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Emerging infectious diseases are caused by unpredictable viruses with the dangerous potential to trigger global pandemics. These viruses typically initiate infection by utilizing the anionic structures of host cell surface receptors to gain entry. Lactoferrin (Lf) is a multifunctional glycoprotein with multiple properties such as antiviral, anti-inflammatory and antioxidant activities. Due to its cationic structure, Lf naturally interacts with certain host cell receptors, such as heparan sulfate proteoglycans, as well as viral particles and other receptors that are targeted by viruses. Therefore, Lf may interfere with virus-host cell interactions by acting as a receptor competitor for viruses. Herein we summarize studies in which this competition was investigated with SARS-CoV-2, Zika, Dengue, Hepatitis and Influenza viruses in vitro. These studies have demonstrated not only Lf's competitive properties, but also its potential intracellular impact on host cells, such as enhancing cell survival and reducing infection efficiency by inhibiting certain viral enzymes. In addition, the immunomodulatory effect of Lf is highlighted, as it can influence the activity of specific immune cells and regulate cytokine release, thereby enhancing the host's response to viral infections. Collectively, these properties promote the potential of Lf as a promising candidate for research in viral infectious diseases. [ABSTRACT FROM AUTHOR]

Introduction: Heparin binding proteins (HBPs) with roles in extracellular matrix assembly are strongly correlated to β-amyloid (Aβ) and tau pathology in Alzheimer's disease (AD) brain and cerebrospinal fluid (CSF). However, it remains challenging to detect these proteins in plasma using standard mass spectrometry-based proteomic approaches. Methods: We employed heparin-affinity chromatography, followed by off-line fractionation and tandem mass tag mass spectrometry (TMT-MS), to enrich HBPs from plasma obtained from AD (n = 62) and control (n = 47) samples. These profiles were then correlated to Aβ, tau and phosphorylated tau (pTau) CSF biomarkers and plasma pTau181 from the same individuals, as well as a consensus brain proteome network to assess the overlap with AD brain pathophysiology. Results: Heparin enrichment from plasma was highly reproducible, enriched well-known HBPs like APOE and thrombin, and depleted high-abundant proteins such as albumin. A total of 2865 proteins, spanning 10 orders of magnitude in abundance, were measured across 109 samples. Compared to the consensus AD brain protein co-expression network, we observed that specific plasma proteins exhibited consistent direction of change in both brain and plasma, whereas others displayed divergent changes, highlighting the complex interplay between the two compartments. Elevated proteins in AD plasma, when compared to controls, included members of the matrisome module in brain that accumulate with Aβ deposits, such as SMOC1, SMOC2, SPON1, MDK, OLFML3, FRZB, GPNMB, and the APOE4 proteoform. Additionally, heparin-enriched proteins in plasma demonstrated significant correlations with conventional AD CSF biomarkers, including Aβ, total tau, pTau, and plasma pTau181. A panel of five plasma proteins classified AD from control individuals with an area under the curve (AUC) of 0.85. When combined with plasma pTau181, the panel significantly improved the classification performance of pTau181 alone, increasing the AUC from 0.93 to 0.98. This suggests that the heparin-enriched plasma proteome captures additional variance in cognitive dementia beyond what is explained by pTau181. Conclusion: These findings support the utility of a heparin-affinity approach coupled with TMT-MS for enriching amyloid-associated proteins, as well as a wide spectrum of plasma biomarkers that reflect pathological changes in the AD brain. [ABSTRACT FROM AUTHOR]

Purpose: Heparan sulfate proteoglycans (HSPGs) are complex molecules found on the cell membrane and within the extracellular matrix, increasingly recognized for their role in tumor progression. This study aimed to investigate the involvement of Heparan sulfate proteoglycan 2 (HSPG2) in the progression of bladder cancer. Methods: We identified HSPG2 as a promoter of bladder tumor progression using single-cell RNA sequencing and transcriptome analysis of sequencing data from seven patient samples obtained from the Gene Expression Omnibus (GEO) database (GSE135337). Transcript profiles of 28 normal tissues and 407 bladder tumor tissues were analyzed for HSPG2 expression and survival outcomes using the Sanger tools and cBioPortal databases. HSPG2-overexpressing T24 and Biu-87 cell lines were generated, and cell proliferation and migration were assessed using CCK-8 and Transwell assays. Western blotting and immunostaining were performed to evaluate the activation of Nidogen-1 (NID1)/protein kinase B (AKT) signaling. Mouse models with patient-derived tumor organoids (HSPG2high and HSPG2low) were established to assess anticancer effects. Results: Our results demonstrated a marked upregulation of HSPG2 in malignant bladder tumors, which correlated significantly with poor patient prognosis. HSPG2 overexpression consistently enhanced bladder tumor cell proliferation and conferred chemotherapy resistance, as shown in both in vitro and in vivo experiments. Mechanistically, HSPG2 upregulated NID1 expression, leading to the activation of the AKT pro-survival signaling pathway and promoting sustained tumor growth in bladder cancer. Conclusion: This study highlights the critical pro-tumor role of HSPG2/NID1/AKT signaling in bladder cancer and suggests its potential as a therapeutic target in clinical treatment. [ABSTRACT FROM AUTHOR]

Background: Idiopathic pulmonary fibrosis (IPF) is a chronic and progressive interstitial lung disease of unknown etiology. Despite the increasing global incidence and poor prognosis, the exact pathogenic mechanisms remain elusive. Currently, effective therapeutic targets and treatment methods for this disease are still lacking. This study tried to explore the pathogenic mechanisms of IPF. We found elevated expression of SULF1 in lung tissues of IPF patients compared to normal control lung tissues. SULF1 is an enzyme that modifies heparan sulfate chains of heparan sulfate proteoglycans, playing a critical role in biological regulation. However, the effect of SULF1 in pulmonary fibrosis remains incompletely understood. Our study aimed to investigate the impact and mechanisms of SULF1 in fibrosis. Methods: We collected lung specimens from IPF patients for transcriptome sequencing. Validation of SULF1 expression in IPF patients was performed using Western blotting and RT-qPCR on lung tissues. ELISA experiments were employed to detect SULF1 concentrations in IPF patient plasma and TGF-β1 levels in cell culture supernatants. We used lentiviral delivery of SULF1 shRNA to knock down SULF1 in HFL1 cells, evaluating its effects on fibroblast secretion, activation, proliferation, migration, and invasion capabilities. Furthermore, we employed Co-Immunoprecipitation (Co-IP) to investigate the regulatory mechanisms involved. Results: Through bioinformatic analysis of IPF transcriptomic sequencing data (HTIPF) and datasets GSE24206, and GSE53845, we identified SULF1 may potentially play a crucial role in IPF. Subsequently, we verified that SULF1 was upregulated in IPF and predominantly increased in fibroblasts. Furthermore, SULF1 expression was induced in HFL1 cells following exposure to TGF-β1. Knockdown of SULF1 suppressed fibroblast secretion, activation, proliferation, migration, and invasion under both TGF-β1-driven and non-TGF-β1-driven conditions. We found that SULF1 catalyzes the release of TGF-β1 bound to TGFβRIII, thereby activating the TGF-β1/SMAD pathway to promote fibrosis. Additionally, TGF-β1 induces SULF1 expression through the TGF-β1/SMAD pathway, suggesting a potential positive feedback loop between SULF1 and the TGF-β1/SMAD pathway. Conclusions: Our findings reveal that SULF1 promotes fibrosis through the TGF-β1/SMAD pathway in pulmonary fibrosis. Targeting SULF1 may offer a promising therapeutic strategy against IPF. [ABSTRACT FROM AUTHOR]

Abstract Introduction Heparin binding proteins (HBPs) with roles in extracellular matrix assembly are strongly correlated to β-amyloid (Aβ) and tau pathology in Alzheimer’s disease (AD) brain and cerebrospinal fluid (CSF). However, it remains challenging to detect these proteins in plasma using standard mass spectrometry-based proteomic approaches. Methods We employed heparin-affinity chromatography, followed by off-line fractionation and tandem mass tag mass spectrometry (TMT-MS), to enrich HBPs from plasma obtained from AD (n = 62) and control (n = 47) samples. These profiles were then correlated to Aβ, tau and phosphorylated tau (pTau) CSF biomarkers and plasma pTau181 from the same individuals, as well as a consensus brain proteome network to assess the overlap with AD brain pathophysiology. Results Heparin enrichment from plasma was highly reproducible, enriched well-known HBPs like APOE and thrombin, and depleted high-abundant proteins such as albumin. A total of 2865 proteins, spanning 10 orders of magnitude in abundance, were measured across 109 samples. Compared to the consensus AD brain protein co-expression network, we observed that specific plasma proteins exhibited consistent direction of change in both brain and plasma, whereas others displayed divergent changes, highlighting the complex interplay between the two compartments. Elevated proteins in AD plasma, when compared to controls, included members of the matrisome module in brain that accumulate with Aβ deposits, such as SMOC1, SMOC2, SPON1, MDK, OLFML3, FRZB, GPNMB, and the APOE4 proteoform. Additionally, heparin-enriched proteins in plasma demonstrated significant correlations with conventional AD CSF biomarkers, including Aβ, total tau, pTau, and plasma pTau181. A panel of five plasma proteins classified AD from control individuals with an area under the curve (AUC) of 0.85. When combined with plasma pTau181, the panel significantly improved the classification performance of pTau181 alone, increasing the AUC from 0.93 to 0.98. This suggests that the heparin-enriched plasma proteome captures additional variance in cognitive dementia beyond what is explained by pTau181. Conclusion These findings support the utility of a heparin-affinity approach coupled with TMT-MS for enriching amyloid-associated proteins, as well as a wide spectrum of plasma biomarkers that reflect pathological changes in the AD brain.

Abstract Background Vascular endothelial growth factor (VEGF) is an important proangiogenic factor and has been considered as a key target of antiangiogenetic therapy in oral squamous cell carcinoma (OSCC). However, clinical application of bevacizumab, a specific VEGF antibody, didn’t improve the survival rate of OSCC patients. One possible explanation is that VEGF gene expresses diverse isoforms, which associate with extracellular vesicles (EVs), and EVs potentially contribute to VEGF resistance to bevacizumab. However, clear solution is lacking in addressing this issue. Methods Expression of VEGF isoforms in OSCC cells was confirmed by reverse transcription and polymerase chain reaction (RT-PCR) and western blot. EVs isolated from OSCC cell’s conditioned medium (CM) were characterized by western blot, transmission electron microscopy (TEM) and nanoparticle tracking analysis (NTA). Flow cytometry, immunogold labeling and western blot were applied to study the VEGF on EVs. Tube formation assay and Matrigel plug angiogenesis assay were used for analyzing the angiogenesis capacity of EV-VEGF. Results The most popular isoforms expressed by VEGF gene are VEGF121, VEGF165 and VEGF189. In this study, we demonstrated that all three isoforms of mRNA could be detected at varying levels in OSCC cells, while only VEGF165 and VEGF189 proteins were found. CM derived from OSCC cells, both soluble and non-soluble forms of VEGF could be detected. We further confirmed the presence of VGEF189 bound to EVs as a non-soluble form. EV-bound VEGF189 presented angiogenic activity, which could not be neutralized by bevacizumab. It was found that VEGF189 bound to EVs by heparan sulfate proteoglycans (HSPG). In addition, the angiogenic effect of EV-VEGF could be reversed by surfen, a kind of HSPG antagonist both in vitro and in vivo. Conclusion Antagonists targeting HSPG might potentially overcome the resistance of EV-VEGF to bevacizumab and serve as an alternative for anti-VEGF therapy in OSCC.

Heparan sulfate proteoglycans (HSPGs) regulate a wide range of biological activities in both physiological and pathological conditions. Altered expression or deregulated function of HSPGs and their heparan sulfate (HS) chains significantly contribute to carcinogenesis as well and crucially depends on the functioning of the complex system of HS biosynthetic/modifying enzymes termed as "GAGosome". Here, we aimed at investigating the expression profile of the system in a cell culture model of stroma-epithelial crosstalk and searching for transcription factors potentially related to the regulation of expression of the genes involved. Coculture of BjTERT-fibroblasts with normal PNT2 human prostate epithelial cells resulted in significant downregulation (2-4-fold) of transcriptional activity of HS metabolism-involved genes (EXT1/2, NDST1/2, GLCE, HS2ST1, HS3ST1/2, HS6ST1/2, SULF1/2, HPSE) in both cell types, whereas coculture with prostate cancer cells (LNCaP, PC3, DU145) demonstrated no significant interchanges. Human Transcription Factor RT2 Profiler PCR array and manual RT-PCR verification supposed FOS, MYC, E2F, SRF, NR3C1 as potential candidates for regulation and/or coordination of HS biosynthesis. Taken together, transcriptional activity of HS biosynthetic system in normal fibroblasts and prostate epithelial cells during their coculture might be controlled by their intercellular communication, reflecting of adaptation of these cells to each other. The regulation is attenuated or abrogated if normal fibroblasts interact with prostate cancer cells making the cancer cells independent of the limiting effects of fibroblasts, thus contributing to possibility of unlimited growth and progression. Overall, these data demonstrate an ability of cell-cell interactions to affect transcriptional activity of HS biosynthesis-involved genes. [ABSTRACT FROM AUTHOR]

Background: Vascular endothelial growth factor (VEGF) is an important proangiogenic factor and has been considered as a key target of antiangiogenetic therapy in oral squamous cell carcinoma (OSCC). However, clinical application of bevacizumab, a specific VEGF antibody, didn't improve the survival rate of OSCC patients. One possible explanation is that VEGF gene expresses diverse isoforms, which associate with extracellular vesicles (EVs), and EVs potentially contribute to VEGF resistance to bevacizumab. However, clear solution is lacking in addressing this issue. Methods: Expression of VEGF isoforms in OSCC cells was confirmed by reverse transcription and polymerase chain reaction (RT-PCR) and western blot. EVs isolated from OSCC cell's conditioned medium (CM) were characterized by western blot, transmission electron microscopy (TEM) and nanoparticle tracking analysis (NTA). Flow cytometry, immunogold labeling and western blot were applied to study the VEGF on EVs. Tube formation assay and Matrigel plug angiogenesis assay were used for analyzing the angiogenesis capacity of EV-VEGF. Results: The most popular isoforms expressed by VEGF gene are VEGF121, VEGF165 and VEGF189. In this study, we demonstrated that all three isoforms of mRNA could be detected at varying levels in OSCC cells, while only VEGF165 and VEGF189 proteins were found. CM derived from OSCC cells, both soluble and non-soluble forms of VEGF could be detected. We further confirmed the presence of VGEF189 bound to EVs as a non-soluble form. EV-bound VEGF189 presented angiogenic activity, which could not be neutralized by bevacizumab. It was found that VEGF189 bound to EVs by heparan sulfate proteoglycans (HSPG). In addition, the angiogenic effect of EV-VEGF could be reversed by surfen, a kind of HSPG antagonist both in vitro and in vivo. Conclusion: Antagonists targeting HSPG might potentially overcome the resistance of EV-VEGF to bevacizumab and serve as an alternative for anti-VEGF therapy in OSCC. [ABSTRACT FROM AUTHOR]

To ensure compositional consistency while mitigating potential immunogenicity for stem cell therapy, synthetic scaffolds have emerged as compelling alternatives to native extracellular matrix (ECM). Substantial progress has been made in emulating specific natural traits featuring consistent chemical compositions and physical structures. However, recapitulating the dynamic responsiveness of the native ECM involving chemical transitions and physical remodeling during differentiation, remains a challenging endeavor. Here, the creation of adaptive scaffolds is demonstrated through sequential protein‐instructed molecular assembly, utilizing stage‐specific proteins, and incorporating in situ assembly technique. The procedure is commenced by introducing a dual‐targeting peptide at the onset of stem cell differentiation. In response to highly expressed integrins and heparan sulfate proteoglycans (HSPGs) on human mesenchymal stem cell (hMSC), the peptides assembled in situ, creating customized extracellular scaffolds that adhered to hMSCs promoting osteoblast differentiation. As the expression of alkaline phosphatase (ALP) and collagen (COL‐1) increased in osteoblasts, an additional peptide is introduced that interacts with ALP, initiating peptide assembly and facilitating calcium phosphate (CaP) deposition. The growth and entanglement of peptide assemblies with collagen fibers efficiently incorporated CaP into the network resulting in an adaptive biphasic scaffold that enhanced healing of bone injuries. [ABSTRACT FROM AUTHOR]

Hepatitis B virus (HBV) expresses co-terminal large (L), middle (M), and small (S) envelope proteins containing preS1/preS2/S, preS2/S, and S domain alone, respectively. S and preS1 domains mediate sequential virion attachment to heparan sulfate proteoglycans and sodium taurocholate cotransporting polypeptide (NTCP), respectively, which can be blocked by anti-S and anti-preS1 antibodies. How anti-preS2 antibodies neutralize HBV infectivity remains enigmatic. The late stage of chronic HBV infection often selects for mutated preS2 translation initiation codon to prevent M protein expression, or in-frame preS2 deletions to shorten both L and M proteins. When introduced to infectious clone of genotype C or D, both M-minus mutations and most 5’ preS2 deletions sustained virion production. Such mutant progeny viral particles were infectious in NTCP-reconstituted HepG2 cells. Neutralization experiments were performed on the genotype D clone. Although remaining susceptible to anti-preS1 and anti-S neutralizing antibodies, M-minus mutants were only partially neutralized by two anti-preS2 antibodies tested while preS2 deletion mutants were resistant. By infection experiments using viral particles with lost versus increased M protein expression, or a neutralization escaping preS2 deletion only present on L or M protein, we found that both full-length L and M proteins contributed to virus neutralization by the two anti-preS2 antibodies. Thus, immune escape could be a driving force for the selection of M-minus mutations, and especially preS2 deletions. The fact that both L and M proteins could mediate neutralization by anti-preS2 antibodies may shed light on the underlying molecular mechanism. IMPORTANCE The large (L), middle (M), and small (S) envelope proteins of hepatitis B virus (HBV) contain preS1/preS2/S, preS2/S, and S domain alone, respectively. The discovery of heparan sulfate proteoglycans and sodium taurocholate cotransporting polypeptide (NTCP) as the low- and high-affinity HBV receptors could explain neutralizing potential of anti-S and anti-preS1 antibodies, respectively, but how anti-preS2 neutralizing antibodies work remains enigmatic. In this study, we found two M-minus mutants in the context of genotype D partially escaped two anti-preS2 neutralizing antibodies in NTCP-reconstituted HepG2 cells, while several naturally occurring preS2 deletion mutants escaped both antibodies. By point mutations to eliminate or enhance M protein expression, and by introducing preS2 deletion selectively to L or M protein, we found binding of anti-preS2 antibodies to both L and M proteins contributed to neutralization of wild-type HBV infectivity. Our finding may shed light on the possible mechanism(s) whereby anti-preS2 antibodies neutralize HBV infectivity. [ABSTRACT FROM AUTHOR]

Antithrombin III (ATIII) is a potent endogenous anticoagulant that binds to heparan sulfate proteoglycans (HSPGs) on endothelial cells' surfaces. Among these HSPGs, syndecans (SDCs) are crucial as transmembrane receptors bridging extracellular ligands with intracellular signaling pathways. Specifically, syndecan-4 (SDC4) has been identified as a key receptor on endothelial cells for transmitting the signaling effects of ATIII. Meanwhile, SDCs have been implicated in facilitating the cellular internalization of SARS-CoV-2. Given the complex interactions between ATIII and SDC4, our study analyzed the impact of ATIII on the virus entry into host cells. While ATIII binds to all SDC isoforms, it shows the strongest affinity for SDC4. SDCs' heparan sulfate chains primarily influence ATIII's SDC attachment, although other parts might also play a role in ATIII's dominant affinity toward SDC4. ATIII significantly reduces SARS-CoV-2′s cellular entry into cell lines expressing SDCs, suggesting a competitive inhibition mechanism at the SDC binding sites, particularly SDC4. Conversely, the virus or its spike protein decreases the availability of SDCs on the cell surface, reducing ATIII's cellular attachment and hence contributing to a procoagulant environment characteristic of COVID-19. [ABSTRACT FROM AUTHOR]

Emerging infectious diseases are caused by unpredictable viruses with the dangerous potential to trigger global pandemics. These viruses typically initiate infection by utilizing the anionic structures of host cell surface receptors to gain entry. Lactoferrin (Lf) is a multifunctional glycoprotein with multiple properties such as antiviral, anti-inflammatory and antioxidant activities. Due to its cationic structure, Lf naturally interacts with certain host cell receptors, such as heparan sulfate proteoglycans, as well as viral particles and other receptors that are targeted by viruses. Therefore, Lf may interfere with virus-host cell interactions by acting as a receptor competitor for viruses. Herein we summarize studies in which this competition was investigated with SARS-CoV-2, Zika, Dengue, Hepatitis and Influenza viruses in vitro. These studies have demonstrated not only Lf’s competitive properties, but also its potential intracellular impact on host cells, such as enhancing cell survival and reducing infection efficiency by inhibiting certain viral enzymes. In addition, the immunomodulatory effect of Lf is highlighted, as it can influence the activity of specific immune cells and regulate cytokine release, thereby enhancing the host’s response to viral infections. Collectively, these properties promote the potential of Lf as a promising candidate for research in viral infectious diseases.

Abstract Objective Bariatric surgery induces a significant loss of both fat mass (FM) and fat-free mass (FFM). The proteoglycan receptor syndecan-4 (SDC4) plays a crucial role in adipose tissue and skeletal muscle functions. Thus, this study was performed (i) to assess plasma SDC4 levels after both Sleeve Gastrectomy (SG) and Roux-en-Y Gastric Bypass (RYGB) surgeries, and (ii) to explore potential associations with changes in body composition variables. Results Twenty-six patients (17 females) with severe obesity underwent SG (n = 13) or RYGB (n = 13) and were followed up to 1 year (1Y). Body weight, FM, FFM, and SCD4 were measured at baseline (BL), and at week 11 (W11) and 1Y after surgery. Independently of procedure, there was a significant body weight loss at W11, with an average FM and FFM reduction of 13.7 ± 0.6 kg and 5.3 ± 0.5 kg, respectively. Participants continued to lose weight afterwards, with a total weigth loss of 38.2 ± 1.5 kg at 1Y. No associations were found at BL between SDC4 levels and any anthropometric variable; however, SDC4 levels were lower than BL at both W11 and 1Y, independently of type of surgery. Additionally, changes in SDC4 between BL and 1Y were positively correlated with weight and FFM loss during the same period. Trial registration ClinicalTrials.gov NCT04051190 on 09/08/2019.

Objective: Various enzymes, reactive oxygen species, inflammatory conditions, and major surgeries cause endothelial glycocalyx breakdown. Inhalation of anaesthetic agents may have protective effects on the endothelium. This study compared syndecan-1 and heparan sulfate levels to evaluate the effects of sevoflurane and desflurane on the endothelial glycocalyx. Methods: This prospective randomized, double-blind study included 46 patients undergoing laparoscopic hysterectomy. The participants were allocated into sevoflurane and desflurane groups. Subsequently, blood samples were drawn at three time points: before anaesthesia induction for a baseline value (T0), after pneumoperitoneum (T1), and after extubation (T2). Heparan sulfate and syndecan-1 levels were measured. Results: There was no statistical difference between the sevoflurane and desflurane groups in terms of heparan sulfate and syndecan-1 levels at any time point. A significant difference was found only in the desflurane group in the intragroup comparisons of the measurements of heparan sulfate levels (χ2=29.826, P < 0.001). Matched pairs of the time points in the desflurane group showed that P=0.036 (Z=-2.099) for T1-T0, P < 0.001 (Z=-3.924) for T2-T0, and P < 0.001 (Z=-4.197) for T2-T1. The change in percentage between T2 and T1 of heparan sulfate in the desflurane group was found to be statistically significant (P=0.034). Conclusion: The damage caused by surgical stress on the endothelial glycocalyx can be reduced by both desflurane and sevoflurane. The protective effect of desflurane is more prominent than that of sevoflurane.

BackgroundPCSK9 (proprotein convertase subtilisin-kexin type 9) chaperones the hepatic LDLR (low-density lipoprotein receptor) for lysosomal degradation, elevating serum LDL (low-density lipoprotein) cholesterol and promoting atherosclerotic heart disease. Though the major effect on the hepatic LDLR comes from secreted PCSK9, the details of PCSK9 reuptake into the hepatocyte remain unclear. In both tissue culture and animal models, HSPGs (heparan sulfate proteoglycans) on hepatocytes act as co-receptors to promote PCSK9 reuptake. We hypothesized that if this PCSK9:HSPG interaction is important in humans, disrupting it with unfractionated heparin (UFH) would acutely displace PCSK9 from the liver and increase plasma PCSK9.MethodsWe obtained remnant plasma samples from 160 subjects undergoing cardiac catheterization before and after administration of intravenous UFH. PCSK9 levels were determined using a commercial enzyme-linked immunosorbent assay.ResultsMedian plasma PCSK9 was 113 ng/mL prior to UFH and 119 ng/mL afterward. This difference was not significant (P=0.83 [95% CI, -6.23 to 6.31 ng/mL]). Equivalence testing provided 95% confidence that UFH would not raise plasma PCSK9 by > 4.7%. Among all subgroups, only subjects with the lowest baseline PCSK9 concentrations exhibited a response to UFH (8.8% increase, adj. P=0.044). A modest correlation was observed between baseline plasma PCSK9 and the change in plasma PCSK9 due to UFH (RS=-0.3634; P

Objective: Bariatric surgery induces a significant loss of both fat mass (FM) and fat-free mass (FFM). The proteoglycan receptor syndecan-4 (SDC4) plays a crucial role in adipose tissue and skeletal muscle functions. Thus, this study was performed (i) to assess plasma SDC4 levels after both Sleeve Gastrectomy (SG) and Roux-en-Y Gastric Bypass (RYGB) surgeries, and (ii) to explore potential associations with changes in body composition variables. Results: Twenty-six patients (17 females) with severe obesity underwent SG (n = 13) or RYGB (n = 13) and were followed up to 1 year (1Y). Body weight, FM, FFM, and SCD4 were measured at baseline (BL), and at week 11 (W11) and 1Y after surgery. Independently of procedure, there was a significant body weight loss at W11, with an average FM and FFM reduction of 13.7 ± 0.6 kg and 5.3 ± 0.5 kg, respectively. Participants continued to lose weight afterwards, with a total weigth loss of 38.2 ± 1.5 kg at 1Y. No associations were found at BL between SDC4 levels and any anthropometric variable; however, SDC4 levels were lower than BL at both W11 and 1Y, independently of type of surgery. Additionally, changes in SDC4 between BL and 1Y were positively correlated with weight and FFM loss during the same period. Trial registration: ClinicalTrials.gov NCT04051190 on 09/08/2019. [ABSTRACT FROM AUTHOR]

Heparan sulfate proteoglycans are highly glycosylated proteins in which heparan sulfate, a glycosaminoglycan sugar chain, is an acidic sugar chain consisting of a repeating disaccharide structure of glucuronic acid and N-acetylglucosamine is locally sulfated. Syndecan, one of the transmembrane HSPGs, functions as a receptor that transmits signals from the extracellular microenvironment to the inside of the cell. In the vascular system, heparan sulfate proteoglycans, a major component of the glycocalyx, enable the binding of various plasma-derived molecules due to their diversity, epimerization of glycosaminoglycans chains, long chains, and sulfation. Heparan sulfate proteoglycans present in the extracellular matrix serve as a reservoir for bioactive molecules such as chemokines, cytokines, and growth factors. Aberrant expression of heparan sulfate proteoglycans, heparanase, and sulfatase is observed in many pathological conditions. Therefore, it can be applied to therapeutic strategies for a wide range of fields including Alzheimer's disease, heart failure, cancer, organ transplants, diabetes, chronic inflammation, aging, and autoimmune diseases. [ABSTRACT FROM AUTHOR]

Numerous viruses have been found to exploit glycoconjugates expressed on human cells as their initial attachment factor for viral entry and infection. The virus-cell glycointeractome, when characterized, may serve as a template for antiviral drug design. Heparan sulfate proteoglycans extensively decorate the human cell surface and were previously described as a primary receptor for human metapneumovirus (HMPV). After respiratory syncytial virus, HMPV is the second most prevalent respiratory pathogen causing respiratory tract infection in young children. To date, there is neither vaccine nor drug available to prevent or treat HMPV infection. Using a multidisciplinary approach, we report for the first time the glycointeractome of the HMPV fusion (F) protein, a viral surface glycoprotein that is essential for target-cell recognition, attachment, and entry. Our glycan microarray and surface plasmon resonance results suggest that Galβ1-3/4GlcNAc moieties that may be sialylated or fucosylated are readily recognized by HMPV F. The bound motifs are highly similar to the N-linked and O-linked glycans primarily expressed on the human lung epithelium. We demonstrate that the identified glycans have the potential to compete with the cellular receptors used for HMPV entry and consequently block HMPV infection. We found that lacto-N-neotetraose demonstrated the strongest HMPV binding inhibition in a cell infection assay. Our current findings offer an encouraging and novel avenue for the design of anti-HMPV drug candidates using oligosaccharide templates. IMPORTANCE All cells are decorated with a dense coat of sugars that makes a sugar code. Many respiratory viruses exploit this sugar code by binding to these sugars to cause infection. Human metapneumovirus is a leading cause for acute respiratory tract infections. Despite its medical importance, there is no vaccine or antiviral drug available to prevent or treat human metapneumovirus infection. This study investigates how human metapneumovirus binds to sugars in order to more efficiently infect the human host. We found that human metapneumovirus binds to a diverse range of sugars and demonstrated that these sugars can ultimately block viral infection. Understanding how viruses can take advantage of the sugar code on our cells could identify new intervention and treatment strategies to combat viral disease. [ABSTRACT FROM AUTHOR]

High-grade gliomas (HGGs) and glioblastoma multiforme (GBM) are characterized by a heterogeneous and aggressive population of tissue-infiltrating cells that promote both destructive tissue remodeling and aberrant vascularization of the brain. The formation of defective and permeable blood vessels and microchannels and destructive tissue remodeling prevent efficient vascular delivery of pharmacological agents to tumor cells and are the significant reason why therapeutic chemotherapy and immunotherapy intervention are primarily ineffective. Vessel-forming endothelial cells and microchannel-forming glial cells that recapitulate vascular mimicry have both infiltration and destructive remodeling tissue capacities. The transmembrane protein TMEM230 (C20orf30) is a master regulator of infiltration, sprouting of endothelial cells, and microchannel formation of glial and phagocytic cells. A high level of TMEM230 expression was identified in patients with HGG, GBM, and U87-MG cells. In this study, we identified candidate genes and molecular pathways that support that aberrantly elevated levels of TMEM230 play an important role in regulating genes associated with the initial stages of cell infiltration and blood vessel and microchannel (also referred to as tumor microtubule) formation in the progression from low-grade to high-grade gliomas. As TMEM230 regulates infiltration, vascularization, and tissue destruction capacities of diverse cell types in the brain, TMEM230 is a promising cancer target for heterogeneous HGG tumors. [ABSTRACT FROM AUTHOR]

Objective: Various enzymes, reactive oxygen species, inflammatory conditions, and major surgeries cause endothelial glycocalyx breakdown. Inhalation of anaesthetic agents may have protective effects on the endothelium. This study compared syndecan-1 and heparan sulfate levels to evaluate the effects of sevoflurane and desflurane on the endothelial glycocalyx. Methods: This prospective randomized, double-blind study included 46 patients undergoing laparoscopic hysterectomy. The participants were allocated into sevoflurane and desflurane groups. Subsequently, blood samples were drawn at three time points: before anaesthesia induction for a baseline value (T0), after pneumoperitoneum (T1), and after extubation (T2). Heparan sulfate and syndecan-1 levels were measured. Results: There was no statistical difference between the sevoflurane and desflurane groups in terms of heparan sulfate and syndecan-1 levels at any time point. A significant difference was found only in the desflurane group in the intragroup comparisons of the measurements of heparan sulfate levels (χ²=29.826, P < 0.001). Matched pairs of the time points in the desflurane group showed that P=0.036 (Z=-2.099) for T1-T0, P < 0.001 (Z=-3.924) for T2-T0, and P < 0.001 (Z=-4.197) for T2-T1. The change in percentage between T2 and T1 of heparan sulfate in the desflurane group was found to be statistically significant (P=0.034). Conclusion: The damage caused by surgical stress on the endothelial glycocalyx can be reduced by both desflurane and sevoflurane. The protective effect of desflurane is more prominent than that of sevoflurane. [ABSTRACT FROM AUTHOR]

Heparan sulfate proteoglycans are highly glycosylated proteins in which heparan sulfate, a glycosaminoglycan sugar chain, is an acidic sugar chain consisting of a repeating disaccharide structure of glucuronic acid and N-acetylglucosamine is locally sulfated. Syndecan, one of the transmembrane HSPGs, functions as a receptor that transmits signals from the extracellular microenvironment to the inside of the cell. In the vascular system, heparan sulfate proteoglycans, a major component of the glycocalyx, enable the binding of various plasma-derived molecules due to their diversity, epimerization of glycosaminoglycans chains, long chains, and sulfation. Heparan sulfate proteoglycans present in the extracellular matrix serve as a reservoir for bioactive molecules such as chemokines, cytokines, and growth factors. Aberrant expression of heparan sulfate proteoglycans, heparanase, and sulfatase is observed in many pathological conditions. Therefore, it can be applied to therapeutic strategies for a wide range of fields including Alzheimer’s disease, heart failure, cancer, organ transplants, diabetes, chronic inflammation, aging, and autoimmune diseases.

Integration of extracellular signals by neurons is pivotal for brain development, plasticity, and repair. Axon guidance relies on receptor-ligand interactions crosstalking with extracellular matrix components. Semaphorin-5A (Sema5A) is a bifunctional guidance cue exerting attractive and inhibitory effects on neuronal growth through the interaction with heparan sulfate (HS) and chondroitin sulfate (CS) glycosaminoglycans (GAGs), respectively. Sema5A harbors seven thrombospondin type-1 repeats (TSR1-7) important for GAG binding, however the underlying molecular basis and functions in vivo remain enigmatic. Here we dissect the structural basis for Sema5A:GAG specificity and demonstrate the functional significance of this interaction in vivo. Using x-ray crystallography, we reveal a dimeric fold variation for TSR4 that accommodates GAG interactions. TSR4 co-crystal structures identify binding residues validated by site-directed mutagenesis. In vitro and cell-based assays uncover specific GAG epitopes necessary for TSR association. We demonstrate that HS-GAG binding is preferred over CS-GAG and mediates Sema5A oligomerization. In vivo, Sema5A:GAG interactions are necessary for Sema5A function and regulate Plexin-A2 dependent dentate progenitor cell migration. Our study rationalizes Sema5A associated developmental and neurological disorders and provides mechanistic insights into how multifaceted guidance functions of a single transmembrane cue are regulated by proteoglycans. Semaphorin 5A (Sema5A) forms complexes with heparan and chondroitin sulfate proteoglycans to regulate neuronal migration. Here, the authors show that the thrombospondin-like repeat 4 (TSR4) of Sema5A enables glycosaminoglycan association, multimerization, and neural progenitor cell distribution. [ABSTRACT FROM AUTHOR]

Wnt proteins are thought to be transported in several ways in the extracellular space. For instance, they are known to be carried by exosomes and by Wnt-carrier proteins, such as sFRP proteins. However, little is known about whether and/or how these two transport systems are related. Here, we show that adding sFRP1 or sFRP2, but not sFRP3 or sFRP4, to culture medium containing Wnt3a or Wnt5a increases re-secretion of exosome-loaded Wnt proteins from cells. This effect of sFRP2 is counteracted by heparinase, which removes sugar chains on heparan sulfate proteoglycans (HSPGs), but is independent of LRP5/6, Wnt co-receptors essential for Wnt signaling. Wnt3a and Wnt5a specifically dimerize with sFRP2 in culture supernatant. Furthermore, a Wnt3a mutant defective in heterodimerization with sFRP2 impairs the ability to increase exosome-mediated Wnt3a re-secretion. Based on these results, we propose that Wnt heterodimerization with its carrier protein, sFRP2, enhances Wnt accumulation at sugar chains on HSPGs on the cell surface, leading to increased endocytosis and exosome-mediated Wnt re-secretion. Our results suggest that the range of action of Wnt ligands is controlled by coordination of different transport systems. Various carriers are involved in extracellular Wnt transport. Here, the authors show that Wnt heterodimerization with its carrier protein, sFRP2, enhances Wnt accumulation on the cell surface, leading to exosome-mediated Wnt re-secretion. [ABSTRACT FROM AUTHOR]

[Display omitted] We have learned from the recent COVID-19 pandemic that the emergence of a new virus can quickly become a global health burden and kill millions of lives. Antiviral drugs are essential in our fight against viral diseases, but most of them are virus-specific and are prone to viral mutations. We have developed broad-spectrum antivirals based on multivalent nanoparticles grafted with ligands that mimic the target of viral attachment ligands (VALs). We have shown that when the ligand has a sufficiently long hydrophobic tail, the inhibition mechanism switches from reversible (virustatic) to irreversible (virucidal). Here, we investigate further how ligand density and particle size affect antiviral efficacy, both in terms of half-inhibitory concentration (IC 50) and of reversible vs irreversible mechanism. We designed antiviral silica nanoparticles modified with 11-mercaptoundecane-1-sulfonic acid (MUS), a ligand that mimics heparan sulfate proteoglycans (HSPG) and we showed that these nanoparticles can be synthesized with different sizes (4–200 nm) and ligand grafting densities (0.59–10.70 /nm2). By testing these particles against herpes simplex virus type 2 (HSV-2), we show that within the size and density ranges studied, the antiviral IC 50 is determined solely by equivalent ligand concentration. The nanoparticles are found to be virucidal at all sizes and densities studied. [ABSTRACT FROM AUTHOR]

Although FOXP3+ regulatory T cells (Treg) depend on IL-2 produced by other cells for their survival and function, the levels of IL-2 in inflamed tissue are low, making it unclear how Treg access this critical resource. Here, we show that Treg use heparanase (HPSE) to access IL-2 sequestered by heparan sulfate (HS) within the extracellular matrix (ECM) of inflamed central nervous system tissue. HPSE expression distinguishes human and murine Treg from conventional T cells and is regulated by the availability of IL-2. HPSE-/- Treg have impaired stability and function in vivo, including in the experimental autoimmune encephalomyelitis (EAE) mouse model of multiple sclerosis. Conversely, endowing monoclonal antibody-directed chimeric antigen receptor (mAbCAR) Treg with HPSE enhances their ability to access HS-sequestered IL-2 and their ability to suppress neuroinflammation in vivo. Together, these data identify a role for HPSE and the ECM in immune tolerance, providing new avenues for improving Treg-based therapy of autoimmunity. Regulatory T cell (Treg) maintenance and function require IL-2, yet this cytokine is only present in low levels in vivo. In this study, the authors demonstrate that that Treg use heparanase to access IL-2 bound to heparan sulfate proteoglycans in the extracellular matrix of inflamed brain tissue in mice. [ABSTRACT FROM AUTHOR]

To identify host factors that affect Bovine Herpes Virus Type 1 (BoHV-1) infection we previously applied a genome wide CRISPR knockout screen targeting all bovine protein coding genes. By doing so we compiled a list of both pro-viral and anti-viral proteins involved in BoHV-1 replication. Here we provide further analysis of those that are potentially involved in viral entry into the host cell. We first generated single cell knockout clones deficient in some of the candidate genes for validation. We provide evidence that Polio Virus Receptor-related protein (PVRL2) serves as a receptor for BoHV-1, mediating more efficient entry than the previously identified Polio Virus Receptor (PVR). By knocking out two enzymes that catalyze HSPG chain elongation, HST2ST1 and GLCE, we further demonstrate the significance of HSPG in BoHV-1 entry. Another intriguing cluster of candidate genes, COG1, COG2 and COG4-7 encode six subunits of the Conserved Oligomeric Golgi (COG) complex. MDBK cells lacking COG6 produced fewer but bigger plaques compared to control cells, suggesting more efficient release of newly produced virions from these COG6 knockout cells, due to impaired HSPG biosynthesis. We further observed that viruses produced by the COG6 knockout cells consist of protein(s) with reduced N-glycosylation, potentially explaining their lower infectivity. To facilitate candidate validation, we also detailed a one-step multiplex CRISPR interference (CRISPRi) system, an orthogonal method to KO that enables quick and simultaneous deployment of three CRISPRs for efficient gene inactivation. Using CRISPR3i, we verified eight candidates that have been implicated in the synthesis of surface heparan sulfate proteoglycans (HSPGs). In summary, our experiments confirmed the two receptors PVR and PVRL2 for BoHV-1 entry into the host cell and other factors that affect this process, likely through the direct or indirect roles they play during HSPG synthesis and glycosylation of viral proteins. [ABSTRACT FROM AUTHOR]

Middle East respiratory syndrome coronavirus (MERS-CoV) is a zoonotic virus with high contagion and mortality rates. Heparan sulfate proteoglycans (HSPGs) are ubiquitously expressed on the surface of mammalian cells. Owing to its high negatively charged property, heparan sulfate (HS) on the surface of host cells is used by many viruses as cofactor to facilitate viral attachment and initiate cellular entry. Therefore, inhibition of the interaction between viruses and HS could be a promising target to inhibit viral infection. In the current study, the interaction between the receptor-binding domain (RBD) of MERS-CoV and heparin was exploited to assess the inhibitory activity of various sulfated glycans such as glycosaminoglycans, marine-sourced glycans (sulfated fucans, fucosylated chondroitin sulfates, fucoidans, and rhamnan sulfate), pentosan polysulfate, and mucopolysaccharide using Surface Plasmon Resonance. We believe this study provides valuable insights for the development of sulfated glycan-based inhibitors as potential antiviral agents. [ABSTRACT FROM AUTHOR]

Persistent high-risk HPV infection is closely associated with cervical cancer development, and there is no drug targeting HPV on the market at present, so it is particularly important to understand the interaction mechanism between HPV and the host which may provide the novel strategies for treating HPV diseases. HPV can hijack cell surface heparan sulfate proteoglycans (HSPGs) as primary receptors. However, the secondary entry receptors for HPV remain elusive. We identify myosin-9 (NMHC-IIA) as a host factor that interacts with HPV L1 protein and mediates HPV internalization. Efficient HPV entry required myosin-9 redistribution to the cell surface regulated by HPV-hijacked MEK-MLCK signaling. Myosin-9 maldistribution by ML-7 or ML-9 significantly inhibited HPV pseudoviruses infection in vitro and in vivo. Meanwhile, N-glycans, especially the galactose chains, may act as the decoy receptors for HPV, which can block the interaction of HPV to myosin-9 and influence the way of HPV infection. Taken together, we identify myosin-9 as a novel functional entry receptor for high-risk HPV both in vitro and in vivo, and unravel the new roles of myosin-9 and N-glycans in HPV entry, which provides the possibilities for host targets of antiviral drugs. [ABSTRACT FROM AUTHOR]

Dpp/BMP acts as a morphogen to provide positional information in the Drosophila wing disc. Key cell-surface molecules to control Dpp morphogen gradient formation and signaling are heparan sulfate proteoglycans (HSPGs). In the wing disc, two HSPGs, the glypicans Division abnormally delayed (Dally) and Dally-like (Dlp) have been suggested to act redundantly to control these processes through direct interaction of their heparan sulfate (HS) chains with Dpp. Based on this assumption, a number of models on how glypicans control Dpp gradient formation and signaling have been proposed, including facilitating or hindering Dpp spreading, stabilizing Dpp on the cell surface, or recycling Dpp. However, how distinct HSPGs act remains largely unknown. Here, we generate genome-engineering platforms for the two glypicans and find that only Dally is critical for Dpp gradient formation and signaling through interaction of its core protein with Dpp. We also find that this interaction is not sufficient and that the HS chains of Dally are essential for these functions largely without interacting with Dpp. We provide evidence that the HS chains of Dally are not essential for spreading or recycling of Dpp but for stabilizing Dpp on the cell surface by antagonizing receptor-mediated Dpp internalization. These results provide new insights into how distinct HSPGs control morphogen gradient formation and signaling during development. [ABSTRACT FROM AUTHOR]

Heparan sulfate proteoglycans (HSPGs) are composed of a core protein and glycosaminoglycan (GAG) chains and serve as coreceptors for many growth factors and morphogens. To understand the molecular mechanisms by which HSPGs regulate morphogen gradient formation and signaling, it is important to determine the relative contributions of the carbohydrate and protein moieties to the proteoglycan function. To address this question, we generated ΔGAG alleles for dally and dally-like protein (dlp), two Drosophila HSPGs of the glypican family, in which all GAG-attachment serine residues are substituted to alanine residues using CRISPR/Cas9 mutagenesis. In these alleles, the glypican core proteins are expressed from the endogenous loci with no GAG modification. Analyses of the dallyΔGAG allele defined Dally functions that do not require heparan sulfate (HS) chains and that need both core protein and HS chains. We found a new, dallyΔGAGspecific phenotype, the formation of a posterior ectopic vein, which we have never seen in the null mutants. Unlike dallyΔGAG, dlpΔGAG mutants do not show most of the dlp null mutant phenotypes, suggesting that HS chains are dispensable for these dlp functions. As an exception, HS is essentially required for Dlp's activity at the neuromuscular junction. Thus, Drosophila glypicans show strikingly different levels of HS dependency. The ΔGAG mutant alleles of the glypicans serve as new molecular genetic toolsets highly useful to address important biological questions, such as molecular mechanisms of morphogen gradient formation. [ABSTRACT FROM AUTHOR]

Neurodegenerative diseases, particularly Alzheimer’s disease (AD), pose a significant challenge in ageing populations. Our current understanding indicates that the onset of toxic amyloid and tau protein pathologies initiates disease progression. However, existing treatments targeting these hallmark symptoms offer symptomatic relief without halting disease advancement. This review offers an alternative perspective on AD, centring on impaired adult hippocampal neurogenesis (AHN) as a potential early aetiological factor. By delving into the intricate molecular events during the initial stages of AD (Braak Stages I–III), a novel hypothesis is presented, interweaving the roles of Notch signalling and heparan sulfate proteoglycans (HSPGs) in compromised AHN. While acknowledging the significance of the amyloid and tau hypotheses, it calls for further exploration beyond these paradigms, suggesting the potential of altered HS sulfation patterns in AD initiation. Future directions propose more detailed investigations into early HS aggregation, aberrant sulfation patterns and examination of their temporal relationship with tau hyperphosphorylation. In challenging the conventional ‘triggers’ of AD and urging their reconsideration as symptoms, this review advocates an alternative approach to understanding this disease, offering new avenues of investigation into the intricacies of AD pathogenesis.

An oral antiviral against SARS-CoV-2 that also attenuates inflammatory instigators of severe COVID-19 is not available to date. Herein, we show that the apoA-I mimetic peptide 4 F inhibits Spike mediated viral entry and has antiviral activity against SARS-CoV-2 in human lung epithelial Calu3 and Vero-E6 cells. In SARS-CoV-2 infected Calu3 cells, 4 F upregulated inducers of the interferon pathway such as MX-1 and Heme oxygenase 1 (HO-1) and downregulated mitochondrial reactive oxygen species (mito-ROS) and CD147, a host protein that mediates viral entry. 4 F also reduced associated cellular apoptosis and secretion of IL-6 in both SARS-CoV-2 infected Vero-E6 and Calu3 cells. Thus, 4 F attenuates in vitro SARS-CoV-2 replication, associated apoptosis in epithelial cells and secretion of IL-6, a major cytokine related to COVID-19 morbidity. Given established safety of 4 F in humans, clinical studies are warranted to establish 4 F as therapy for COVID-19.

Abstract Background Syndecan-4 (SDC4) is a member of the heparan sulfate proteoglycan family of cell-surface receptors. We and others previously reported that variation in the SDC4 gene was associated with several components of the metabolic syndrome, including intra-abdominal fat, fasting glucose and triglyceride levels, and hypertension, in human cohorts. Additionally, we demonstrated that high fat diet (HFD)-induced obese female mice with a Sdc4 genetic deletion had higher visceral adiposity and a worse metabolic profile than control mice. Here, we aimed to first investigate whether the mouse Sdc4 null mutation impacts metabolic phenotypes in a sex- and diet-dependent manner. We then tested whether SDC4 polymorphisms are related to the metabolic syndrome (MetS) in humans. Methods For the mouse experiment, Sdc4-deficient (Sdc4 −/− ) and wild-type (WT) mice were treated with 14-weeks of low-fat diet (LFD). Body composition, energy balance, and selected metabolic phenotypes were assessed. For the human genetic study, we used logistic regression models to test 11 SDC4 SNPs for association with the MetS and its components in a cohort of 274 (113 with MetS) elderly subjects from southern Italy. Results Following the dietary intervention in mice, we observed that the effects of the Sdc4 null mutation on several phenotypes were different from those previously reported in the mice kept on an HFD. Nonetheless, LFD-fed female Sdc4 −/− mice, but not males, displayed higher levels of triglycerides and lower insulin sensitivity at fasting than WT mice, as seen earlier in the HFD conditions. In the parallel human study, we found that carriers of SDC4 rs2228384 allele C and rs2072785 allele T had reduced risk of MetS. The opposite was true for carriers of the SDC4 rs1981429 allele G. Additionally, the SNPs were found related to fasting triglyceride levels and triglyceride glucose (TyG) index, a reliable indicator of insulin resistance, with sex-stratified analysis detecting the association of rs1981429 with these phenotypes only in females. Conclusions Altogether, our results suggest that SDC4 is an evolutionary conserved genetic determinant of MetS and that its genetic variation is associated with fasting triglyceride levels in a female-specific manner.

The human sulfatase HSulf-2 is one of only two known endosulfatases that play a decisive role in modulating the binding properties of heparan sulfate proteoglycans on the cell surface and in the extracellular matrix. Recently, HSulf-2 was shown to exhibit an unusual post-translational modification consisting of a sulfated glycosaminoglycan chain. This study describes the structural characterization of this glycosaminoglycan (GAG) and provides new data on its impact on the catalytic properties of HSulf-2. The unrevealed nature of this GAG chain is identified as a chondroitin/dermatan sulfate (CS/DS) mixed chain, as shown by mass spectrometry combined with NMR analysis. It consists primarily of 6-O and 4-O monosulfated disaccharide units, with a slight predominance of the 4-O-sulfation. Using atomic force microscopy, we show that this unique post-translational modification dramatically impacts the enzyme hydrodynamic volume. We identified human hyaluronidase-4 as a secreted hydrolase that can digest HSulf-2 GAG chain. We also showed that HSulf-2 is able to efficiently 6-O-desulfate antithrombin III binding pentasaccharide motif, and that this activity was enhanced upon removal of the GAG chain. Finally, we identified five N-glycosylation sites on the protein and showed that, although required, reduced N-glycosylation profiles were sufficient to sustain HSulf-2 integrity. [ABSTRACT FROM AUTHOR]

Why apolipoprotein AV (APOA5) deficiency causes hypertriglyceridemia has remained unclear, but we have suspected that the underlying cause is reduced amounts of lipoprotein lipase (LPL) in capillaries. By routine immunohistochemistry, we observed reduced LPL staining of heart and brown adipose tissue (BAT) capillaries in Apoa5–/– mice. Also, after an intravenous injection of LPL-, CD31-, and GPIHBP1-specific mAbs, the binding of LPL Abs to heart and BAT capillaries (relative to CD31 or GPIHBP1 Abs) was reduced in Apoa5–/– mice. LPL levels in the postheparin plasma were also lower in Apoa5–/– mice. We suspected that a recent biochemical observation — that APOA5 binds to the ANGPTL3/8 complex and suppresses its capacity to inhibit LPL catalytic activity — could be related to the low intracapillary LPL levels in Apoa5–/– mice. We showed that an ANGPTL3/8-specific mAb (IBA490) and APOA5 normalized plasma triglyceride (TG) levels and intracapillary LPL levels in Apoa5–/– mice. We also showed that ANGPTL3/8 detached LPL from heparan sulfate proteoglycans and GPIHBP1 on the surface of cells and that the LPL detachment was blocked by IBA490 and APOA5. Our studies explain the hypertriglyceridemia in Apoa5–/– mice and further illuminate the molecular mechanisms that regulate plasma TG metabolism. [ABSTRACT FROM AUTHOR]

Hepatocellular carcinoma (HCC) is a highly heterogeneous malignancy and lacks effective treatment. Bulksequencing of different gene transcripts by comparing HCC tissues and adjacent normal tissues provides some clues for investigating the mechanisms or identifying potential targets for tumor progression. However, genes that are exclusively expressed in a subpopulation of HCC may not be enriched or detected through such a screening. In the current study, we performed a single cell-clone-based screening and identified galectin-14 as an essential molecule in the regulation of tumor growth. The aberrant expression of galectin-14 was significantly associated with a poor overall survival of liver cancer patients with database analysis. Knocking down galectin-14 inhibited the proliferation of tumor growth, whereas overexpressing galectin-14 promoted tumor growth in vivo. Nontargeted metabolomics analysis indicated that knocking down galectin-14 decreased glycometabolism; specifically that glycoside synthesis was significantly changed. Further study found that galectin-14 promoted the expression of cell surface heparan sulfate proteoglycans (HSPGs) that functioned as co-receptors, thereby increasing the responsiveness of HCC cells to growth factors, such as epidermal growth factor and transforming growth factoralpha. In conclusion, the current study identifies a novel HCC-specific molecule galectin-14, which increases the expression of cell surface HSPGs and the uptake of growth factors to promote HCC cell proliferation. [ABSTRACT FROM AUTHOR]

Neovascularization is a key feature of ischemic retinal diseases and the wet form of age-related macular degeneration (AMD), all leading causes of severe vision loss. Vascular endothelial growth factor (VEGF) inhibitors have transformed the treatment of these disorders. Millions of patients have been treated with these drugs worldwide. However, in real-life clinical settings, many patients do not experience the same degree of benefit observed in clinical trials, in part because they receive fewer anti-VEGF injections. Therefore, there is an urgent need to discover and identify novel long-acting VEGF inhibitors. We hypothesized that binding to heparan-sulfate proteoglycans (HSPG) in the vitreous, and possibly other ocular structures, may be a strategy to promote intraocular retention, ultimately leading to a reduced burden of intravitreal injections. We designed a series of VEGF receptor 1 variants and identified some with strong heparin-binding characteristics and ability to bind to vitreous matrix. Our data indicate that some of our variants have longer duration and greater efficacy in animal models of intraocular neovascularization than current standard of care. Our study represents a systematic attempt to exploit the functional diversity associated with heparin affinity of a VEGF receptor.

Heparan sulfate proteoglycans consist of a small family of proteins decorated with one or more covalently attached heparan sulfate glycosaminoglycan chains. These chains have intricate structural patterns based on the position of sulfate groups and uronic acid epimers, which dictate their ability to engage a large repertoire of heparan sulfate-binding proteins, including extracellular matrix proteins, growth factors and morphogens, cytokines and chemokines, apolipoproteins and lipases, adhesion and growth factor receptors, and components of the complement and coagulation system. This review highlights recent progress in the characterization of the so-called heparan sulfate interactome, with a major focus on systems-wide strategies as a tool for discovery and characterization of this subproteome. In addition, we compiled all heparan sulfate-binding proteins reported in the literature to date and grouped them into a few major functional classes by applying a networking approach.

Heparan sulfate proteoglycans (HSPGs) surround the surface of odontoblasts, and their modification affects their affinity for Wnt ligands. This study proposes applying Matching Transformation System® (MA-T), a novel chlorinated oxidant, to enhance dentinogenesis. MA-T treatment in odontoblasts decreased sulfation of HSPG and upregulated the expression of dentin sialophosphoprotein (Dspp) and Dentin Matrix Protein 1 (Dmp1) via activation of canonical Wnt signaling in vitro. Ex vivo application of MA-T also enhanced dentin matrix formation in developing tooth explants. Reanalysis of a public single-cell RNA-seq dataset revealed significant Wnt activity in the odontoblast population, with enrichment for Wnt10a and Wnt6. Silencing assays showed that Wnt10a and Wnt6 were redundant in inducing Dspp and Dmp1 mRNA expression. These Wnt ligands’ expression was upregulated by MA-T treatment, and TCF/LEF binding sites are present in their promoters. Furthermore, the Wnt inhibitors Notum and Dkk1 were enriched in odontoblasts, and their expression was also upregulated by MA-T treatment, together suggesting autonomous maintenance of Wnt signaling in odontoblasts. This study provides evidence that MA-T activates dentinogenesis by modifying HSPG and through subsequent activation of Wnt signaling.

Parkinson's disease (PD) is the second-most common neurodegenerative disease and is largely caused by the death of dopaminergic (DA) cells. Dopamine loss occurs in the substantia nigra pars compacta and leads to dysfunctions in motor functions. Death of DA cells can occur with oxidative stress and dysfunction of glial cells caused by Parkinson-related gene mutations. Lactoferrin (Lf) is a multifunctional glycoprotein that is usually known for its presence in milk, but recent research shows that Lf is also found in the brain regions. 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) is a known mitochondrial toxin that disturbs themitochondrial electron transport chain (ETC) systemand increases the rate of reactive oxygen species. Lf's high affinity for metals decreases the required iron for the Fenton reaction, reduces the oxidative damage to DA cells caused by MPTP, and increases their surveillance rate. Several studies also investigated Lf's effect on neurons that are treated with MPTP. The results pointed out that Lf's protective effect can also be observed without the presence of oxidative stress; thus, several potential mechanisms are currently being researched, starting with a potential HSPG-Lf interaction in the cellular membrane of DA cells. The presence of Lf activity in the brain region also showed that lactoferrin initiates receptor-mediated transcytosis in the blood-brain barrier (BBB) with the existence of lactoferrin receptors in the endothelial cells. The existence of Lf receptors both in endothelial cells and DA cells created the idea of using Lf as a secondary molecule in the transport of therapeutic agents across the BBB, especially in nanoparticle development. [ABSTRACT FROM AUTHOR]

SARS-CoV-2 variants evolve to rely more on heparan sulfate (HS) for viral attachment and subsequent infection. In our earlier work, we demonstrated that the Delta variant's spike protein binds more strongly to HS compared to WT SARS-CoV-2, leading to enhanced cell internalization via syndecans (SDCs), a family of transmembrane HS proteoglycans (HSPGs) facilitating the cellular entry of the original strain. Using our previously established ACE2- or SDC-overexpressing cellular models, we now compare the ACE2- and SDC-dependent cellular uptake of heat-inactivated WT SARS-CoV-2 with the Delta and Omicron variants. Internalization studies with inactivated virus particles showed that ACE2 overexpression could not compensate for the loss of HS in Omicron's internalization, suggesting that this variant primarily uses HSPGs to enter cells. Although SDCs increased the internalization of all three viruses, subtle differences could be detected between their SDC isoform preferences. The Delta variant particularly benefitted from SDC1, 2, and 4 overexpression for cellular entry, while SDC4 had the most prominent effect on Omicron internalization. The SDC4 knockdown (KD) in Calu-3 cells reduced the cellular uptake of all three viruses, but the inhibition was the most pronounced for Omicron. The polyanionic heparin also hindered the cellular internalization of all three viruses with a dominant inhibitory effect on Omicron. Omicron's predominant HSPG affinity, combined with its preference for the universally expressed SDC4, might account for its efficient transmission yet reduced pathogenicity. [ABSTRACT FROM AUTHOR]

Secreted modular calcium-binding proteins (SMOCs) are conserved matricellular proteins found in organisms from Caenorhabditis elegans to humans. SMOC homologs characteristically contain 1 or 2 extracellular calcium (EC)-binding domain(s) and 1 or 2 thyroglobulin type-1 (TY) domain(s). SMOC proteins in Drosophila and Xenopus have been found to interact with cell surface heparan sulfate proteoglycans (HSPGs) to exert both positive and negative influences on the conserved bone morphogenetic protein (BMP) signaling pathway. In this study, we used a combination of biochemical, structural modeling, and molecular genetic approaches to dissect the functions of the sole SMOC protein in C. elegans. We showed that CeSMOC-1 binds to the heparin sulfate proteoglycan GPC3 homolog LON-2/glypican, as well as the mature domain of the BMP2/4 homolog DBL-1. Moreover, CeSMOC-1 can simultaneously bind LON-2/glypican and DBL-1/BMP. The interaction between CeSMOC-1 and LON-2/glypican is mediated specifically by the EC domain of CeSMOC-1, while the full interaction between CeSMOC-1 and DBL-1/BMP requires full-length CeSMOC-1. We provide both in vitro biochemical and in vivo functional evidence demonstrating that CeSMOC-1 functions both negatively in a LON-2/glypican-dependent manner and positively in a DBL-1/BMP-dependent manner to regulate BMP signaling. We further showed that in silico, Drosophila and vertebrate SMOC proteins can also bind to mature BMP dimers. Our work provides a mechanistic basis for how the evolutionarily conserved SMOC proteins regulate BMP signaling. Cell-cell signaling needs to be precisely controlled to ensure proper development and homeostasis. This study shows that the conserved secreted modular calcium binding protein, SMOC, regulates bone morphogenetic protein (BMP) signaling in the nematode C. elegans by binding to both BMP and glypican. [ABSTRACT FROM AUTHOR]

Extracellular matrix (ECM) plays a pivotal and dynamic role in the construction of tumor microenvironment (TME), becoming the focus in cancer research and treatment. Multiple cell signaling in ECM remodeling contribute to uncontrolled proliferation, metastasis, immune evasion and drug resistance of cancer. Targeting trilogy of ECM remodeling could be a new strategy during the early-, middle-, advanced-stages of cancer and overcoming drug resistance. Currently nearly 60% of the alternative anticancer drugs are derived from natural products or active ingredients or structural analogs isolated from plants. According to the characteristics of ECM, this manuscript proposes three phases of whole-process management of cancer, including prevention of cancer development in the early stage of cancer (Phase I); prevent the metastasis of tumor in the middle stage of cancer (Phase II); provide a novel method in the use of immunotherapy for advanced cancer (Phase III), and present novel insights on the contribution of natural products use as innovative strategies to exert anticancer effects by targeting components in ECM. Herein, we focus on trilogy of ECM remodeling and the interaction among ECM, cancer-associated fibroblasts (CAFs) and tumor-associated macrophages (TAMs), and sort out the intervention effects of natural products on the ECM and related targets in the tumor progression, provide a reference for the development of new drugs against tumor metastasis and recurrence. [ABSTRACT FROM AUTHOR]

Background: Syndecan-4 (SDC4) is a member of the heparan sulfate proteoglycan family of cell-surface receptors. We and others previously reported that variation in the SDC4 gene was associated with several components of the metabolic syndrome, including intra-abdominal fat, fasting glucose and triglyceride levels, and hypertension, in human cohorts. Additionally, we demonstrated that high fat diet (HFD)-induced obese female mice with a Sdc4 genetic deletion had higher visceral adiposity and a worse metabolic profile than control mice. Here, we aimed to first investigate whether the mouse Sdc4 null mutation impacts metabolic phenotypes in a sex- and diet-dependent manner. We then tested whether SDC4 polymorphisms are related to the metabolic syndrome (MetS) in humans. Methods: For the mouse experiment, Sdc4-deficient (Sdc4−/−) and wild-type (WT) mice were treated with 14-weeks of low-fat diet (LFD). Body composition, energy balance, and selected metabolic phenotypes were assessed. For the human genetic study, we used logistic regression models to test 11 SDC4 SNPs for association with the MetS and its components in a cohort of 274 (113 with MetS) elderly subjects from southern Italy. Results: Following the dietary intervention in mice, we observed that the effects of the Sdc4 null mutation on several phenotypes were different from those previously reported in the mice kept on an HFD. Nonetheless, LFD-fed female Sdc4−/− mice, but not males, displayed higher levels of triglycerides and lower insulin sensitivity at fasting than WT mice, as seen earlier in the HFD conditions. In the parallel human study, we found that carriers of SDC4 rs2228384 allele C and rs2072785 allele T had reduced risk of MetS. The opposite was true for carriers of the SDC4 rs1981429 allele G. Additionally, the SNPs were found related to fasting triglyceride levels and triglyceride glucose (TyG) index, a reliable indicator of insulin resistance, with sex-stratified analysis detecting the association of rs1981429 with these phenotypes only in females. Conclusions: Altogether, our results suggest that SDC4 is an evolutionary conserved genetic determinant of MetS and that its genetic variation is associated with fasting triglyceride levels in a female-specific manner. [ABSTRACT FROM AUTHOR]

In Australia, 13% of women are diagnosed with breast cancer (BC) in their lifetime with approximately 20,000 women diagnosed with the disease in 2021. BC is characterised by complex histological and genomic influences with recent advances in cancer biology improving early diagnosis and personalised treatment interventions. The Phosphatidyl-inositol-3-kinase/Protein kinase B (PI3K/AKT) pathway is essential in apoptosis resistance, cell survival, activation of cellular responses to DNA damage and DNA repair. Heparan sulfate proteoglycans (HSPGs) are ubiquitous molecules found on the cell surface and in the extracellular matrix with essential functions in regulating cell survival, growth, adhesion and as mediators of cell differentiation and migration. HSPGs, particularly the syndecans (SDCs), have been linked to cancers, making them an exciting target for anticancer treatments. In the PI3K/AKT pathway, syndecan-4 (SDC4) has been shown to downregulate AKT Serine/Threonine Kinase (AKT1) gene expression, while the ATM Serine/Threonine Kinase (ATM) gene has been found to inhibit this pathway upstream of AKT. We investigated single-nucleotide polymorphisms (SNPs) in HSPG and related genes SDC4, AKT1 and ATM and their influence on the prevalence of BC. SNPs were genotyped in the Australian Caucasian Genomics Research Centre Breast Cancer (GRC-BC) population and in the Griffith University–Cancer Council Queensland Breast Cancer Biobank (GU-CCQ BB) population. We identified that SDC4-rs1981429 and ATM-rs228590 may influence the development and progression of BC, having the potential to become biomarkers in early BC diagnosis and personalised treatment. [ABSTRACT FROM AUTHOR]

The glomerular basement membrane (GBM) is an important tissue structure in kidney function. It is the membrane through which filtrate and solutes must pass to reach the nephron tubules. This review focuses on the spatial location of the main extracellular matrix components of the GBM. It also attempts to explain this organization in terms of their synthesis, transport, and loss. The picture that emerges is that the collagen IV and laminin content of GBM are in a very slow dynamic disequilibrium, leading to GBM thickening with age, and in contrast, some heparan sulfate proteoglycans are in a dynamic equilibrium with a very rapid turnover (i.e. half-life measured in ~hours) and flow direction against the flow of filtrate. The highly rapid heparan sulfate turnover may serve several roles, including an unclogging mechanism for the GBM, compressive stiffness of the GBM fiber network, and/or enabling podocycte-endothelial crosstalk against the flow of filtrate. [ABSTRACT FROM AUTHOR]

Proteoglycans are a specific subset of glycoproteins found at the cell surface and in the extracellular matrix, where they interact with a plethora of proteins involved in metabolic homeostasis and meta-inflammation. Over the last decade, new insights have emerged on the mechanism and biological significance of these interactions in the context of diet-induced disorders such as obesity and type-2 diabetes. Complications of energy metabolism drive most diet-induced metabolic disorders, which results in low-grade chronic inflammation, thereby affecting proper function of many vital organs involved in energy homeostasis, such as the brain, liver, kidney, heart and adipose tissue. Here, we discuss how heparan, chondroitin and keratan sulfate proteoglycans modulate obesity-induced metabolic dysfunction and low-grade inflammation that impact the initiation and progression of obesity-associated morbidities.

Heparan sulfate proteoglycans (HSPGs) form essential components of the extracellular matrix (ECM) and basement membrane (BM) and have both structural and signaling roles. Perlecan is a secreted ECM-localized HSPG that contributes to tissue integrity and cell-cell communication. Although a core component of the ECM, the role of Perlecan in neuronal structure and function is less understood. Here, we identify a role for Drosophila Perlecan in the maintenance of larval motoneuron axonal and synaptic stability. Loss of Perlecan causes alterations in the axonal cytoskeleton, followed by axonal breakage and synaptic retraction of neuromuscular junctions. These phenotypes are not prevented by blocking Wallerian degeneration and are independent of Perlecan's role in Wingless signaling. Expression of Perlecan solely in motoneurons cannot rescue synaptic retraction phenotypes. Similarly, removing Perlecan specifically from neurons, glia, or muscle does not cause synaptic retraction, indicating the protein is secreted from multiple cell types and functions non-cell autonomously. Within the peripheral nervous system, Perlecan predominantly localizes to the neural lamella, a specialized ECM surrounding nerve bundles. Indeed, the neural lamella is disrupted in the absence of Perlecan, with axons occasionally exiting their usual boundary in the nerve bundle. In addition, entire nerve bundles degenerate in a temporally coordinated manner across individual hemi-segments throughout larval development. These observations indicate disruption of neural lamella ECM function triggers axonal destabilization and synaptic retraction of motoneurons, revealing a role for Perlecan in axonal and synaptic integrity during nervous system development. [ABSTRACT FROM AUTHOR]

Gastric cancer is a dominating cause of cancer-associated mortality with limited therapeutic options. Here, we show that syndecan-4 (SDC4), a transmembrane proteoglycan, is highly expressed in intestinal subtype gastric tumors and that this signature associates with patient poor survival. Further, we mechanistically demonstrate that SDC4 is a master regulator of gastric cancer cell motility and invasion. We also find that SDC4 decorated with heparan sulfate is efficiently sorted in extracellular vesicles (EVs). Interestingly, SDC4 in EVs regulates gastric cancer cell-derived EV organ distribution, uptake, and functional effects in recipient cells. Specifically, we show that SDC4 knockout disrupts the tropism of EVs for the common gastric cancer metastatic sites. Our findings set the basis for the molecular implications of SDC4 expression in gastric cancer cells and provide broader perspectives on the development of therapeutic strategies targeting the glycan-EV axis to limit tumor progression. [ABSTRACT FROM AUTHOR]

Enterovirus A71 (EV-A71) can elicit a wide variety of human diseases such as hand, foot, and mouth disease and severe or fatal neurological complications. It is not clearly understood what determines the virulence and fitness of EV-A71. It has been observed that amino acid changes in the receptor binding protein, VP1, resulting in viral binding to heparan sulfate proteoglycans (HSPGs) may be important for the ability of EV-A71 to infect neuronal tissue. In this study, we identified that the presence of glutamine, as opposed to glutamic acid, at VP1-145 is key for viral infection in a 2D human fetal intestinal model, consistent with previous findings in an airway organoid model. Moreover, pre-treatment of EV-A71 particles with low molecular weight heparin to block HSPG-binding significantly reduced the infectivity of two clinical EV-A71 isolates and viral mutants carrying glutamine at VP1-145. Our data indicates that mutations in VP1 leading to HSPG-binding enhances viral replication in the human gut. These mutations resulting in increased production of viral particles at the primary replication site could lead to a higher risk of subsequent neuroinfection. Importance: With the near eradication of polio worldwide, polio-like illness (as is increasingly caused by EV-A71 infections) is of emerging concern. EV-A71 is indeed the most neurotropic enterovirus that poses a major threat globally to public health and specifically in infants and young children. Our findings will contribute to the understanding of the virulence and the pathogenicity of this virus. Further, our data also supports the identification of potential therapeutic targets against severe EVA71 infection especially among infants and young children. Furthermore, our work highlights the key role of HSPG-binding mutations in the disease outcome of EVA71. Additionally, EV-A71 is not able to infect the gut (the primary replication site in humans) in traditionally used animal models. Thus, our research highlights the need for human-based models to study human viral infections. [ABSTRACT FROM AUTHOR]