Serum Hyaluronan Research Articles

Elevated Cancer-Associated Hyaluronan Correlates With Diagnosis and Lymph Node Metastasis of Papillary Thyroid Cancer

The glycosaminoglycan hyaluronan (HA) plays an important role in tumor progression. However, its biological and clinical significance in papillary thyroid cancer (PTC) remains unknown. Immunohistochemistry was performed to examine HA expression in tissues from PTC patients. Two PTC cell lines were treated with HA synthesized inhibitor against HA production to assess its function. Serum HA levels from 107 PTC patients, 30 Hashimoto thyroiditis patients, and 45 normal controls (NC) were measured by chemiluminescence immunoassay. HA levels in fine needle aspiration (FNA) washouts obtained from thyroid nodules and lymph nodes (LNs) were measured by chemiluminescence immunoassay. Area under the curve (AUC) was computed to evaluate HA’s clinical value. HA was highly expressed in PTC. Reducing HA production significantly inhibited PTC cell proliferation and invasion. Importantly, serum HA levels in PTC were significantly higher than those in NCs and Hashimoto thyroiditis and allowed distinguishing of thyroid cancers from NCs with high accuracy (AUC = 0.782). Moreover, elevated serum HA levels in PTC correlate with LN metastasis. HA levels in FNA washouts from PTC patients were significantly higher than those in benign controls, with a high AUC value (0.8644) for distinguishing PTC from benign controls. Furthermore, HA levels in FNA washouts from metastatic LN were significantly higher than those in nonmetastatic LN, with a high AUC value (0.8007) for distinguishing metastatic LNs from nonmetastatic LNs. HA levels in serum and FNA washout exhibited a potential significance for PTC diagnosis and an indicator for LN metastasis in patients with PTC.

Development of Therapeutic Agent for Osteoarthritis via Inhibition of KIAA1199 Activity: Effect of Ipriflavone In Vivo.

This study aimed to clarify the effects of ipriflavone, which effectively reduces KIAA1199 activity, on osteoarthritis (OA) development and progression in an in vivo OA mouse model. The OA model mice were divided into the ipriflavone (200 mg/kg/day) group and the control group. OA onset and progression were evaluated with the Mankin score, and KIAA1199 expression and hyaluronan (HA) accumulation were analyzed by immunostaining. The molecular weight of HA in the cartilage tissue and serum HA concentration were analyzed by chromatography and competitive HA enzyme-linked immunoassay. The effects of ipriflavone on the bovine cartilage explant culture under the influence of IL-1β were also investigated. In the ipriflavone group, Safranin-O stainability was well-preserved, resulting in significant reduction of the Mankin score (p = 0.027). KIAA1199 staining positivity decreased and HA stainability was preserved in the ipriflavone group. The serum HA concentration decreased, and the molecular weight of HA in the cartilage tissue increased in the ipriflavone group. The results of the cartilage explant culture indicated that ipriflavone could reduce GAG losses and increase the molecular weight of HA. Thus, ipriflavone may have an inhibitory effect on OA development/progression. Ipriflavone could be a therapeutic drug for OA by targeting KIAA1199 activity.

Increased urinary albumin leakage is related to injuries of glomerular glycocalyx and podocytes, and associated with tubular dysfunction in preeclampsia.

The pathogenesis of preeclampsia (PE) is known to be endothelial cell damage; however, the existence of dysfunction in glomerular endothelial glycocalyx, podocytes and tubules remains unclear. The glomerular endothelial glycocalyx, basement membrane, podocytes, and tubules are permeability barriers against albumin excretion. This study aimed to assess the relationship between urinary albumin leakage and injuries of the glomerular endothelial glycocalyx, podocytes, and tubules in patients with PE. A total of 81 women with uncomplicated pregnancies (control, n=22), PE (PE, n=36), or gestational hypertension (GH) (GH, n=23) were enrolled. We assessed urinary albumin and serum hyaluronan for glycocalyx injuries, podocalyxin for podocytes injuries, and urinary N-acetyl-β-d-glucosaminidase (NAG) and liver-type fatty acid-binding protein (l-FABP) for renal tubular dysfunctions. The serum hyaluronan and the urinary podocalyxin levels were higher in the PE and GH groups. The urinary NAG and l-FABP levels were higher in the PE group. Urinary NAG and l-FABP levels positively correlated with urinary albumin excretion. Our findings suggest that increased urinary albumin leakage is related to injuries of the glycocalyx and podocytes, and associated with tubular dysfunction in pregnant women with PE. The clinical trial described in this paper was registered at the UMIN Clinical Trials Registry under registration number UMIN000047875. URL of registration: https://centre6.umin.ac.jp/cgi-open-bin/ctr_e/ctr_view.cgi?recptno=R000054437.

Changes in glycocalyx-related biochemical parameters during lung resection in non-small cell carcinoma cases : A pilot study

Objective: Non-small cell lung cancer (NSCLC) is one of the most common neoplasms with high mortality rates, and new studies are needed to understand its characteristics better. This study aimed to determine the changes in the glycocalyx structure related to surgery regarding histopathologic subtypes and to evaluate the correlation of these changes on the development of metastasis and mortality. Material and methods: Serum levels of hyaluronan, VEGF-A, FGF-10, BMP-2, and BMP-4 were measured before and after surgery in 42 patients with NSCLC. The alterations in serum levels of studied markers were evaluated as related to metastasis status and mortality in post-operative 18-24 months. Results: Our study included 15 adenocarcinoma and 27 squamous cell carcinoma cases. Pre- and post-operative values of serum hyaluronan, VEGF-A, FGF-10, and BMP-2 showed significant differences for the whole group (p=0.006, p=0.001, p=0.002, and p=0.004, respectively). Post-operative BMP-2 values also correlated with hyaluronan and VEGF-A values. Post-operative values of hyaluronan and VEGF-A values found incorrelation with metastasis and mortality, while BMP-2 with metastasis and FGF-10 with mortality. Serum values of hyaluronan, VEGF-A, FGF-10 and BMP-2 differed significantly in-between the pre- and post-operative measurements in adenocarcinoma cases (p=0.020, p=0.009, p=0.003, and p=0.011, respectively) , but not that of squamous cell carcinoma. Conclusion: Pre- and post-operative changes in serum hyaluronan, VEGF-A, FGF-10 and BMP-2 values may be associated with metastasis and/or mortality in NSCLC. These findings were also more prominent in adenocarcinoma cases, though further extended studies are needed for a better conclusion.

Circulating hyaluronan as a marker of endothelial glycocalyx damage in dogs with myxomatous mitral valve disease and dogs in a hypercoagulable state

The endothelial glycocalyx (eGlx) lines the luminal surface of endothelial cells, maintaining vascular health. Glycocalyx damage is pathophysiologically important in many diseases across species however few studies have investigated its breakdown in naturally occurring disease in dogs. The aims of the study were to investigate eGlx damage in dogs with myxomatous mitral valve disease (MMVD) diagnosed on echocardiography, and dogs in a hypercoagulable state diagnosed using thromboelastography (TEG), by measuring serum hyaluronan concentrations. Serum hyaluronan was quantified in dogs with MMVD (n = 27), hypercoagulability (n = 21), and in healthy controls dogs (n = 18). Serum hyaluronan concentrations were measured using a commercially-available ELISA validated for use in dogs. Hyaluronan concentrations were compared among groups using Kruskal-Wallis tests, and post-hoc with Dunn’s tests. Serum hyaluronan concentrations (median [range]) were significantly increased in dogs with MMVD (62.4 [22.8–201] ng/mL; P = 0.031) and hypercoagulability (92.40 [16.9–247.6] ng/mL; P < 0.001) compared to controls (45.7 [8.7–80.2] ng/mL). Measurement of serum hyaluronan concentration offers a clinically applicable marker of eGlx health and suggests the presence of eGlx damage in dogs with MMVD and dogs in a hypercoagulable state.

Roles of glomerular endothelial hyaluronan in the development of proteinuria.

Vascular endothelial cells are covered with glycocalyx comprising heparan sulfate, hyaluronan, chondroitin sulfate, and associated proteins. Glomerular endothelial glycocalyx is involved in protecting against induction of proteinuria and structural damage, but the specific components in glycocalyx that represent therapeutic targets remain unclear. Anti‐vascular endothelial growth factor (VEGF) therapy is associated with an increased risk of glomerular endothelial injury. This study investigated whether hyaluronan could provide a therapeutic target to protect against proteinuria. We conducted ex vivo and in vivo experiments to explore the effects of degrading glomerular hyaluronan by administering hyaluronidase and of supplementation with hyaluronan. We investigated hyaluronan expression using biotin‐labeled hyaluronan‐binding protein (HABP) in human kidney specimens or serum hyaluronan in endothelial injuries under inhibition of VEGF signaling. We directly demonstrated hyaluronan in glomerular endothelial layers using HABP staining. Ex vivo and in vivo experiments showed the development of proteinuria after digestion of hyaluronan in glomerular capillaries. Supplementation with hyaluronan after hyaluronidase treatment suppressed proteinuria. Mice in the in vivo study developed albuminuria after intraperitoneal injection of hyaluronidase with decreased glomerular hyaluronan and increased serum hyaluronan. In human kidneys with endothelial cell dysfunction and proteinuria due to inhibition of VEGF, glomerular expression of hyaluronan was reduced even in normal‐appearing glomeruli. Serum hyaluronan levels were elevated in patients with pre‐eclampsia with VEGF signaling inhibition. Our data suggest that hyaluronan itself plays crucial roles in preventing proteinuria and preserving the integrity of endothelial cells. Hyaluronan could provide a therapeutic target for preventing glomerular endothelial glycocalyx damage, including VEGF signaling inhibition.

4‐Methylumbelliferone treatment and hyaluronan inhibition as a therapeutic strategy for chronic prostatitis

Hyaluronan (HA), an extracellular matrix component, accumulates in most chronic inflammatory tissues. Here, we studied the impact of HA on the pathogenesis of chronic prostatitis. First, we sorted demographic characteristics and peripheral blood serum samples from patients with chronic prostatitis/chronic pelvic pain syndrome (CP/CPPS) to assess the relationship between the levels of HA in peripheral blood serum and the severity of inflammation in patients. Second, we induced an experimental autoimmune prostatitis (EAP) mouse model and treated the mice with 4-methylumbelliferone (4-MU) (200 mg/kg/day). After the mice were sacrificed, RNA from Th1 cells of the mouse spleens was extracted for RNA sequencing. We used weighted gene co-expression network analysis (WGCNA) to identify co-expressed gene modules and hub-gene related to the pathogenesis of EAP. The expression of critical genes associated with the identified pathway was confirmed by using western blot analysis. HA was significantly more highly expressed in CP/CPPS patients than in healthy volunteers and positively correlated with the severity of pain, urination symptoms, and quality of life. Besides, the protein expression of HA was significantly higher in prostate tissues derived from EAP models than in those derived from controls. 4-MU, an oral inhibitor of HA synthesis, relieved immunocyte infiltration to the prostate and significantly reduced the proportion of Th1 cells. Based on the WGCNA, we identified 18 co-expression modules and identified that the Grey60 and brown modules were positively associated with the EAP and negatively associated with the Control and 4-MU-treated groups. Pathway enrichment analyses and western blot assays proved that HA potentially activated the cell cycle pathway, increasing the proportion of Th1 cells promoting chronic prostatitis pathogenesis, while these processes were reversed by 4-MU treatment. Our results suggest that HA is elevated in patients with CP/CPPS compared with healthy controls and that targeting HA through 4-MU suppresses the activity of the cell cycle-related pathway, potentially by decreasing the proportion of Th1 cells and relieving chronic prostatitis. Our findings might inspire the clinical treatment of chronic prostatitis.

Diagnostic Accuracy of Serum Hyaluronan for Detecting HCV Infection and Liver Fibrosis in Asymptomatic Blood Donors.

Background: The disease caused by hepatitis C virus (HCV) is asymptomatic, silent, and progressive liver disease. In HCV-infected patients the increase in serum HA is associated with the development of hepatic fibrosis and disease progression. Methods: HCV-RNA detection was performed in all serological samples of blood donors that tested positive using HCV Ultra ELISA. Determination of hyaluronan (HA) was performed in positive HCV samples using ELISA-like fluorometric method. The HA content was compared to HCV viral load, genotype of the virus, liver fibrosis as well as ALT and GGT liver biomarkers. Results: Persistently normal ALT (<40 U/L) and GGT (<50 U/L) serum levels were detected in 75% and 69% of the HCV-Infected blood donors, respectively. Based on ROC analysis, the HA value < 34.2 ng/mL is an optimal cut-off point to exclude HCV viremia (specificity = 91%, NPV = 99%). Applying HA value ≥34.2 ng/mL significant liver fibrosis (≥F2) can be estimated in 46% of the HCV-infected blood donors. HA serum level (≥34.2 ng/mL) associated with a high ALT level (>40 U/mL) can correctly identify HCV infection and probable liver fibrosis (sensitivity = 96% and specificity = 90%) in asymptomatic blood donors. Conclusions: A high level of HA (≥34.2 ng/mL) in association with ALT (≥40 U/L) in serum can provide a good clinical opportunity to detect HCV-infected asymptomatic persons that potentially require a liver biopsy confirmation and antiviral treatment to prevent the development of advanced liver fibrosis or cirrhosis.

The Diagnostic Usefulness of Circulating Profile of Extracellular Matrix Components: Sulfated Glycosaminoglycans (sGAG), Hyaluronan (HA) and Extracellular Part of Syndecan-1 (sCD138) in Patients with Crohn's Disease and Ulcerative Colitis.

The described research focused on the diagnostic usefulness of sulfated glycosaminoglycans (sGAG), hyaluronan (HA), and extracellular part of syndecan-1 (sCD138) as new markers related to extracellular matrix (ECM) remodeling in the intestine during the two most common forms of inflammatory bowel diseases (IBD), i.e., ulcerative colitis (UC) and Crohn’ disease (CD). Inflammatory markers belonging to ECM components were assessed in serum of patients with IBD using an immunoenzymatic method (HA and sCD138) and a method based on the reaction with dimethylmethylene blue (sulfated GAG). Measurements were carried out twice: at baseline and after one year of therapy with prednisone (patients with CD) or adalimumab (patients with UC). No quantitative changes were observed in serum sGAG, HA, and sCD138 concentrations between patients newly diagnosed with CD and the healthy group. In the case of patients with UC, the parameter which significantly differentiated healthy subjects and patients with IBD before biological therapy was HA. Significant correlation between serum HA level and inflammation activity, expressed as Mayo score, was also observed in patients with UC. Moreover, the obtained results have confirmed that steroid therapy with prednisone significantly influenced the circulating profile of all examined ECM components (sGAG, HA, and sCD138), whereas adalimumab therapy in patients with UC led to a significant change in only circulating sGAG levels. Moreover, the significant differences in serum HA levels between patients with UC and CD indicate that quantification of circulating HA may be useful in the differential diagnosis of CD and UC.

Abundance and size of hyaluronan in naked mole-rat tissues and plasma

Large amounts of ultra-high molecular weight hyaluronan (HA) have been described as the main cause of cancer resistance in naked mole-rats (Heterocephalus glaber, NMR). Our work examined HA metabolism in these rodents more closely. HA was localized and quantified using HA binding proteins. Its molecular weight was determined using size exclusion chromatography and gel electrophoresis, HA family gene expression using RNAseq analysis, and hyaluronidase activity using zymography. Guinea pigs (Cavia porcellus) and mice (Mus musculus) were used as controls for some experiments. We found that HA localization was similar in NMR, guinea pig, and mouse tissues but NMR had larger amounts and higher molecular weight (maximum, around 2.5 MDa) of HA in serum and almost all tissues tested. We could not find ultra-high molecular weight HA (≥ 4 MDa) in NMR samples, in contrast to previous descriptions. Hyaluronidase-1 had lower expression and activity in NMR than mouse lymph nodes. RNAseq results showed that, among HA family genes, Tnfaip6 and hyaluronidase-3 (Hyal3) were systematically overexpressed in NMR tissues. In conclusion, NMR samples, contrary to expectations, do not harbor ultra-high molecular weight HA, although its amount and average molecular weight are higher in NMR than in guinea pig tissues and serum. Although hyaluronidase expression and activity are lower in NMR than mouse lymph nodes, this not sufficient to explain the presence of high molecular weight HA. A different activity of the NMR HA synthases remains possible. These characteristics, together with extremely high Hyal3 and Tnfaip6 expression, may provide the NMR with a bespoke, and perhaps protective, HA metabolism.

Biosensor for electrochemical detection of biomarkers of osteoarthritis

Biosensor for electrochemical detection of biomarkers of osteoarthritis

An exploratory study to investigate the association between age, physical activity, femoral trochlear cartilage thickness and biomarkers of tissue metabolism in adult males

PurposeTo investigate the association between age, physical activity, femoral trochlear cartilage thickness and biomarkers of tissue metabolism in a cross-sectional sample of adult males. This study utilizes several emerging biomarkers that have been associated with early joint degenerative changes; serum COMP (cartilage oligomeric matrix protein), HA (hyaluronan) and lubricin.MethodsEighty-one males (age: mean (range): 43(18–70) years; body mass index: 25.2 (21.0–30.6) kg/m2) volunteered. Resting serum COMP, HA and lubricin concentrations were determined via commercially available enzyme-linked immunosorbent assay (ELISA) and femoral trochlear cartilage thickness via supra-patellar ultrasound imaging. Physical activity levels were assessed using questionnaires. Statistical analyses were performed using correlation and regression analyses.ResultsAge was correlated with lateral trochlear cartilage thickness (r = − 0.372; p < 0.01) and serum COMP (r = 0.342; p < 0.01). 7-day physical activity was correlated with serum COMP (r = 0.357, p < 0.01), and 12-month physical activity with both lateral trochlear cartilage thickness (r = 0.340, p = 0.01) and serum HA (r = 0.296, p < 0.05). Regression analyses revealed that age significantly accounted for the variability in lateral cartilage thickness and serum COMP, following the adjustment for potential cofounders. However, the association between age and lateral trochlear cartilage thickness was not moderated by physical activity levels (all p > 0.05).ConclusionThis study indicates that older age may be associated with thinner lateral trochlear cartilage and higher cartilage turnover. Being physically active may also be positive for lateral trochlear cartilage thickness. However, overall, both age and physical activity level only account for a small amount of the variability in cartilage thickness and serum biomarkers.

Epidermal growth factor promotes stromal cells migration and invasion via up-regulation of hyaluronate synthase 2 and hyaluronan in endometriosis

To investigate the role(s) of hyaluronan synthase 2 (HAS2) and hyaluronan in disease progression of endometriosis and epidermal growth factor (EGF)-induced motility changes of endometriotic cells. A case-control experimental study and invitro primary cell culture study. University hospital-affiliated research centers. A total of 21 women with stage I/II endometriosis, 33 women with stage III/IV endometriosis with endometrioma, and 32 women without endometriosis were included in our study. Serum, eutopic endometrial tissues, and/or ectopic endometriotic tissues were collected. Primary eutopic endometrial stromal cells (EuESCs) and ectopic ovarian endometriotic stromal cells (OvESCs) were isolated and cultured from women with ovarian endometrioma, and then treated with or without EGF. The concentrations of EGF and hyaluronan in serum were analyzed by enzyme-linked immunosorbent assay. The expressions and localizations of EGF receptor (EGFR), phosphorylated-(p)EGFR, HAS2, and hyaluronan receptor CD44 in tissues were examined by immunohistochemistry. The mRNA and protein levels of HAS2 in EuESCs and OvESCs were examined by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blot, respectively, and the concentrations of hyaluronan in conditioned medium were examined by enzyme-linked immunosorbent assay (ELISA). Cell motility was evaluated by transwell migration/invasion assays. Serum EGF and hyaluronan concentrations were higher in women with stage III/IV endometriosis than in women with stage I/II or without endometriosis. EGFR, pEGFR, HAS2, and CD44 were immunolocalized in eutopic endometrium and ectopic endometriotic lesions, and the expressions of pEGFR and HAS2 were elevated in ectopic endometriotic lesions compared to eutopic endometrium. Treatment with EGF upregulated HAS2 and hyaluronan expression as well as cell migration and invasion in both EuESCs and OvESCs, and pharmaceutical blocking of EGFR abolished these effects. In addition, knockdown of HAS2 by small interfering RNA attenuated both basal and EGF-induced hyaluronan expression and cell motility changes. Notably, ERK1/2 and AKT signaling pathways were shown to be downstream of EGF in regulating HAS2 and hyaluronan expression as well as cell migration and invasion. EGF increased the expression of endometriosis-associated hyaluronan and its synthase HAS2, both of which mediated EGF-induced stromal cell migration and invasion in women with endometriosis.

Proteoglycan-4 and hyaluronan composition in synovial fluid and serum from clinical equine subjects: relationship to cartilage boundary lubrication and viscosity of synovial fluid

ABSTRACT Purpose: In experimental models of equine joint-injury and osteoarthritis synovial fluid (SF) composition (proteoglycan-4, hyaluronan) can vary, along with changes to SF mechanical function (lubrication, viscosity). The study hypotheses were a) clinical equine joint-injury and disease results in altered SF composition and diminished mechanical function, and b) serum composition (proteoglycan-4 or hyaluronan) changes concurrently. The objectives were to characterize composition (proteoglycan-4, hyaluronan), and function of SF and serum from normal horses compared to clinical groups: osteoarthritis, acute-joint-injury, and osteochondrosis. Materials and Methods: Equine samples of SF (from various joints) and blood were collected at the point-of-care. Proteoglycan-4 concentrations were measured by amplified-luminescence-proximity-assay and enzyme-linked-immunosorbent-assay in SF and serum, respectively. Molecular-weight of hyaluronan was characterized by agarose-gel-electrophoresis, and concentrations were measured by enzyme-linked-immunosorbent-assay kit. Biomechanical function of SF was characterized by an in vitro cartilage-on-cartilage friction test, and viscosity test. Results: SF proteoglycan-4 concentration increased in acute-joint-injury (1185 ± 276 versus normal 205 ± 106 µg/mL, µ± SEM, p < 0.01), with increased percentage of lower molecular-weight hyaluronan in acute-joint-injury and osteochondrosis. SF and serum proteoglycan-4 concentrations were correlated in normal horses (r2 = 0.85, p < 0.05), but not in clinical groups. Cartilage-lubricating ability was unchanged, although steady-shear viscosity of acute-joint-injury SF decreased from normal. Conclusion: Composition of SF from cases of equine acute-joint-injury changed; both proteoglycan-4 concentration and hyaluronan molecular-weight were altered, with decreased SF viscosity, but no associated changes to serum. Serum proteoglycan-4 and hyaluronan concentrations alone may not be useful biomarkers for equine joint-injury or disease.

Correlation analysis of the severity and clinical prognosis of 32 cases of patients with COVID-19.

Correlation analysis of the severity and clinical prognosis of 32 cases of patients with COVID-19.

Clinical value of serum hyaluronan and propeptide of type III collagen in patients with pancreatic cancer.

Hyaluronan (HA) and collagen are highly expressed in pancreatic cancer (PC) stroma. HA and collagen accumulation increase tumor interstitial fluid pressure, compromising blood flow and drug penetration. The aim of this biomarker study was to determine the clinical utility of serum HA and the propeptide of type III collagen (PRO-C3) in patients with PC. A cohort from the Danish BIOPAC study (NCT03311776) including patients with histologically confirmed pancreatic ductal adenocarcinoma (n = 809), ampullary carcinoma (n = 44), distal biliary tract cancer (n = 31), chronic pancreatitis (n = 15), intraductal papillary mucinous neoplasm (n = 41), duodenal adenoma (n = 7) and no cancer (n = 25). Healthy controls were available for serum HA (n = 141) and PRO-C3 (n = 8). The main outcome was overall survival (OS) of patients with PC in relation to pretreatment serum HA and PRO-C3 levels. Patients with PC had higher baseline serum HA and PRO-C3 than healthy subjects and patients with benign conditions. Pretreatment serum baseline HA and PRO-C3 in patients with PC were associated with poorer survival and PRO-C3 remained prognostic also after adjusting for age, performance status, stage, the presence of liver and peritoneum metastasis, and CA19-9. Detection of HA and PRO-C3 may be useful in differentiating between malignant and benign pancreatic conditions. Serum HA and PRO-C3 were prognostic for OS in patients with PC.

Usefulness of serum Mac-2 binding protein glycosylation isomer in patients undergoing hepatectomy: A case controlled study.

BackgroundTo evaluate the clinical significance of Mac-2 binding protein glycosylation isomer (M2BPGi), we investigated the relationship between M2BPGi and clinicopathological and surgical parameters and posthepatectomy complications.Materials and methodsWe examined M2BPGi in 115 patients with hepatic malignancies undergoing hepatectomy. Significance as an independent prognostic marker was determined with multivariate logistic regression analysis.ResultsThe mean serum M2BPGi level was 1.14 ± 1.03 C.O.I. (range 0.2–5.79). M2BPGi in the chronic viral hepatitis group (1.42 ± 1.25) was significantly higher than that in the other disease groups (p < 0.05). The M2BPGi level correlated negatively with platelet count, LHL15 and GSA-Rmax (r = −0.36, −0.69 and −0.56, respectively; p < 0.01) but correlated positively with serum hyaluronate level (fibrotic marker), ICGR15 and HH15 (r = 0.52, 0.63 and 0.57, respectively; p < 0.01). In 53 patients examined for histological hepatic fibrosis, the M2BPGi level was highest for hepatic fibrosis stage 4, indicating cirrhosis (2.15 ± 1.56), and was significantly higher than that for stages 0–2 (p < 0.05). M2BPGi level did not correlate significantly with any surgical parameters. The preoperative level correlated significantly only with increased alanine aminotransferase level (r = −0.21, p < 0.05) and was significantly higher in patients with (1.35 ± 0.78) than without (1.11 ± 1.07) hepatectomy-related complications (p < 0.05). Area under the ROC curve analysis for prediction of hepatic fibrosis score 4 showed a cut-off value of 0.78 for M2BPGi to have high sensitivity (90%) and specificity (58%). For postoperative hepatectomy-related complications, only the M2BPGi level (at a cut-off value 0.90) tended to show significance (p = 0.06).ConclusionsThe non-invasively measured serum level of M2BPGi reflected impaired liver function or cirrhosis and hepatectomy-related complications after surgery, making it potentially useful as a complementary parameter accompanying other liver function parameters.

High levels of serum hyaluronan is an early predictor of dengue warning signs and perturbs vascular integrity.

BackgroundA main pathological feature of severe dengue virus infection is endothelial hyper-permeability. The dengue virus nonstructural protein 1 (NS1) has been implicated in the vascular leakage that characterizes severe dengue virus infection, however, the molecular mechanisms involved are not known.MethodsA cohort of 250 dengue patients has been followed from the onset of symptoms to the recovery phase. Serum hyaluronan levels and several other clinical parameters were recorded. The effect of NS1 treatment of cultured fibroblasts and endothelial cells on the expressions of hyaluronan synthetic and catabolic enzymes and the hyaluronan receptor CD44, were determined, as have the effects on the formation of hyaluronan-rich matrices and endothelial permeability.FindingsElevated serum hyaluronan levels (≥70 ng/ml) during early infection was found to be an independent predictor for occurrence of warning signs, and thus severe dengue fever. High circulating levels of the viral protein NS1, indicative of disease severity, correlated with high concentrations of serum hyaluronan. NS1 exposure decreased the expression of CD44 in differentiating endothelial cells impairing the integrity of vessel-like structures, and promoted the synthesis of hyaluronan in dermal fibroblasts and endothelial cells in synergy with dengue-induced pro-inflammatory mediators. Deposited hyaluronan-rich matrices around cells cultured in vitro recruited CD44-expressing macrophage-like cells, suggesting a mechanism for enhancement of inflammation. In cultured endothelial cells, perturbed hyaluronan-CD44 interactions enhanced endothelial permeability through modulation of VE-cadherin and cytoskeleton re-organization, and exacerbated the NS1-induced disruption of endothelial integrity.InterpretationPharmacological targeting of hyaluronan biosynthesis and/or its CD44-mediated signaling may limit the life-threatening vascular leakiness during moderate-to-severe dengue virus infection.FundThis work was supported in part by grants from the Swedish Cancer Society (2018/337; 2016/445), the Swedish Research Council (2015-02757), the Ludwig Institute for Cancer Research, Uppsala University, the Ministry of Science and Technology, Taiwan (106–2314-B-037-088- and 106–2915-I-037-501-), Kaohsiung Medical University Hospital (KMUH103-3 T05) and Academy of Finland. The funders played no role in the design, interpretation or writing of the manuscript.

SAT-123 THE FRACTIONAL EXCRETION OF HYALURONAN AND SYNDECAN-1 IN DETECTING ENDOTHELIAL GLYCOCALYX DAMAGE IN KIDNEY DISEASE

The endothelial glycocalyx is a glycoprotein layer in the luminal surface on blood vessels with protective properties. Damage to this layer can be detected as an increase in the levels of its shed components such as hyaluronan and syndecan-1. Previous studies from our group have demonstrated elevated hyaluronan and syndecan-1 in patients with kidney disease. However, it is not clear whether an increase in these constituents in this population reflects damage to the glycocalyx or reduced renal clearance. Hyaluronan has a molecular weight of 4000kDa and is predominantly hepatically metabolised but is degraded into smaller fragments which can be detected in urine. The metabolism of syndecan-1 is unknown and has a variable molecular weight (32-200kDa) due to its attachment to variable heparan sulphate chain lengths. We aimed to determine the fractional excretion of hyaluronan and syndecan-1 in people with different levels of kidney function. We recruited healthy controls, CKD patients (eGFR 15-60), and kidney transplant recipients. Blood and spot urine samples were collected for enzyme-linked immunosorbent assays for hyaluronan (R&D Systems, Minneapolis) and syndecan-1 (Diaclone, France). Blood samples were centrifuged at 1000g for 15 minutes. Urine samples were centrifuged at 3000g for 10 minutes as well as 1000g for 15 minutes for comparison. Serum used for hyaluronan and syndecan-1 assays were diluted at 1:4 and 1:1, respectively. The fractional excretion of hyaluronan and syndecan were calculated from serum and urine levels using the formula: 100 × (urine level × serum creatinine)/(serum level × urine creatinine). Healthy controls (n=32), CKD patients (n=32) and transplants (n=36) had median eGFRs of 98 (70-127), 31 (10-59) and 59 (32-90) mL/min/1.73m2, p<0.0001, respectively. Liver function as measured by alanine transferase was normal in all groups (see table). No difference was detected in urinary hyaluronan and syndecan-1 values between urine centrifuged at 3000g or 1000g. Serum hyaluronan levels were highest in the CKD group, followed by transplant and healthy controls, whereas urine hyaluronan levels were unchanged between groups. The fractional excretion of hyaluronan was higher in healthy controls compared to CKD patients, and eGFR correlated negatively with serum hyaluronan (r=-0.47, p<0.0001); both indicating a higher clearance of hyaluronan with a higher GFR. Serum and urine syndecan-1 levels as well as the fractional excretion of syndecan were not different between groups. Furthermore, there was no correlation between serum syndecan-1 and eGFR. Both hyaluronan and syndecan-1 are detectable in urine which may represent the detection of smaller fragments of these molecules. The higher fractional excretion of hyaluronan in people with better kidney function suggests an element of renal excretion whereas this was not evident with syndecan-1. Given the normal liver function of this cohort and the known relatively small contribution of renal clearance of hyaluronan, the high serum levels in this study likely represent increased release from the endothelial glycocalyx. Syndecan-1 is present in renal tubular cells which may have affected the calculation of its fractional excretion. However, the absence of association between serum syndecan-1 and eGFR indicates little contribution of renal function to serum concentrations.

Hyaluronan and LYVE-1 and allograft function in lung transplantation recipients

Hyaluronan (HA) is associated with innate immune response activation and may be a marker of allograft dysfunction in lung transplant recipients. This was a prospective, single center study comparing levels of bronchioalveolar lavage (BAL) and serum HA and the HA immobilizer LYVE-1 in lung transplant recipients with and without acute cellular rejection (ACR). Chronic lung allograft dysfunction (CLAD)-free survival was also evaluated based on HA and LYVE-1 levels. 78 recipients were enrolled with a total of 115 diagnostic biopsies and 1.5 years of median follow-up. Serum HA was correlated with BAL HA (r = 0.25, p = 0.01) and with serum LYVE-1 (r = 0.32, p = 0.002). There was significant variation in HA and LYVE-1 over time, regardless of ACR status. Levels of serum HA (median 74.7 vs 82.7, p = 0.69), BAL HA (median 149.4 vs 134.5, p = 0.39), and LYVE-1 (mean 190.2 vs 183.8, p = 0.72) were not associated with ACR. CLAD-free survival was not different in recipients with any episode of elevated serum HA (HR = 1.5, 95% CI = 0.3–7.7, p = 0.61) or BAL HA (HR = 0.94, 95% CI = 0.2–3.6, p = 0.93). These results did not differ when stratified by bilateral transplant status. In this small cohort, serum HA, BAL HA, and LYVE-1 levels are not associated with ACR or CLAD-free survival in lung transplant recipients.