PDZ scaffolds regulate extracellular vesicle production, composition, and uptake.

Extracellular vesicles (EVs) are membrane-limited organelles mediating cell-to-cell communication in health and disease. EVs are of high medical interest, but their rational use for diagnostics or therapies is restricted by our limited understanding of the molecular mechanisms governing EV biology. Here, we tested whether PDZ proteins, molecular scaffolds that support the formation, transport, and function of signal transduction complexes and that coevolved with multicellularity, may represent important EV regulators. We reveal that the PDZ proteome (ca. 150 proteins in human) establishes a discrete number of direct interactions with the tetraspanins CD9, CD63, and CD81, well-known EV constituents. Strikingly, PDZ proteins interact more extensively with syndecans (SDCs), ubiquitous membrane proteins for which we previously demonstrated an important role in EV biogenesis, loading, and turnover. Nine PDZ proteins were tested in loss-of-function studies. We document that these PDZ proteins regulate both tetraspanins and SDCs, differentially affecting their steady-state levels, subcellular localizations, metabolism, endosomal budding, and accumulations in EVs. Importantly, we also show that PDZ proteins control the levels of heparan sulfate at the cell surface that functions in EV capture. In conclusion, our study establishes that the extensive networking of SDCs, tetraspanins, and PDZ proteins contributes to EV heterogeneity and turnover, highlighting an important piece of the molecular framework governing intracellular trafficking and intercellular communication.

Mutual Inhibition of Antithrombin III and SARS-CoV-2 Cellular Attachment to Syndecans: Implications for COVID-19 Treatment and Vaccination.

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.

Syndecan-4 Mediates the Cellular Entry of Adeno-Associated Virus 9.

Due to their low pathogenicity, immunogenicity, and long-term gene expression, adeno-associated virus (AAV) vectors emerged as safe and efficient gene delivery tools, over-coming setbacks experienced with other viral gene delivery systems in early gene therapy trials. Among AAVs, AAV9 can translocate through the blood-brain barrier (BBB), making it a promising gene delivery tool for transducing the central nervous system (CNS) via systemic administration. Recent reports on the shortcomings of AAV9-mediated gene delivery into the CNS require reviewing the molecular base of AAV9 cellular biology. A more detailed understanding of AAV9's cellular entry would eradicate current hurdles and enable more efficient AAV9-based gene therapy approaches. Syndecans, the transmembrane family of heparan-sulfate proteoglycans, facilitate the cellular uptake of various viruses and drug delivery systems. Utilizing human cell lines and syndecan-specific cellular assays, we assessed the involvement of syndecans in AAV9's cellular entry. The ubiquitously expressed isoform, syndecan-4 proved its superiority in facilitating AAV9 internalization among syndecans. Introducing syndecan-4 into poorly transducible cell lines enabled robust AAV9-dependent gene transduction, while its knockdown reduced AAV9's cellular entry. Attachment of AAV9 to syndecan-4 is mediated not just by the polyanionic heparan-sulfate chains but also by the cell-binding domain of the extracellular syndecan-4 core protein. Co-immunoprecipitation assays and affinity proteomics also confirmed the role of syndecan-4 in the cellular entry of AAV9. Overall, our findings highlight the universally expressed syndecan-4 as a significant contributor to the cellular internalization of AAV9 and provide a molecular-based, rational explanation for the low gene delivery potential of AAV9 into the CNS.

The microRNA-cell surface proteoglycan axis in cancer progression.

Proteoglycans consist one of the major extracellular matrix class of biomolecules that demonstrate nodal roles in cancer progression. Modern diagnostic and therapeutic approaches include proteoglycan detection and pharmacological targeting in various cancer types. Proteoglycans orchestrate critical signaling pathways for cancer development and progression through dynamic interactions with matrix components. It is well established that the epigenetic signatures of cancer cells play critical role in guiding their functional properties and metastatic potential. Secreted microRNAs (miRNAs) reside in a complex network with matrix proteoglycans, thus affecting cell-cell and cell-matrix communication. This mini-review aims to highlight current knowledge on the cell-surface proteoglycan-mediated signaling cascades that regulate miRNA biogenesis in cancer. Moreover, the miRNA-mediated proteoglycan regulation during cancer progression and mechanistic aspects on the way that proteoglycans affect miRNA expression are presented. Recent advances on the role of cell surface proteoglycans in exosome biogenesis and miRNA packaging and expression are also discussed.

Paradoxical effects of osteoprotegerin on vascular function: inhibiting inflammation while promoting oxidative stress?

Osteoprotegerin (OPG), also known as osteoclastogenesis inhibitory factor or tumor necrosis factor receptor superfamily member 11B, is well known as a modulator of bone remodeling. The contribution of OPG to cardiovascular disease (CVD) has been suggested, but its molecular mechanism is complex and remains unclear. In the present study, Alves-Lopes et al. (Clin. Sci. (Lond.) (2021) 135(20): https://doi.org/10.1042/CS20210643) reported the critical role of syndecan-1 (SDC-1, also known as CD138), a surface protein part of the endothelial glycocalyx, in OPG-induced vascular dysfunction. The authors found that in endothelial cells (ECs), through SDC-1, OPG increased eNOS Thr495 phosphorylation, thereby inhibiting eNOS activity. Furthermore, the OPG-SDC-1 interaction increased reactive oxygen species (ROS) production through NOX1/4 activation. Both the reduced eNOS activity and induced ROS production inhibited NO production and impaired EC function. In vascular smooth muscle cells (VSMCs), the OPG-SDC-1 interaction increased ROS production through NOX1/4 activation, subsequently increased MLC phosphorylation-mediated Rho kinase-MYPT1 regulation, leading to increased vascular contraction. Ultilizing wire myography and mechanistic studies, the authors nicely provide the evidence that SDC-1 plays a crucial role in OPG-induced vascular dysfunction. As we mentioned above, the molecular mechanism and roles of OPG in cardiovascular system are complex and somewhat confusing. In this commentary, we briefly summarize the OPG-mediated signaling pathways in cardiovascular system.

Pathological Angiogenesis Requires Syndecan-4 for Efficient VEGFA-Induced VE-Cadherin Internalization.

[Figure: see text].

Biodistribution and Cellular Internalization of Inactivated SARS-CoV-2 in Wild-Type Mice.

Despite the growing list of identified SARS-CoV-2 receptors, the human angiotensin-converting enzyme 2 (ACE2) is still viewed as the main cell entry receptor mediating SARS-CoV-2 internalization. It has been reported that wild-type mice, like other rodent species of the Muridae family, cannot be infected with SARS-CoV-2 due to differences in their ACE2 receptors. On the other hand, the consensus heparin-binding motif of SARS-CoV-2's spike protein, PRRAR, enables the attachment to rodent heparan sulfate proteoglycans (HSPGs), including syndecans, a transmembrane HSPG family with a well-established role in clathrin- and caveolin-independent endocytosis. As mammalian syndecans possess a relatively conserved structure, we analyzed the cellular uptake of inactivated SARS-CoV-2 particles in in vitro and in vivo mice models. Cellular studies revealed efficient uptake into murine cell lines with established syndecan-4 expression. After intravenous administration, inactivated SARS-CoV-2 was taken up by several organs in vivo and could also be detected in the brain. Internalized by various tissues, inactivated SARS-CoV-2 raised tissue TNF-α levels, especially in the heart, reflecting the onset of inflammation. Our studies on in vitro and in vivo mice models thus shed light on unknown details of SARS-CoV-2 internalization and help broaden the understanding of the molecular interactions of SARS-CoV-2.

Proteoglycans: Systems-Level Insight into Their Expression in Healthy and Diseased Placentas.

Proteoglycan macromolecules play key roles in several physiological processes (e.g., adhesion, proliferation, migration, invasion, angiogenesis, and apoptosis), all of which are important for placentation and healthy pregnancy. However, their precise roles in human reproduction have not been clarified. To fill this gap, herein, we provide an overview of the proteoglycans' expression and role in the placenta, in trophoblast development, and in pregnancy complications (pre-eclampsia, fetal growth restriction), highlighting one of the most important members of this family, syndecan-1 (SDC1). Microarray data analysis showed that of 34 placentally expressed proteoglycans, SDC1 production is markedly the highest in the placenta and that SDC1 is the most upregulated gene during trophoblast differentiation into the syncytiotrophoblast. Furthermore, placental transcriptomic data identified dysregulated proteoglycan genes in pre-eclampsia and in fetal growth restriction, including SDC1, which is supported by the lower concentration of syndecan-1 in maternal blood in these syndromes. Overall, our clinical and in vitro studies, data analyses, and literature search pointed out that proteoglycans, as important components of the placenta, may regulate various stages of placental development and participate in the maintenance of a healthy pregnancy. Moreover, syndecan-1 may serve as a useful marker of syncytialization and a prognostic marker of adverse pregnancy outcomes. Further studies are warranted to explore the role of proteoglycans in healthy and complicated pregnancies, which may help in diagnostic or therapeutic developments.

Hyperglycemia-induced effects on glycocalyx components in the retina.

PURPOSE: Diabetic retinopathy is a vision-threatening complication of diabetes characterized by endothelial injury and vascular dysfunction. The loss of the endothelial glycocalyx, a dynamic layer lining all endothelial cells, contributes to several microvascular pathologies, including an increase in vascular permeability, leukocyte plugging, and capillary occlusion, and may drive the progression of retinopathy. Previously, a significant decrease in glycocalyx thickness has been observed in diabetic retinas. However, the effects of diabetes on specific components of the retinal glycocalyx have not yet been studied. Therefore, the aim of our study was to investigate changes in synthesis, expression, and shedding of retinal glycocalyx components induced by hyperglycemia, which could provide a novel therapeutic target for diabetic retinopathy. METHODS: Primary rat retinal microvascular endothelial cells (RRMECs) were grown under normal glucose (5 mM) or high-glucose (25 mM) conditions for 6 days. The mRNA and protein levels of the glycocalyx components were examined using qRT-PCR and Western blot analysis, respectively. Further, mass spectrometry was used to analyze protein intensities of core proteins. In addition, the streptozotocin-induced Type 1 diabetic rat model was used to study changes in the expression of the retinal glycocalyx in vivo. The shedding of the glycocalyx was studied in both culture medium and in plasma using Western blot analysis. RESULTS: A significant increase in the shedding of syndecan-1 and CD44 was observed both in vitro and in vivo under high-glucose conditions. The mRNA levels of syndecan-3 were significantly lower in the RRMECs grown under high glucose conditions, whereas those of syndecan-1, syndecan-2, syndecan-4, glypican-1, glypican-3, and CD44 were significantly higher. The protein expression of syndecan-3 and glypican-1 in RRMECs was reduced considerably following exposure to high glucose, whereas that of syndecan-1 and CD44 increased significantly. In addition, mass spectrometry data also suggests a significant increase in syndecan-4 and a significant decrease in glypican-3 protein levels with high glucose stimulation. In vivo, our data also suggest a significant decrease in the mRNA transcripts of syndecan-3 and an increase in mRNA levels of glypican-1 and CD44 in the retinas of diabetic rats. The diabetic rats exhibited a significant reduction in the retinal expression of syndecan-3 and CD44. However, the expression of syndecan-1 and glypican-1 increased significantly in the diabetic retina. CONCLUSIONS: One of the main findings of our study was the considerable diversity of glucose-induced changes in expression and shedding of various components of endothelial glycocalyx, for example, increased endothelial and retinal syndecan-1, but decreased endothelial and retinal syndecan-3. This indicates that the reported decrease in the retinal glycocalyx in diabetes in not a result of a non-specific shedding mechanism. Moreover, mRNA measurements indicated a similar diversity, with increases in endothelial and/or retinal levels of syndecan-1, glypican-1, and CD44, but a decrease for syndecan-3, with these increases in mRNA potentially a compensatory reaction to the overall loss of glycocalyx.

An investigation of genetic polymorphisms in heparan sulfate proteoglycan core proteins and key modification enzymes in an Australian Caucasian multiple sclerosis population.

Multiple sclerosis (MS) is a chronic inflammatory demyelinating disease affecting the central nervous system in young adults. Heparan sulfate proteoglycans (HSPGs) are ubiquitous to the cell surface and the extracellular matrix. HSPG biosynthesis is a complex process involving enzymatic attachment of heparan sulfate (HS) chains to a core protein. HS side chains mediate specific ligand and growth factor interactions directing cellular processes including cell adhesion, migration and differentiation. Two main families of HSPGs exist, the syndecans (SDC1-4) and glypicans (GPC1-6). The SDCs are transmembrane proteins, while the GPC family are GPI linked to the cell surface. SDC1 has well-documented interactions with numerous signalling pathways. Genome-wide association studies (GWAS) have identified regions of the genome associated with MS including a region on chromosome 13 containing GPC5 and GPC6. International studies have revealed significant associations between this region and disease development. The exostosin-1 (EXT1) and sulfatase-1 (SULF1) are key enzymes contributing to the generation of HS chains. EXT1, with documented tumour suppressor properties, is involved in the initiation and polymerisation of the growing HS chain. SULF1 removes 6-O-sulfate groups from HS chains, affecting protein-ligand interactions and subsequent downstream signalling with HS modification potentially having significant effects on MS progression. In this study, we identified significant associations between single nucleotide polymorphisms in SDC1, GPC5 and GPC6 and MS in an Australian Caucasian case-control population. Further significant associations in these genes were identified when the population was stratified by sex and disease subtype. No association was found for EXT1 or SULF1.

Osteoprotegerin regulates vascular function through syndecan-1 and NADPH oxidase-derived reactive oxygen species.

Osteogenic factors, such as osteoprotegerin (OPG), are protective against vascular calcification. However, OPG is also positively associated with cardiovascular damage, particularly in pulmonary hypertension, possibly through processes beyond effects on calcification. In the present study, we focused on calcification-independent vascular effects of OPG through activation of syndecan-1 and NADPH oxidases (Noxs) 1 and 4. Isolated resistance arteries from Wistar-Kyoto (WKY) rats, exposed to exogenous OPG, studied by myography exhibited endothelial and smooth muscle dysfunction. OPG decreased nitric oxide (NO) production, eNOS activation and increased reactive oxygen species (ROS) production in endothelial cells. In VSMCs, OPG increased ROS production, H2O2/peroxynitrite levels and activation of Rho kinase and myosin light chain. OPG vascular and redox effects were also inhibited by the syndecan-1 inhibitor synstatin (SSNT). Additionally, heparinase and chondroitinase abolished OPG effects on VSMCs-ROS production, confirming syndecan-1 as OPG molecular partner and suggesting that OPG binds to heparan/chondroitin sulphate chains of syndecan-1. OPG-induced ROS production was abrogated by NoxA1ds (Nox1 inhibitor) and GKT137831 (dual Nox1/Nox4 inhibitor). Tempol (SOD mimetic) inhibited vascular dysfunction induced by OPG. In addition, we studied arteries from Nox1 and Nox4 knockout (KO) mice. Nox1 and Nox4 KO abrogated OPG-induced vascular dysfunction. Vascular dysfunction elicited by OPG is mediated by a complex signalling cascade involving syndecan-1, Nox1 and Nox4. Our data identify novel molecular mechanisms beyond calcification for OPG, which may underlie vascular injurious effects of osteogenic factors in conditions such as hypertension and/or diabetes.

The Interplay of Apoes with Syndecans in Influencing Key Cellular Events of Amyloid Pathology.

Apolipoprotein E (ApoE) isoforms exert intricate effects on cellular physiology beyond lipid transport and metabolism. ApoEs influence the onset of Alzheimer's disease (AD) in an isoform-dependent manner: ApoE4 increases AD risk, while ApoE2 decreases it. Previously we demonstrated that syndecans, a transmembrane proteoglycan family with increased expression in AD, trigger the aggregation and modulate the cellular uptake of amyloid beta (Aß). Utilizing our previously established syndecan-overexpressing cellular assays, we now explore how the interplay of ApoEs with syndecans contributes to key events, namely uptake and aggregation, in Aß pathology. The interaction of ApoEs with syndecans indicates isoform-specific characteristics arising beyond the frequently studied ApoE-heparan sulfate interactions. Syndecans, and among them the neuronal syndecan-3, increased the cellular uptake of ApoEs, especially ApoE2 and ApoE3, while ApoEs exerted opposing effects on syndecan-3-mediated Aß uptake and aggregation. ApoE2 increased the cellular internalization of monomeric Aß, hence preventing its extracellular aggregation, while ApoE4 decreased it, thus helping the buildup of extracellular plaques. The contrary effects of ApoE2 and ApoE4 remained once Aß aggregated: while ApoE2 reduced the uptake of Aß aggregates, ApoE4 facilitated it. Fibrillation studies also revealed ApoE4's tendency to form fibrillar aggregates. Our results uncover yet unknown details of ApoE cellular biology and deepen our molecular understanding of the ApoE-dependent mechanism of Aß pathology.

Contribution of Syndecans to the Cellular Entry of SARS-CoV-2.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a novel emerging pathogen causing an unprecedented pandemic in 21st century medicine. Due to the significant health and economic burden of the current SARS-CoV-2 outbreak, there is a huge unmet medical need for novel interventions effectively blocking SARS-CoV-2 infection. Unknown details of SARS-CoV-2 cellular biology hamper the development of potent and highly specific SARS-CoV-2 therapeutics. Angiotensin-converting enzyme-2 (ACE2) has been reported to be the primary receptor for SARS-CoV-2 cellular entry. However, emerging scientific evidence suggests the involvement of additional membrane proteins, such as heparan sulfate proteoglycans, in SARS-CoV-2 internalization. Here, we report that syndecans, the evolutionarily conserved family of transmembrane proteoglycans, facilitate the cellular entry of SARS-CoV-2. Among syndecans, the lung abundant syndecan-4 was the most efficient in mediating SARS-CoV-2 uptake. The S1 subunit of the SARS-CoV-2 spike protein plays a dominant role in the virus's interactions with syndecans. Besides the polyanionic heparan sulfate chains, other parts of the syndecan ectodomain, such as the cell-binding domain, also contribute to the interaction with SARS-CoV-2. During virus internalization, syndecans colocalize with ACE2, suggesting a jointly shared internalization pathway. Both ACE2 and syndecan inhibitors exhibited significant efficacy in reducing the cellular entry of SARS-CoV-2, thus supporting the complex nature of internalization. Data obtained on syndecan specific in vitro assays present syndecans as novel cellular targets of SARS-CoV-2 and offer molecularly precise yet simple strategies to overcome the complex nature of SARS-CoV-2 infection.

Syndecan-1 modulates the invasive potential of endometrioma via TGF-ß signalling in a subgroup of women with endometriosis.

STUDY QUESTION: What is the physiological role of transforming growth factor-beta (TGF-ß1) and syndecans (SDC1, SDC4) in endometriotic cells in women with endometriosis? SUMMARY ANSWER: We observed an abnormal, pro-invasive phenotype in a subgroup of samples with ovarian endometriosis, which was reversed by combining gene silencing of SDC1 with the TGF-ß1 treatment. WHAT IS KNOWN ALREADY: Women with endometriosis express high levels of TGF-ß1 and the proteoglycan co-receptors SDC1 and SDC4 within endometriotic cysts. However, how SDC1 and SDC4 expression is regulated by TGF-ß1 and the physiological significance of the high expression in endometriotic cysts remains unknown as does the potential role in disease severity. STUDY DESIGN, SIZE, DURATION: We utilized a pre-validated panel of stem- and cancer cell-associated markers on endometriotic tissue (n = 15) to stratify subgroups of women with endometriosis. Furthermore, CD90+CD73+CD105+ (SC+) endometriotic stromal cells from these patient subgroups were explored for their invasive behaviour in vitro by transient gene inhibition of SDC1 or SDC4, both in the presence or absence of TGF-ß1 treatment. PARTICIPANTS/MATERIALS, SETTING, METHODS: Endometriotic cyst biopsies (n = 15) were obtained from women diagnosed with ovarian endometriosis (ASRM Stage III-IV). Gene expression variability was assessed on tissue samples by applying gene clustering tools for the dataset generated from the pre-validated panel of markers. Three-dimensional (3D) spheroids from endometriotic SC+ were treated in vitro with increasing doses of TGF-ß1 or the TGFBRI/II inhibitor Ly2109761 and assessed for SDC1, SDC4 expression and in vitro 3D-spheroid invasion. Transcriptomic signatures from the invaded 3D spheroids were evaluated upon combining transient gene silencing of SDC1 or SDC4, both in presence or absence of TGF-ß1 treatment. Furthermore, nanoscale changes on the surface of endometriotic cells were analysed after treatment with TGF-ß1 or TGFBRI/II inhibitor using atomic force microscopy. MAIN RESULTS AND THE ROLE OF CHANCE: Gene clustering analysis revealed that endometriotic tissues displayed variability in their gene expression patterns; a small subgroup of samples (2/15, Endo-hi) exhibited high levels of SDC1, SDC4 and molecules involved in TGF-ß signalling (TGF-ß1, ESR1, CTNNB1, SNAI1, BMI1). The remaining endometriotic samples (Endo-lo) showed a uniform, low gene expression profile. Three-dimensional spheroids derived from Endo-hi SC+ but not Endo-lo SC+ samples showed an aberrant expression of SDC1 and exhibited enhanced 3D-spheroid invasion in vitro, upon rhTGF-ß1 treatment. However, this abnormal, pro-invasive response of Endo-hi SC+ was reversed upon gene silencing of SDC1 with the TGF-ß1 treatment. Interestingly, transcriptomic signatures of 3D spheroids silenced for SDC1 and consecutively treated with TGF-ß1, showed a down-regulation of cancer-associated pathways such as WNT and GPCR signalling. LARGE SCALE DATA: Transcriptomic data were deposited in NCBI's Gene Expression Omnibus (GEO) and could be retrieved using GEO series accession number: GSE135122. LIMITATIONS, REASONS FOR CAUTION: It is estimated that about 2.5% of endometriosis patients have a potential risk for developing ovarian cancer later in life. It is possible that the pro-oncogenic molecular changes observed in this cohort of endometriotic samples may not correlate with clinical occurrence of ovarian cancer later in life, thus a validation will be required. WIDER IMPLICATIONS OF THE FINDINGS: This study emphasizes the importance of interactions between syndecans and TGF-ß1 in the pathophysiology of endometriosis. We believe that this knowledge could be important in order to better understand endometriosis-associated complications such as ovarian cancer or infertility. STUDY FUNDING/COMPETING INTEREST(S): This study was funded by Cancerfonden (CAN 2016/696), Radiumhemmets Forskningsfonder (Project no. 154143 and 184033), EU MSCA-RISE-2015 project MOMENDO (691058), Estonian Ministry of Education and Research (IUT34-16), Enterprise Estonia (EU48695) and Karolinska Institute. Authors do not have any conflict of interest.

Plasma and Urinary Glycosaminoglycans as Evidence for Endotheliopathy in a Swine Burn Model.

BACKGROUND: The endothelial glycocalyx controls vascular permeability, cellular signaling, blood-endothelial cell adhesion, extravasation, and transmission of shear stress signals. Burn injury compromises integrity of this layer increasing vascular permeability, which is further exacerbated by large volumes of (intravenous) crystalloids. We have shown that enteral resuscitation is able to reverse burn-induced acute kidney injury (AKI), and herein, we present a follow-up examination of the integrity of the glycocalyx layer and its relationship with renal dysfunction after burn injury. MATERIALS AND METHODS: Anesthetized Yorkshire pigs sustained 40% total body surface area full-thickness contact burns and recovered in metabolic cages for one of three treatments: no fluids (oral or intravenous); (n = 6), ad libitum water (n = 6), or volume-matched oral rehydration solution (ORS; n = 6) for 48 h. Urine and blood were collected at baseline (BL), 6, 12, 24, 32, and 48 h after burn at which point kidneys were harvested. RESULTS: In no fluid and water groups (but not ORS), plasma levels of glycosaminoglycans (GAGs) were elevated after burn (P ≤ 0.031). Syndecan-1 was elevated by 6 h after burn in all animals, but levels declined by 24 h with enteral fluids. Urinary GAGs in the no-fluid group were elevated after burn. No differences among treatments were detected in syndecan-1 levels, or glomerular lectin within the kidney. CONCLUSIONS: Collectively, these data demonstrate that ORS prevented increases in circulating GAGs. Furthermore, an inexpensive and simple method for detecting GAGs provides a sensitive measure of endotheliopathy after burn.

Calcium in Cell-Extracellular Matrix Interactions.

In multicellular organisms, the cells are surrounded by persistent, dynamic extracellular matrix (ECM), the largest calcium reservoir in animals. ECM regulates several aspects of cell behavior including cell migration and adhesion, survival, gene expression and differentiation, thus playing a significant role in health and disease. Calcium is reported to be important in the assembly of ECM, where it binds to many ECM proteins. While serving as a calcium reservoir, ECM macromolecules can directly interact with cell surface receptors resulting in calcium transport across the membrane. This chapter mainly focusses on the role of cell-ECM interactions in cellular calcium regulation and how calcium itself mediates these interactions.

Mesoglycan induces keratinocyte activation by triggering syndecan-4 pathway and the formation of the annexin A1/S100A11 complex.

Wound healing is a dynamic process comprising multiple events, such as inflammation, re-epithelialization, and tissue remodeling. Re-epithelialization phase is characterized by the engagement of several cell populations, mainly of keratinocytes that sequentially go through cycles of migration, proliferation, and differentiation to restore skin functions. Troubles can arise during the re-epithelialization phase of skin wound healing particularly in keratinocyte migration, resulting in chronic non-healing lesions, which represent a serious clinical problem. Over the last decades, the efforts aimed to find new pharmacological approaches for wound care were made, yet almost all current therapeutic strategies used remain inadequate or even ineffective. As such, it is crucial to identify new drugs that can enable a proper regeneration of the epithelium in wounded skin. Here, we have investigated the effects of the fibrinolytic drug mesoglycan, a glycosaminoglycans mixture derived from porcine intestinal mucosa on HaCaT human keratinocytes that were used as in vitro experimental model of skin re-epithelialization. We found that mesoglycan induces keratinocyte migration and early differentiation by triggering the syndecan-4/PKCα pathway and that these effects were at least in part, because of the formation of the annexin A1/S100A11 complex. Our data suggest that mesoglycan may be useful as a new pro-healing drug for skin wound care.

Physiological and anatomical aspects of the reproduction of mice with reduced Syndecan-1 expression.

BACKGROUND: Syndecan-1 is a heparan sulfate proteoglycan acting as a co-receptor for cytokines and growth factors mediating developmental, immunological and angiogenic processes. In human, the uteroplacental localization of Syndecan-1 and its reduced expression in pregnancy-associated pathologies, such as the intrauterine growth restriction, suggests an influence of Syndecan-1 in embryo-maternal interactions. The aim of the present study was to identify the effect of a reduced expression of Syndecan-1 on the reproductive phenotype of mice and their progenies. METHODS: Reproductive characteristics have been investigated using animals with reduced Syndecan-1 and their wildtype controls after normal mating and after vice versa embryo transfers. Female mice were used to measure the estrus cycle length and the weight gain during pregnancy, as well as for histological examination of ovaries. Male mice were examined for the concentration, motility, viability and morphology of spermatozoa. Organs like heart, lung, liver, kidney, spleen, brain and ovaries or testes and epididymis of 6-month-old animals were isolated and weighed. Statistical analyses were performed using two-tailed students t-test with P < .05 and P < .02, chi square test (P < .05) and Fisher's Exact Test (P < .05). A linear and a non-linear mixed-effects model were generated to analyze the weight gain of pregnant females and of the progenies. RESULTS: Focusing on the pregnancy outcome, the Syndecan-1 reduced females gave birth to larger litters. However, regarding the survival of the offspring, a higher percentage of pups with less Syndecan-1 died during the first postnatal days. Even though the ovaries and the testes of Syndecan-1 reduced mice showed no histological differences and the ovaries showed a similar number of primary and secondary follicles and corpora lutea, the spermatozoa of Syndecan-1 reduced males showed more tail and midpiece deficiencies. Concerning the postnatal and juvenile development the pups with reduced Syndecan-1 expression remained lighter and smaller regardless whether carried by mothers with reduced Syndecan-1 or wildtype foster mothers. With respect to anatomical differences kidneys of both genders as well as testes and epididymis of male mice with reduced syndecan-1 expression weighed less compared to controls. CONCLUSIONS: These data reveal that the effects of Syndecan-1 reduction are rather genotype- than parental-dependent.

Acute exercise increases syndecan-1 and -4 serum concentrations.

The effects of acute and long-term exercise on syndecans and the relationship to insulin sensitivity are not fully explored. We aimed to examine the effects of acute and 12 weeks of exercise on (1) serum levels of syndecan-1 and -4, (2) gene expression related to syndecan synthesis and modification in skeletal muscle and adipose tissue, and (3) the relationship to insulin sensitivity. Sedentary men with (n = 13) or without (n = 13) dysglycemia underwent two 45 min acute bicycle tests interspersed by 12 weeks of exercise intervention. Euglycemic hyperinsulinemic clamp and mRNA-sequencing of skeletal muscle and adipose tissue biopsies were performed before and after intervention. Serum syndecan-1 and -4 levels were quantified before, immediately after and 2 h after bicycling. Syndecan-1 and -4 serum concentrations increased in response to acute physical exercise. Baseline syndecan-4 but not syndecan-1 concentrations were higher in dysglycemic compared to normoglycemic men, and correlated to change in insulin sensitivity, but did not change during the 12 weeks exercise intervention. Only syndecan-4 was expressed in skeletal muscle and adipose tissue. Adipose tissue mRNA levels of transcripts affecting syndecan structure and shedding were upregulated in dysglycemia, and muscle mRNA responded to long-term physical activity. The increase in serum syndecan-1 and -4 due to acute exercise suggest increased syndecan shedding and disruption of glycocalyx in response to increased blood flow. The higher syndecan-4 baseline serum levels in dysglycemia, association to insulin sensitivity, and changes in mRNA transcripts may suggest syndecan-4 involvement in muscle and adipose tissue response to exercise.

Syndecans in chronic inflammatory and autoimmune diseases: Pathological insights and therapeutic opportunities.

Syndecans (SDCs) are a family of heparan sulfate proteoglycans (HSPGs) glycoproteins ubiquitously expressed on the cell surfaces and extracellular matrix of all mammalian tissues. There are four mammalian syndecans, SDC-1 thorough 4, which play a critical role in cell adhesion, migration, proliferation, differentiation, and angiogenesis through independent and growth factor mediated signaling. An altered expression of SDCs is often observed in autoimmune disorders, cancer, HIV infection, and many other pathological conditions. SDCs modulate disease progression by interacting with a diverse array of ligands, receptors, and other proteins, including extracellular matrix, glycoproteins, integrins, morphogens, and various growth factors and chemokines, along with their receptors and kinases. Specifically, SDCs present on cell surface can bind directly to chemokines to enhance their binding to receptors, downstream signaling, and migration. Alternatively, SDCs can be cleaved and shed to mediate negative regulation of chemokine and growth factor signaling pathways and ligand sequestration. Importantly, SDC shedding may be a biomarker of inflammation, especially in chronic inflammatory diseases. While the current therapies for cancer and several autoimmune disorders have revolutionized treatment outcomes, understanding the pathophysiological role of SDCs and the use of HSPG mimetic or antagonists on cytokine signaling networks may uncover potentially novel targeted therapeutic approaches. This review mainly summarizes the current findings on the role of individual SDCs in disease processes, mechanisms through which SDCs mediate their biological functions, and the possibility of targeting SDCs as future potential therapeutic approaches.