Syndecan-1 Shedding Research Articles

Circulating glycocalyx shedding products as biomarkers for evaluating prognosis of patients with out-of-hospital cardiac arrest after return of spontaneous circulation

The endothelial glycocalyx is damaged in postcardiac arrest syndrome (PCAS), but the prognostic value is unknown. We aimed to observe the expression and prognostic value of glycocalyx shedding products, including syndecan-1 (SDC-1), hyaluronan (HA), and heparan sulfate (HS) in PCAS. Data on clinical and 28-day outcomes of seventy-one consecutive patients with out-of-hospital cardiac arrest (OHCA) after the return of spontaneous circulation (ROSC) were collected. SDC-1, HA, and HS were measured on days 0, 1, and 3 after ROSC. Thirty healthy individuals were controls. Glycocalyx shedding was observed in human umbilical vein endothelial cells (HUVECs) stimulated during hypoxia and reoxygenation in vitro. Within 4 h of ROSC, SDC-1 and HA levels, significantly increased. In the 28-day non-survivors, HA levels showed a gradual upward trend, SDC-1 remained at a high level, and HS levels first increased, then decreased. Kaplan–Meier curves and binary logistic regression analysis showed the prognostic value of SDC-1 levels on days 0, 1, and 3, HA levels on days 1 and 3, and HS levels on day 1. Only HS levels on day 1 showed a prognostic value for 28-day neurological outcomes. SDC-1 and HA levels were positively correlated with the no-flow time. In vitro, HUVECs showed shedding of SDC-1 and HS during a prolonged duration of hypoxia. After ROSC, SDC-1, HA, and HS levels may predict the 28-day survival after PCAS, and HS levels are associated with functional outcomes.

The effects of metabolites from three vaginal bacteria on the Syndecan-1 of cervical epithelial cells

This study aims to explore the impact of metabolites from three vaginal bacteria on the expression of Syndecan 1 (SDC-1). Human cervical epithelial cells (HcerEpic) were separately incubated with the cell-free supernatants of Lactobacillus crispatus (LCS group), Gardnerella vaginalis (GVS group), and Atopobium vaginalis (AVS group). LCS showed a proliferative effect on HcerEpic, with the most significant effect observed at a concentration of 30 % (P < 0.001). GVS and AVS exhibited some cytotoxicity, with significant growth inhibitory effects observed at concentrations of 30 % and 40 % (P < 0.01). Therefore, subsequent experiments were conducted using 30 % LCS, 40 % GVS, and 40 % AVS. In terms of cellular morphology, compared to the Control group, the LCS group showed more frequent fusion of cell sheets, with no obvious changes in the morphology of individual cells. In the GVS and AVS groups, some individual cells became round and smaller, with reduced protrusions and even a small amount of floating cells. The metabolic products of the three vaginal bacteria significantly upregulated the expression of IL-1β, IL-6, and TNF-α in HcerEpic (P < 0.05). In the GVS and AVS groups, the level of SDC-1 on the surface of HcerEpic was significantly decreased (P < 0.01), while the concentration of SDC-1 in the cell culture supernatant was significantly increased (P < 0.0001). Additionally, the level of SDC-1 mRNA was significantly downregulated (P < 0.01). In the LCS group, no significant changes were observed in SDC-1 protein and mRNA expression (P > 0.05). LCS promotes HcerEpic proliferation, without significant impact on SDC-1 expression and shedding. This provides molecular evidence for LCS as a protective factor against human papillomavirus infection in the cervix. Metabolites of GV and AV inhibit HcerEpic proliferation, induce cytokine secretion, suppress SDC-1 transcription and expression, and promote SDC-1 shedding.

Endothelial dysfunction is dampened by early administration of fresh frozen plasma in a rodent burn shock model.

Endothelial dysfunction has been implicated in the pathogenesis of burn shock affecting patients with large thermal injury. In response to injury, glycocalyx components like Syndecan-1 (SDC-1) are shed into circulation and have been used as markers of endothelial damage. Previous work in our laboratory has shown that plasma inclusive resuscitation (PIR) with fresh frozen plasma (FFP) ameliorates endothelial damage. However, there remains a paucity of information regarding optimal timing and dosing of PIR as well as organ-specific endothelial responses to shock. We aimed to examine the impact of PIR on endothelial dysfunction using clinically translatable timing and dosing. Sprague-Dawley rats were used to create thermal burns. Rats were subjected to 40% total body surface area scald burns and were resuscitated with lactated Ringer's (LR) only, LR plus albumin, and LR plus early 1 mL boluses of FFP at 0, 2, 4, and 8 hours postinjury. A late group also received LR plus FFP starting at hour 10 postinjury. Syndecan-1 levels were quantified by enzyme-linked immunosorbent assay, and quantitative real-time polymerase chain reaction analysis characterized transcription of glycocalyx components and inflammatory cytokines in the lung and spleen. Evan's blue dye was used to quantify amount of vascular leakage. Lactated Ringer's plus early FFP reduced Evan's blue dye extravasation when compared with LR only groups, while late FFP did not. When comparing LR only versus LR plus early FFP, SDC-1 levels were reduced in the LR plus early FFP group at hours 8, 12, and 24 (5.23 vs. 2.07, p < 0.001; 4.49 vs. 2.05, p < 0.01; and 3.82 vs. 2.08, p < 0.05, respectively). Lactated Ringer's only groups had upregulation of Exostosin-1 and SDC-1 in the lung compared with LR plus early FFP groups ( p < 0.01 and p < 0.05) and upregulation of cytokines interluekin-10 and interferon γ ( p < 0.001 and p < 0.001). Early administration of LR plus FFP reduces the magnitude of SDC-1 shedding and dampens the cytokine response to injury. The upregulation of glycocalyx components as a response to endothelial injury is also decreased in the lung and spleen by LR plus early FFP administration.

DMPP attenuates lipopolysaccharide-induced lung injury by inhibiting glycocalyx degradation through activation of the cholinergic anti-inflammatory pathway.

The study aimed to investigate the therapeutic potential of 1,1-dimethyl-4-phenylpiperazinium iodide (DMPP), an agonist of nicotinic acetylcholine receptor (nAChR), in treating acute lung injury (ALI) induced by lipopolysaccharide (LPS). A murine ALI model was developed utilizing intraperitoneal injection of LPS. We evaluated the therapeutic efficacy of DMPP treatment in LPS-induced lung injury using various approaches, including pathohistological evaluation, appraisal of pulmonary edema, and measurement of inflammatory cytokine levels and their associated pathways within lung tissues. The gene chip data of LPS-induced acute lung injury mice were retrieved from the Gene Expression Omnibus (GEO) database for gene differential expression analysis and Gene Set Enrichment Analysis (GSEA) analysis. The impact of DMPP on glycocalyx shedding was assessed by measuring the expression levels of syndecan-1 (SDC-1) and matrix metalloproteinase-9 (MMP-9). DMPP treatment significantly improved pathomorphological changes and pathological lung injury scores in the LPS-induced ALI mouse model. The genes expressed differentially in the LPS-induced ALI group in GSE2411 were found to be involved in multiple processes, including the NF-κB signaling pathway, NOD-like receptor signaling pathway, Toll-like receptor signaling pathway, as well as the JAK-STAT signaling pathway. DMPP treatment effectively downregulated pro-inflammatory cytokines, suppressed the NF-κB signaling pathway, and effectively restrained the LPS-induced upregulation of MMP-9 and shedding of syndecan-1, thereby contributing to the preservation of endothelial glycocalyx and attenuation of endothelial barrier dysfunction. The administration of DMPP has been shown to confer protection against LPS-induced acute lung injury via a cholinergic anti-inflammatory pathway, which effectively inhibits endothelial glycocalyx degradation.

Syndecan-1 shedding destroys epithelial adherens junctions through STAT3 after renal ischemia/reperfusion injury

Syndecan-1 shedding destroys epithelial adherens junctions through STAT3 after renal ischemia/reperfusion injury

Dynamic Interplay in Tumor Ecosystems: Communication between Hepatoma Cells and Fibroblasts.

Tumors are intricate ecosystems where cancer cells and non-malignant stromal cells, including cancer-associated fibroblasts (CAFs), engage in complex communication. In this study, we investigated the interaction between poorly (HLE) and well-differentiated (HuH7) hepatoma cells and LX2 fibroblasts. We explored various communication channels, including soluble factors, metabolites, extracellular vesicles (EVs), and miRNAs. Co-culture with HLE cells induced LX2 to produce higher levels of laminin β1, type IV collagen, and CD44, with pronounced syndecan-1 shedding. Conversely, in HuH7/LX2 co-culture, fibronectin, thrombospondin-1, type IV collagen, and cell surface syndecan-1 were dominant matrix components. Integrins α6β4 and α6β1 were upregulated in HLE, while α5β1 and αVβ1 were increased in HuH7. HLE-stimulated LX2 produced excess MMP-2 and 9, whereas HuH7-stimulated LX2 produced excess MMP-1. LX2 activated MAPK and Wnt signaling in hepatoma cells, and conversely, hepatoma-derived EVs upregulated MAPK and Wnt in LX2 cells. LX2-derived EVs induced over tenfold upregulation of SPOCK1/testican-1 in hepatoma EV cargo. We also identified liver cancer-specific miRNAs in hepatoma EVs, with potential implications for early diagnosis. In summary, our study reveals tumor type-dependent communication between hepatoma cells and fibroblasts, shedding light on potential implications for tumor progression. However, the clinical relevance of liver cancer-specific miRNAs requires further investigation.

Sulodexide improves vascular permeability via glycocalyx remodelling in endothelial cells during sepsis.

Degradation of the endothelial glycocalyx is critical for sepsis-associated lung injury and pulmonary vascular permeability. We investigated whether sulodexide, a precursor for the synthesis of glycosaminoglycans, plays a biological role in glycocalyx remodeling and improves endothelial barrier dysfunction in sepsis. The number of children with septic shock that were admitted to the PICU at Children's Hospital of Fudan University who enrolled in the study was 28. On days one and three after enrollment, venous blood samples were collected, and heparan sulfate, and syndecan-1 (SDC1) were assayed in the plasma. We established a cell model of glycocalyx shedding by heparinase III and induced sepsis in a mouse model via lipopolysaccharide (LPS) injection and cecal ligation and puncture (CLP). Sulodexide was administrated to prevent endothelial glycocalyx damage. Endothelial barrier function and expression of endothelial-related proteins were determined using permeability, western blot and immunofluorescent staining. The survival rate, histopathology evaluation of lungs and wet-to-dry lung weight ratio were also evaluated. We found that circulating SDC1 levels were persistently upregulated in the non-alive group on days 1 and 3 and were positively correlated with IL-6 levels. Receiver operating characteristic curve analysis showed that SDC1 could distinguish patients with mortality. We showed that SDC1-shedding caused endothelial permeability in the presence of heparinase III and sepsis conditions. Mechanistically, sulodexide (30 LSU/mL) administration markedly inhibited SDC1 shedding and prevented endothelial permeability with zonula occludens-1 (ZO-1) upregulation via NF-κB/ZO-1 pathway. In mice with LPS and CLP-induced sepsis, sulodexide (40 mg/kg) administration decreased the plasma levels of SDC1 and increased survival rate. Additionally, sulodexide alleviated lung injury and restored endothelial glycocalyx damage. In conclusion, our data suggest that SDC1 predicts prognosis in children with septic shock and sulodexide may have therapeutic potential for the treatment of sepsis-associated endothelial dysfunction.

Myeloperoxidase inhibition may protect against endothelial glycocalyx shedding induced by COVID-19 plasma

BackgroundSARS-CoV-2, the causative agent of COVID-19, is a threat to public health. Evidence suggests increased neutrophil activation and endothelial glycocalyx (EG) damage are independently associated with severe COVID-19. Here, we hypothesised that an increased level of blood neutrophil myeloperoxidase (MPO) is associated with soluble EG breakdown, and inhibiting MPO activity may reduce EG damage.MethodsAnalysing a subset of acute and convalescent COVID-19 plasma, 10 from severe and 15 from non-severe COVID-19 cases, and 9 from pre-COVID-19 controls, we determined MPO levels, MPO activity and soluble EG proteins (syndecan-1 and glypican-1) levels by enzyme-linked immunosorbent assay. In vitro primary human aortic endothelial cells were cultured with plasma untreated or treated with specific MPO inhibitors (MPO-IN-28, AZD5904) to determine EG shedding. We then investigated whether inhibiting MPO activity decreased EG degradation.ResultsIn COVID-19 plasma, MPO levels, MPO activity and levels of soluble EG proteins are significantly raised compared to controls, and concentrations increase in proportion to disease severity. Despite clinical recovery, protein concentrations remain significantly elevated. Interestingly, there is a trend of increasing MPO activity in convalescent plasma in both severe and non-severe groups. MPO levels and MPO activity correlate significantly with soluble EG levels and inhibiting MPO activity leads to reduced syndecan-1 shedding, in vitro.ConclusionsNeutrophil MPO may increase EG shedding in COVID-19, and inhibiting MPO activity may protect against EG degradation. Further research is needed to evaluate the utility of MPO inhibitors as potential therapeutics against severe COVID-19.

Fibrinogen Inhibits Metalloproteinase-9 Activation and Syndecan-1 Cleavage to Protect Lung Function in ApoE Null Mice After Hemorrhagic Shock

IntroductionObesity is associated with higher mortality following trauma, although the pathogenesis is unclear. Both obesity and trauma are associated with syndecan-1 shedding and metalloproteinase-9 (MMP-9) activation, which can adversely affect endothelial cell function. We recently demonstrated that fibrinogen stabilizes endothelial cell surface syndecan-1 to reduce shedding and maintain endothelial barrier integrity. We thus hypothesized that MMP-9 activation and syndecan-1 shedding would be exacerbated by obesity after trauma but attenuated by fibrinogen-based resuscitation. Materials and methodsApoE null (−/−) mice were fed a Western diet to induce obesity. Mice were subjected to hemorrhage shock and laparotomy then resuscitated with Lactated Ranger's (LR) or LR containing fibrinogen and compared to null and lean sham wild type mice. Mean arterial pressure (MAP) was monitored. Bronchial alveolar lavage protein as an indicator of permeability and lung histopathologic injury were assessed. Syndecan-1 protein and active MMP-9 protein were measured. ResultsMAP was similar between lean sham and ApoE−/− sham mice. However, following hemorrhage, ApoE−/− mice resuscitated with fibrinogen had significantly higher MAP than LR mice. Lung histopathologic injury and permeability were increased in LR compared to fibrinogen resuscitated animals. Compared with lean sham mice, both active MMP-9 and cleaved syndecan-1 level were significantly higher in ApoE−/− sham mice. Resuscitation with fibrinogen but not lactated Ringers largely reduced these changes. ConclusionsFibrinogen as a resuscitative adjunct in ApoE−/− mice after hemorrhage shock augmented MAP and reduced histopathologic injury and lung permeability, suggesting fibrinogen protects the endothelium by inhibiting MMP-9–mediated syndecan-1 cleavage in obese mice.

Blood component resuscitative strategies to mitigate endotheliopathy in a murine hemorrhagic shock model.

Resuscitation with plasma components has been shown to improve endotheliopathy induced by hemorrhagic shock, but the optimal resuscitation strategy to preserve the endothelial glycocalyx has yet to be defined. The aim of this study was to determine if resuscitation with lactated Ringer's (LR), whole blood (WB), packed red blood cells (RBCs), platelet-rich plasma (PRP), platelet poor plasma, balanced RBC:PRP (1:1), or day 14 (d14) RBC would best minimize endothelial damage following shock. Male C57BL/6 mice were hemorrhaged to a goal mean arterial pressure of 25 mm Hg for 1 hour. Unshocked sham mice served as controls. Mice were then resuscitated with equal volumes of LR, WB, RBC, PRP, platelet poor plasma, 1:1, or d14 RBC and then sacrificed at 1, 4, or 24 hours (n = 5). Serum was analyzed for syndecan-1, ubiquitin C-terminal hydrolase L1, and cytokine concentrations. Lungs underwent syndecan-1 immunostaining, and lung injury scores were calculated after hematoxylin and eosin. Proteolytic cleavage of the endothelial glycocalyx was assessed by serum matrix metalloprotease 9 levels. Serum syndecan-1 and ubiquitin C-terminal hydrolase L1 levels were significantly increased following resuscitation with d14 RBC compared with other groups. Early elevation in lung syndecan-1 staining was noted in LR-treated mice, while d14 mice showed decreased staining compared with sham mice following shock. Lung injury scores were significantly elevated 4 hours after resuscitation with LR and d14 RBC compared with WB. Serum matrix metalloprotease 9 levels were significantly increased at 1 and 4 hours in d14 mice compared with sham mice. Systemic inflammation was increased in animals receiving LR, 1:1, or d14 RBC. Resuscitation with WB following hemorrhagic shock reduces endothelial syndecan-1 shedding and mitigates lung injury. Aged RBC and LR fail to attenuate endothelial injury following hemorrhagic shock. Further research will be necessary to determine the effect of each of these resuscitative fluids in a hemorrhagic shock model with the addition of tissue injury.

Inflammation and Syndecan-4 Shedding from Cardiac Cells in Ischemic and Non-Ischemic Heart Disease

Circulating biomarkers reflecting cardiac inflammation are needed to improve the diagnostics and guide the treatment of heart failure patients. The cardiac production and shedding of the transmembrane proteoglycan syndecan-4 is upregulated by innate immunity signaling pathways. Here, we investigated the potential of syndecan-4 as a blood biomarker of cardiac inflammation. Serum syndecan-4 was measured in patients with (i) non-ischemic, non-valvular dilated cardiomyopathy (DCM), with (n = 71) or without (n = 318) chronic inflammation; (ii) acute myocarditis (n = 15), acute pericarditis (n = 3) or acute perimyocarditis (23) and (iii) acute myocardial infarction (MI) at day 0, 3 and 30 (n = 119). Syndecan-4 was investigated in cultured cardiac myocytes and fibroblasts (n = 6-12) treated with the pro-inflammatory cytokines interleukin (IL)-1β and its inhibitor IL-1 receptor antagonist (IL-1Ra), or tumor necrosis factor (TNF)α and its specific inhibitor infliximab, an antibody used in treatment of autoimmune diseases. The levels of serum syndecan-4 were comparable in all subgroups of patients with chronic or acute cardiomyopathy, independent of inflammation. Post-MI, syndecan-4 levels were increased at day 3 and 30 vs. day 0. IL-1Ra attenuated IL-1β-induced syndecan-4 production and shedding in vitro, while infliximab had no effect. In conclusion, syndecan-4 shedding from cardiac myocytes and fibroblasts was attenuated by immunomodulatory therapy. Although its circulating levels were increased post-MI, syndecan-4 did not reflect cardiac inflammatory status in patients with heart disease.

Endothelial damage occurs early after inhalation injury as measured by increased syndecan-1 levels.

Inhalation injury is a significant cause of morbidity and mortality in the burn patient population. However, the pathogenesis of inhalation injury and its potential involvement in burn shock is not well understood. Preclinical studies have shown endothelial injury, as measured by syndecan-1 (SDC-1) levels, to be involved in the increased vascular permeability seen in shock states. Furthermore, the lung has been identified as a site of significant SDC-1 shedding. Here we aim to characterize the contribution of endotheliopathy caused by inhalation alone in a swine model. When comparing injured animals, the fold change of circulating SDC-1 levels from preinjury was significantly higher at 2, 4, and 6 hours postinjury (P = .0045, P = .0017, and P < .001, respectively). When comparing control animals, the fold change of SDC-1 from preinjury was not significant at any timepoint. When comparing injured animals versus controls, the fold change of SDC-1 injured animals was significantly greater at 2, 4, 6, and 18 hours (P = .004, P = .03, P < .001, and P = .03, respectively). Histological sections showed higher lung injury severity compared to control uninjured lungs (0.56 vs 0.38, P < .001). This novel animal model shows significant increases in SDC-1 levels that provide evidence for the connection between smoke inhalation injury and endothelial injury. Further understanding of the mechanisms underlying inhalation injury and its contribution to shock physiology may aid in development of early, more targeted therapies.

Acute pyelonephritis in pregnancy and plasma syndecan-1: evidence of glycocalyx involvement

Background Acute pyelonephritis, a risk factor for maternal sepsis, adult respiratory distress syndrome, and preterm labor, is a frequent cause of hospitalization. This condition is characterized by excessive intravascular inflammation and endothelial cell activation and dysfunction. Syndecan-1, a major component of the glycocalyx, is a gel-like layer that covers the luminal surface of healthy endothelial cells, preserving and mediating many endothelial functions. During pregnancy, there is an additional potential source of syndecan-1, the “syncytiotrophoblast glycocalyx,” which lines the intervillous space. Insults that damage the glycocalyx lead to a shedding of syndecan-1 into the circulation. Hence, syndecan-1 has been proposed as a marker of endothelial injury in conditions such as sepsis, trauma, cardiovascular disease, and diabetes mellitus. Objective The objective of this study was to determine whether the plasma syndecan-1 concentration changes in women with acute pyelonephritis in the presence or absence of bacteremia. Study design This cross-sectional study included three groups: (1) non-pregnant women (n = 25); (2) women with an uncomplicated pregnancy from whom samples were collected preterm (n = 61) or at term (n = 69); and (3) pregnant women diagnosed with acute pyelonephritis from whom samples were collected at the time of diagnosis during the second and third trimesters (n = 33). The diagnosis of acute pyelonephritis was based on clinical findings and a positive urine culture for bacteria. Blood culture results were available in 85% (28/33) of women with acute pyelonephritis. Plasma concentrations of syndecan-1 were determined by a validated immunoassay. Results (1) Women with an uncomplicated pregnancy had a higher plasma concentration of syndecan-1 than non-pregnant women. The geometric mean (95% confidence interval [CI]) of syndecan-1 concentration was 51.0 (12.1–216.1) ng/mL in non-pregnant controls; 1280 (365–4487) ng/mL in normal preterm gestations; and 1786 (546–5834) ng/mL in normal term gestations (adjusted p < .005 for all three between group comparisons); (2) plasma syndecan-1 concentrations increased with gestational age among women with a normal pregnancy (p < .001, R 2 = 0.27); (3) syndecan-1 multiple of the mean (MoM) values in pregnant patients with acute pyelonephritis were higher than those in normal pregnant women based on second- and third-trimester samples (p = .048, 1.26-fold change). The increase was driven primarily by cases with a positive blood culture (p = .009, 1.74-fold change); (4) when data from third-trimester samples were compared, overall differences in syndecan-1 MoM values between cases and controls were slightly larger (p = .03, 1.36- fold change), which were especially contributed to by cases with a positive blood culture (p = .023, fold change 1.79-fold change). Conclusions Plasma syndecan-1 concentration is higher in pregnant women and increases as a function of gestational age. Patients with acute pyelonephritis have a higher plasma concentration of syndecan-1, and this is particularly the case in the presence of bacteremia.

HIF-1α promotes the expression of syndecan-1 and inhibits the NLRP3 inflammasome pathway in vascular endothelial cells under hemorrhagic shock

Hemorrhagic shock (HS) is a global life-threatening matter that causes massive mortality annually worldwide. Syndecan-1 (SDC1) is an important predictor and evaluation index for HS, but its mechanism involved in the HS development remain unclear. HS mice model and human umbilical vein endothelial cells (HUVECs) under hypoxia were applied to explore the relationship of SDC1 with HIF-1α and NLRP3 inflammasome in vascular ECs under HS. Transcriptome sequencing of isolated vascular ECs were conduct to search for hub genes. Dual luciferase assay was adopted to prove the binding effects of the HIF-1α on SDC1 promoter in HUVECs. Molecular expression was evaluated through routine experiments. Here, HS led to aggravated lung injury and inflammatory response with the shedding of SDC1 on the lung vascular ECs in mice. Circulatory SDC1 and proinflammatory cytokines were significantly increased after HS. HIF-1α and IL-1β were identified as hub genes in vascular ECs of HS mice. Meanwhile, HIF-1α-mediaed hypoxia and IL-1β-involved NLRP3 inflammasome pathways were activated following HS. The transcriptional factor HIF-1α promoted the expression of SDC1 through binding to the SDC1 promoter. SDC1 had an inhibitory effect on the NLRP3 inflammasome activity. An exogenous increase of HIF-1α upregulated SDC1 and restrained the activation of the NLRP3 inflammasome under hypoxia, while further interference of SDC1 weakened this effect. Hence, SDC1 is an intermediate connecting HIF-1α and NLRP3 inflammasome in the vascular ECs under hypoxia. HIF-1α promotes the expression of SDC1 and inhibits the NLRP3 inflammasome pathway in vascular ECs under HS.

Fingolimod does not prevent syndecan-4 shedding from the endothelial glycocalyx in a cultured human umbilical vein endothelial cell model of vascular injury

BackgroundShedding of the endothelial glycocalyx (EG) is associated with poor outcomes in a range of conditions including sepsis. Fresh frozen plasma (FFP) restores the damaged EG to baseline thickness, however the mechanism for this effect is unknown, and some components of FFP have adverse effects unrelated to the EG. There is some limited evidence that sphingosine-1-phosphate (S1P) within FFP restores the EG by activating the endothelial cell S1P receptor 1 (S1PR1). However, there are disadvantages to using S1P clinically as an EG restorative therapy. A potential alternative is the S1PR agonist fingolimod (FTY720). The aim of this study was to assess whether FTY720 prevents EG shedding in injured cultured human umbilical vein endothelial cells.MethodsShedding of the EG was induced in cultured human umbilical vein endothelial cells (HUVECs) by exposure to adrenaline, TNF-α and H2O2. The cells were then assigned to one of six conditions for 4 h: uninjured and untreated, injured and untreated, injured and treated with FTY720 with and without the S1PR1 inhibitor W146, and injured and treated with 25% FFP with and without W146. Syndecan-4, a component of the EG, was measured in cell supernatants, and syndecan-4 and thrombomodulin mRNA expression was quantitated in cell lysates.ResultsThe injury resulted in a 2.1-fold increase in syndecan-4 (p < 0.001), consistent with EG shedding. Syndecan-4 and thrombomodulin mRNA expression was increased (p < 0.001) and decreased (p < 0.05), respectively, by the injury. Syndecan-4 shedding was not affected by treatment with FTY720, whereas FFP attenuated syndecan-4 shedding back to baseline levels in the injured cells and this was unaffected by W146. Neither treatment affected syndecan-4 or thrombomodulin mRNA expression.ConclusionsFTY720 did not prevent syndecan-4 shedding from the EG in the HUVEC model of endothelial injury, suggesting that activation of S1PR does not prevent EG damage. FFP prevented syndecan-4 shedding from the EG via a mechanism that was independent of S1PR1 and upregulation of SDC-4 production. Further studies to examine whether FTY720 or another S1PR agonist might have EG-protective effects under different conditions are warranted, as are investigations seeking the mechanism of EG protection conferred by FFP in this experimental model.

Serum syndecan-4 correlates with blood pressure and cardiovascular parameters but not proinflammatory markers in healthy older women.

Aging is accompanied by a low-grade proinflammatory status that plays a role in age-related vascular alterations. Syndecan-4 (SDC4) is a key component of the endothelial glycocalyx, and its extracellular domain can be shed by matrix metalloproteinase-9 (MMP-9). In vitro studies demonstrated that MMP-9-mediated shedding of SDC4 is induced by tumor necrosis factor-α (TNF- α) in human endothelial cells. However, the relationship between circulating shed SDC4, systemic inflammation, and age-related vascular alterations remains unknown. Here, we used linear regression models to examine the associations of serum SDC4 levels with cardiovascular hemodynamic phenotypes, serum MMP-9, and serum TNF-α and inteleukin-6 in healthy older women (n = 74). Serum SDC4 was not associated with proinflammatory cytokines or arterial elasticity. Nevertheless, we found significant correlations of SDC4 with MMP-9, heart rate, left ventricular ejection time, systemic vascular resistance, and blood pressure. Our preliminary evidence suggests that systemic inflammation might not induce SDC4 shedding in healthy aging.

Doxycycline can reduce glycocalyx shedding by inhibiting matrix metalloproteinases in patients undergoing cardiopulmonary bypass: A randomized controlled trial.

Cardiopulmonary bypass (CPB) leads to shedding of the glycocalyx of endothelial cells, resulting in a series of complications such as tissue edema and coagulatory and microcirculatory dysfunctions. Matrix metalloproteinases (MMPs) can cause glycocalyx shedding in a variety of pathological processes, but their role in the process of CPB is still unclear. We hypothesized that the MMPs inhibitor doxycycline would reduce glycocalyx shedding by inhibiting MMPs during CPB. Thirty-six patients were randomized to receive either 100mg oral doxycycline (an MMPs inhibitor) or a matching placebo pill twice a day for three days before CPB. The primary outcome was the concentration of plasma syndecan-1. Secondary outcomes included heparan sulphate, MMP-2, MMP-9, ratio of urinary albumin to creatinine, and short-term clinical outcomes. In order to further prove that MMPs in plasma caused the glycocalyx shedding, human umbilical vein endothelial cells were cultured with plasma obtained from cardiac surgery patients before or after CPB (with or without MMPs inhibitor GM6001). The change in glycocalyx content was detected by immunofluorescence. CPB resulted in an increase of MMPs and shedding of the glycocalyx. Plasma syndecan-1 was higher in the control group than in the doxycycline group (median difference:15.04μg/L; 95% CI: 9.14-20.94μg/L; P<0.001). Similar to syndecan-1, plasma heparan sulphate, MMP-2, and MMP-9 concentrations in the doxycycline group were significantly lower than those in the control group during CPB. Doxycycline was also correlated with a reduction in the ratio of urinary albumin to creatinine and improved the short-term clinical outcomes of patients. Endothelial cells cultured with plasma from patients after CPB showed significant shedding of syndecan-1 and heparan sulphate (post-CPB group vs pre-CPB group, P<0.001). GM6001 was shown to reduce shedding of syndecan-1 and heparan sulphate by inhibiting MMPs (post-CPB+GM6001 group vs post-CPB group, P<0.001). Doxycycline can reduce glycocalyx shedding by inhibiting MMPs during CPB.

45 Endothelial Damage Occurs Early After Inhalation Injury as Measured by Increased syndecan-1 Levels

IntroductionInhalation injury is a significant cause of morbidity and mortality in the burn patient population. However, the pathogenesis of inhalation injury and its potential involvement in burn shock is not well understood. Pre-clinical studies have shown endothelial injury, as measured by syndecan-1 levels, to be involved in the increased vascular permeability seen in shock states. Furthermore, the lung has been identified as a site of significant syndecan-1 shedding. Here we aim to characterize the contribution of endotheliopathy caused by inhalation alone in a swine model.MethodsEight female Yorkshire pigs were used in this experiment. A custom-made smoke box was employed to deliver smoke via endotracheal tube directly into the swine lungs. Carboxyhemoglobin levels were then titrated to a level of 50-75%. Blood was collected at induction of anesthesia, pre-injury, 30 minutes, and at hours 1, 2, 4, 6, and 12 post-injury and was stored in EDTA tubes from which plasma was separated and stored for future analysis. Pigs were necropsied immediately after completion of the experiment and lung samples were placed in all-protect and flash frozen. Histology was performed on lung sections and a validated, published scoring system composed of 5 parameters (neutrophils in the alveolar space, neutrophils in the interstitial space, hyaline membrane formation, protein detritus in the alveolar space and septum thickening) was used to assess lung injury severity (between 0 and 1).Plasma Syndecan-1 (SDC-1) was quantified by ELISA. All data was compared to Syndecan-1 levels measured at induction. Conditions were analyzed with one-way ANOVA with multiple comparisons and Dunnett’s correction for multiple comparisons.ResultsSyndecan-1 levels at induction were 13.74 ± 2.03 ng/ml. Pre-injury and 30 minutes post-injury levels remained similar. Syndecan-1 levels at hour 2 post-injury increased 37% from induction (18.36 ± 1.28 ng/ml, p=0.0057). This trend continued with a 47% percent increase from induction at hour 4 post-injury (19.62 ± 2.15 ng/ml, p=0.0033) and a 49% increase from induction at hour 6 post-injury (20.42 ± 2.43 ng/ml, p=0.0011). Histological sections showed higher lung injury severity compared to control pigs (0.1-0.3 vs. 0.5-0.74, p< .05).ConclusionsSignificant increases in syndecan-1 levels in this animal model provide evidence for a connection between smoke inhalation injury and endothelial injury. Furthermore, the endotheliopathy that leads to burn shock could be exacerbated by inhalation injury, leading to the poor clinical outcomes that are often seen in patients with combined burn and inhalation injuries. Future research should focus on the mechanisms underlying inhalation injury and its contribution to shock physiology.

Vitamin C Improves the Outcomes of Cardiopulmonary Resuscitation and Alters Shedding of Syndecan-1 and p38/MAPK Phosphorylation in a Rat Model.

BackgroundPost‐resuscitation syndrome, involves a severe inflammatory response following successful cardiopulmonary resuscitation. The potential mechanism of Vitamin C (VitC) after cardiopulmonary resuscitation on myocardial and cerebral function, duration of survival is undefined.Methods and ResultsA first set of experiments were done in 18 male Sprague‐Dawley rats for the investigation of short‐term follow‐up, randomized into 3 groups: (1) sham; (2) controls; (3) VitC. Ventricular fibrillation was electrically induced and untreated for 6 minutes. Cardiopulmonary resuscitation including chest compression and mechanical ventilation were then initiated and continued for 8 minutes followed by defibrillation. At 5 minutes after return of spontaneous circulation, either VitC (200 mg/kg) or placebo was administered by intravenous infusion with a syringe pump for half an hour. There were significant improvements in myocardial function and buccal microcirculation in rats treated with VitC after return of spontaneous circulation 4 hours compared with controls. VitC inhibited proinflammatory cytokines (interleukin‐6 and tumor necrosis factor‐α), SDC‐1 (Syndecan‐1), and hyaluronic acid in plasma compared with controls (P<0.01). VitC decreased reactive oxygen species production and inhibited p38/MAPK (mitogen‐activated protein kinase) pathway phosphorylation. A second set with 20 animals was used for assessing the neurological deficit score after return of spontaneous circulation 72 hours, randomized into 2 groups: 1) controls; 2) VitC. The survival rate and neurological deficit score after return of spontaneous circulation 72 hours were improved in VitC‐treated animals compared with those of the control group.ConclusionsVitC reduces the severity of post‐resuscitation myocardial and cerebral dysfunction and improves the survival. The mechanisms may involve inhibiting transcription of inflammatory cytokines and oxidative stress, thus protecting the integrity of the vascular endothelium. Meanwhile VitC reduces shedding of SDC‐1 and alters p38/MAPK phosphorylation and microcirculation.

Prevention of Triglyceridemia by (Non-)Anticoagulant Heparin(oids) Does Not Preclude Transplant Vasculopathy and Glomerulosclerosis.

Background: In renal transplantation, chronic transplant dysfunction (CTD) is associated with increased PCSK9 and dyslipidemia. PCSK9 is an enzyme that increases plasma cholesterol levels by downregulating LDLR expression. We recently showed increased PCSK9–syndecan-1 interaction in conditions of proteinuria and renal function loss. Treatment with heparin(oids) might be a therapeutic option to improve dyslipidemia and CTD. We investigated the effects of (non-)anticoagulant heparin(oids) on serum lipids, syndecan-1 and PCSK9 levels, and CTD development. Methods: Kidney allotransplantation was performed from female Dark Agouti to male Wistar Furth recipients. Transplanted rats received daily subcutaneous injections of saline, unfractionated heparin, and RO-heparin or NAc-heparin (2 mg heparin(oid)/kg BW) until sacrifice after 9 weeks of treatment. Results: Saline-treated recipients developed hypertension, proteinuria, and loss of creatinine clearance (all p < 0.05 compared to baseline), along with glomerulosclerosis and arterial neo-intima formation. Saline-treated recipients showed significant increase in plasma triglycerides (p < 0.05), borderline increase in non-HDLc/HDLc (p = 0.051), and ∼10-fold increase in serum syndecan-1 (p < 0.05), without significant increase in serum PCSK9 at 8 weeks compared to baseline. Heparin and non-anticoagulant RO-heparin administration in transplanted rats completely prevented an increase in triglycerides compared to saline-treated recipients at 8 weeks (both p < 0.05). Heparin(oids) treatment did not influence serum total cholesterol (TC), plasma syndecan-1 and PCSK9 levels, creatinine clearance, proteinuria, glomerulosclerosis, and arterial neo-intima formation, 8 weeks after transplantation. Combining all groups, increased syndecan-1 shedding was associated with TC (r = 0.5; p = 0.03) and glomerulosclerosis (r = 0.53; p = 0.021), whereas the non-HDLc/HDLc ratio was associated with the neo-intimal score in the transplanted kidneys (r = 0.65; p < 0.001). Conclusion: Prevention of triglyceridemia by (non-)anticoagulant heparin(oids) neither influenced PCSK9/syndecan-1 nor precluded CTD, which however did associate with the shedding of lipoprotein clearance receptor syndecan-1 and the unfavorable cholesterol profile.