.8 pg/mL) in children with MASLD was 80.0 %, specificity 65.7 %, AUC 0.787 (p < 0.001). Threshold value of serum GAG (4.24 mmol/L) demonstrated a sensitivity of 70.6 % and a specificity of 69.6 %, AUC 0.743 (p < 0.01). CK-18, TGF-β1, GAG shown a positive correlation with liver stiffness and elasticity, body composition of MASLD children and had high levels of diagnostic accuracy, which allows them to be used in children when screening for MASLD-associated liver fibrosis. Conclusions. Children with liver fibrosis are characterized by elevated serum levels of CK-18, VEGF, TGF-β1, HPp/b and GAG. The threshold values of CK-18 (more than 90.3 U/l), TGF-β1 (above 96.8 pg/mL) and GAG (more than 4.24 mmol/l) have high sensitivity and specificity, which allows them to be used for the diagnosis of liver fibrosis in children with MASLD. [ABSTRACT FROM AUTHOR]

The temporomandibular joint (TMJ) disc complex (i.e., the TMJ disc and its six attachments) is crucial to everyday functions such as mastication and speaking. The TMJ can be afflicted by many conditions, including disc displacement and defects. Pathologies of the TMJ disc complex most commonly present first as anterior disc displacement, which the field hypothesizes may implicate the two posterior attachments. As a result of anterior disc displacement, defects may develop in the lateral disc complex. Tissue engineering is poised to improve treatment paradigms for these indications of the TMJ disc complex by engineering biomimetic implants, but, first, gold-standard design criteria for such implants should be established through characterization studies. This study's objective was to characterize the structural, mechanical, biochemical, and crosslinking differences among the two posterior attachments and the lateral disc in the Yucatan minipig, a well-accepted TMJ animal model. In tension, it was found that the posterior inferior attachment (PIA) was significantly stiffer and stronger by 2.13 and 2.30 times, respectively, than the posterior superior attachment (PSA). It was found that collagen in both attachments was primarily aligned mediolaterally; however, the lateral disc was much more aligned and anisotropic than either attachment. Among the three locations, the PSA exhibited the greatest degree of heterogeneity and highest proportion of fat vacuoles. The PIA and lateral disc were 1.93 and 1.91 times more collagenous, respectively, by dry weight (DW) than the PSA. The PIA also exhibited 1.78 times higher crosslinking per DW than the PSA. Glycosaminoglycan per DW was significantly higher in the lateral disc by 1.48 and 5.39 times than the PIA and PSA, respectively. Together, these results establish design criteria for tissue-engineering of the TMJ disc complex and indicate that the attachments are less fibrocartilaginous than the disc, while still significantly contributing to the mechanical stability of the TMJ disc complex during articulation. These results also support the biomechanical function of the PIA and PSA, suggesting that the stiffer PIA anchors the disc to the mandibular condyle during articulation, while the softer PSA serves to allow translation over the articular eminence. Impact Statement Characterization of the temporomandibular joint (TMJ) disc complex (i.e., the disc and its attachments) has important implications for those aiming to tissue-engineer functional replacements and can help elucidate its biomechanical function. For example, the findings shown here suggest that the stiffer posterior inferior attachment anchors the disc during articulation, while the softer posterior superior attachment allows translation over the articular eminence.

Abstract Background Syndecan‐1 (SDC1) is an established marker of endothelial glycocalyx shedding. Most research on SDC1 has focused on plasma or serum concentrations, and little is known about urine concentrations. Objectives Measure urinary SDC1 concentrations in dogs undergoing anesthesia with either sevoflurane or isoflurane and assess the effects of anesthesia duration and IV crystalloids on urinary SDC1 concentrations. Animals Thirty‐one client‐owned dogs undergoing anesthesia for magnetic resonance imaging (MRI) with or without surgery for suspected intervertebral disk disease (IVDD) were used. Methods Dogs with suspected IVDD were randomized to undergo anesthesia with either sevoflurane or isoflurane. Urine was collected before and immediately after anesthesia for the analysis of SDC1. Urinary creatinine concentrations also were measured, and the ratio of urinary SDC1 to urinary creatinine (USCR) was used to account for dilution. Results Median (range) USCR was significantly higher after anesthesia compared with baseline for all groups combined (P

The existing in vitro and in vivo models for studying osteoarthritis have significant limitations in replicating the complexity of joint tissues. This research aims to validate a Tissue-On-a-Chip system for osteoarthritis research. Osteochondral tissues obtained from knee replacement surgeries of patients with osteoarthritis were cultured in an Organ-On-a-Chip system. This system was designed to supply oxygen and glucose to the cartilage from the bone. The distribution of oxygen and glucose was evaluated by fluorescence using Image-iT Green Hypoxia and 2-NBDG, respectively. Cytotoxicity was measured using lactate dehydrogenase (LDH) levels in chip cultures compared to plate cultures (12 tissues per method). Glycosaminoglycans (GAGs), alkaline phosphatase (ALP), Coll2-1, and procollagen type II N-terminal propeptide (PIINP) were measured in the perfused medium of the Tissue-On-a-Chip over a period of 70 days. Fluorescence of Image-iT Green Hypoxia was observed only in the cartilage area, while 2-NBDG was distributed throughout the tissue. An increase in LDH levels was noted in the plate cultures on day 24 and in the Tissue-On-a-Chip cultures on day 63. Compared to the start of the culture, GAG content increased on day 52, while ALP showed variations. A notable increase in GAG, ALP, and Coll2-1 levels was observed on day 59. PIINP levels remained stable throughout the experiment. The validated osteochondral Tissue-On-a-Chip system can replicate the joint microenvironment, with hypoxic conditions in cartilage and normoxic conditions in bone. Tissue survival and component stability were maintained for approximately two months. This platform is a useful tool for evaluating new drugs and represents a viable alternative to animal models. [ABSTRACT FROM AUTHOR]

We aimed to investigate the use of free glycosaminoglycan profiles (GAGomes) and cfDNA in plasma to differentiate between lung cancer and benign lung disease, in a cohort of 113 patients initially suspected of lung cancer. GAGomes were analyzed in all samples using the MIRAM® Free Glycosaminoglycan Kit with ultra-high-performance liquid chromatography and electrospray ionization triple quadrupole mass spectrometry. In a subset of samples, cfDNA concentration and NGS-data was available. We detected two GAGome features, 0S chondroitin sulfate (CS), and 4S CS, with cancer-specific changes. Based on the observed GAGome changes, we devised a model to predict lung cancer. The model, named the GAGome score, could detect lung cancer with 41.2% sensitivity (95% CI: 9.2–54.2%) at 96.4% specificity (95% CI: 95.2–100.0%, n = 113). When we combined the GAGome score with a cfDNA-based model, the sensitivity increased from 42.6% (95% CI: 31.7–60.6%, cfDNA alone) to 70.5% (95% CI: 57.4–81.5%) at 95% specificity (95% CI: 75.1–100%, n = 74). Notably, the combined GAGome and cfDNA testing improved the sensitivity, compared to cfDNA alone, especially in ASCL stage I (55.6% vs 11.1%). Our findings show that plasma GAGome profiles can enhance cfDNA testing performance, highlighting the applicability of a multiomics approach in lung cancer diagnostics. [ABSTRACT FROM AUTHOR]

Background: Premature rupture of the membranes (PROM) is a key cause of preterm birth and represents a major cause of neonatal mortality and morbidity. Natural products N-acetyl-d-galactosamine (GalNAc), which are basic building blocks of important polysaccharides in biological cells or tissues, such as chitin, glycoproteins, and glycolipids, may improve possible effects of wound healing. Methods: An in vitro inflammation and oxidative stress model was constructed using tumor necrosis-α (TNF-α) and lipopolysaccharide (LPS) action on WISH cells. Human amniotic epithelial cells (hAECs) were primarily cultured by digestion to construct a wound model. The effects of GalNAc on anti-inflammatory and anti-oxidative stress, migration and proliferation, epithelial-mesenchymal transition (EMT), glycosaminoglycan (GAG)/hyaluronic acid (HA) production, and protein kinase B (Akt) pathway in hAECs and WISH cells were analyzed using the DCFH-DA fluorescent probe, ELISA, CCK-8, scratch, transwell migration, and western blot to determine the mechanism by which GalNAc promotes amniotic wound healing. Results: GalNAc decreased IL-6 expression in TNF-α-stimulated WISH cells and ROS expression in LPS-stimulated WISH cells (P < 0.05). GalNAc promoted the expression of Gal-1 and Gal-3 with anti-inflammatory and anti-oxidative stress effects. GalNAc promoted the migration of hAECs (50% vs. 80%) and WISH cells through the Akt signaling pathway, EMT reached the point of promoting fetal membrane healing, and GalNAc did not affect the activity of hAECs and WISH cells (P > 0.05). GalNAc upregulated the expression of sGAG in WISH cells (P < 0.05) but did not affect HA levels (P > 0.05). Conclusions: GalNAc might be a potential target for the prevention and treatment of PROM through the galectin pathway, including (i) inflammation; (ii) epithelial-mesenchymal transition; (iii) proliferation and migration; and (iv) regression, remodeling, and healing. [ABSTRACT FROM AUTHOR]

Glycosaminoglycans (GAGs) play a key role in a variety of biological processes in the extracellular matrix (ECM) via interactions with their protein targets. Due to their high flexibility, periodicity and electrostatics-driven interactions, GAG-containing complexes are very challenging to characterize both experimentally and in silico. In this study, we, for the first time, systematically analyzed the interactions of endostatin, a proteolytic fragment of collagen XVIII known to be anti-angiogenic and anti-tumoral, with heparin (HP) and representative heparan sulfate (HS) oligosaccharides of various lengths, sequences and sulfation patterns. We first used conventional molecular docking and a docking approach based on a repulsive scaling–replica exchange molecular dynamics technique, as well as unbiased molecular dynamic simulations, to obtain dynamically stable GAG binding poses. Then, the corresponding free energies of binding were calculated and the amino acid residues that contribute the most to GAG binding were identified. We also investigated the potential influence of Zn2+ on endostatin–HP complexes using computational approaches. These data provide new atomistic details of the molecular mechanism of HP's binding to endostatin, which will contribute to a better understanding of its interplay with proteoglycans at the cell surface and in the extracellular matrix. [ABSTRACT FROM AUTHOR]

Several years ago, dozens of cases were described in patients with symptoms very similar to mucopolysaccharidosis (MPS). This new disease entity was described as mucopolysaccharidosis-plus syndrome (MPSPS). The name of the disease indicates that in addition to the typical symptoms of conventional MPS, patients develop other features such as congenital heart defects and kidney and hematopoietic system disorders. The symptoms are highly advanced, and patients usually do not survive past the second year of life. MPSPS is inherited in an autosomal recessive manner and is caused by a homozygous-specific mutation in the gene encoding the VPS33A protein. To date, it has been described in 41 patients. Patients with MPSPS exhibited excessive excretion of glycosaminoglycans (GAGs) in the urine and exceptionally high levels of heparan sulfate in the plasma, but the accumulation of substrates is not caused by a decrease in the activity of any lysosomal enzymes. Here, we discuss the pathomechanisms and symptoms of MPSPS, comparing them to those of MPS. Moreover, we asked the question whether MPSPS should be classified as a type of MPS or a separate disease, as contrary to 'classical' MPS types, despite GAG accumulation, no defects in lysosomal enzymes responsible for degradation of these compounds could be detected in MPSPS. The molecular mechanism of the appearance of GAG accumulation in MPSPS is suggested on the basis of results available in the literature. [ABSTRACT FROM AUTHOR]

Inflammation models are widely used in the in vitro investigation of new therapeutic approaches for osteoarthritis. TNFα (tumor necrosis factor alpha) plays an important role in the inflammatory process. Current inflammation models lack uniformity and make comparisons difficult. Therefore, this study aimed to systematically investigate whether the effects of TNFα are concentration-dependent and whether chondrocyte expansion has an effect on the inflammatory model. Bovine chondrocytes were enzymatically isolated, expanded to passages 1–3, and transferred into a 3D pellet culture. Chondrocyte pellets were stimulated with recombinant bovine TNFα at different concentrations for 48 h to induce inflammation. Gene expression of anabolic (collagen 2, aggrecan, cartilage oligomeric protein (COMP)), catabolic (matrix metalloproteinases (MMP3, MMP13)), dedifferentiation (collagen 1) markers, inflammation markers (interleukin-6 (IL-6), nuclear factor kappa B (NFkB), cyclooxygenase-2 (COX), prostaglandin-E-synthase-2 (PTGES2)), and the apoptosis marker caspase 3 was determined. At the protein level, concentrations of IL-6, nitric oxide (NO), and sulfated glycosaminoglycans (GAG) were evaluated. Statistical analysis was performed using the independent t-test, and significance was defined as p < 0.05. In general, TNFα caused a decrease in anabolic markers and an increase in the expression of catabolic and inflammatory markers. There was a concentration-dependent threshold of 10 ng/mL to induce significant inflammatory effects. Most of the markers analyzed showed TNFα concentration-dependent effects (COMP, PRG4, AGN, Col1, MMP3, and NFkB). There was a statistical influence of selected gene expression markers from different passages on the TNFα chondrocyte inflammation model, including Col2, MMP13, IL-6, NFkB, COX2, and PTGES2. Considering the expression of collagen 2 and MMP3, passage 3 chondrocytes showed a higher sensitivity to TNFα stimulation compared to passages 1 and 2. On the other hand, MMP13, IL-6, NFkB, and caspase 3 gene expression were lower in P3 chondrocytes compared to the other passages. On the protein level, inflammatory effects showed a similar pattern, with cytokine effects starting at 10 ng/mL and differences between the passages. TNFα had a detrimental effect on cartilage, with a clear threshold observed at 10 ng/mL. Although TNFα effects showed concentration-dependent patterns, this was not consistent for all markers. The selected passage showed a clear influence, especially on inflammation markers. Further experiments were warranted to explore the effects of TNFα concentration and passage in long-term stimulation. [ABSTRACT FROM AUTHOR]

Snails exhibit remarkable adaptability, allowing them to flourish in diverse environmental conditions and resulting in thriving populations in specific regions. This abundance has led communities to harness snails for various purposes, including their use as animal feed, daily dietary source, and in traditional wound-healing practices with historical roots. The primary objective of this systematic review is to identify the snail species commonly employed in wound healing and evaluate the bioactivity of compounds derived from different snail species. This review was conducted using literature review method, drawing from international databases such as Scopus, and encompassed publications from 2013 to 2023. A total of 22 articles met the inclusion and exclusion criteria. Snail body parts that have been explored for wound-healing purposes include both the body and the shell, along with snail secretions, particularly their mucus. Various methods have been employed to extract mucus, involving manual stimulation of the snail's body, spraying with a saline solution (NaCl), application of electric shock, and the use of ozone gas through nebulisation. Prominent snail species found to be beneficial for wound healing include Achatina fulica, Helix aspersa, Eobania desertorum, Helix lucurus, Cornu bistrialis, Theba pisana, and Megalobulimus lopesi. These snail species demonstrate potential applications in the treatment of burns, excision wounds, incision wounds, and diabetic ulcers. Key compounds within snail secretions encompass mucopolysaccharides, polyphenols, peptides, and glycosaminoglycans. These compounds exert significant effects on haemostasis, inflammation control, cellular proliferation, and re-epithelialisation, significantly contributing to the wound healing process. [ABSTRACT FROM AUTHOR]

Glycosaminoglycans (GAGs) and proteoglycans (PGs) are essential components of the extracellular matrix (ECM) with pivotal roles in cellular mechanosensing pathways. GAGs, such as heparan sulfate (HS) and chondroitin sulfate (CS), interact with various cell surface receptors, including integrins and receptor tyrosine kinases, to modulate cellular responses to mechanical stimuli. PGs, comprising a core protein with covalently attached GAG chains, serve as dynamic regulators of tissue mechanics and cell behavior, thereby playing a crucial role in maintaining tissue homeostasis. Dysregulation of GAG/PG-mediated mechanosensing pathways is implicated in numerous pathological conditions, including cancer and inflammation. Understanding the intricate mechanisms by which GAGs and PGs modulate cellular responses to mechanical forces holds promise for developing novel therapeutic strategies targeting mechanotransduction pathways in disease. This comprehensive overview underscores the importance of GAGs and PGs as key mediators of mechanosensing in maintaining tissue homeostasis and their potential as therapeutic targets for mitigating mechano-driven pathologies, focusing on cancer and inflammation. [ABSTRACT FROM AUTHOR]