Missense Mutation in Human CHD4 Causes Ventricular Noncompaction by Repressing ADAMTS1.

BACKGROUND: Left ventricular noncompaction (LVNC) is a prevalent cardiomyopathy associated with excessive trabeculation and thin compact myocardium. Patients with LVNC are vulnerable to cardiac dysfunction and at high risk of sudden death. Although sporadic and inherited mutations in cardiac genes are implicated in LVNC, understanding of the mechanisms responsible for human LVNC is limited. METHODS: We screened the complete exome sequence database of the Pediatrics Cardiac Genomics Consortium and identified a cohort with a de novo CHD4 (chromodomain helicase DNA-binding protein 4) proband, CHD4M202I, with congenital heart defects. We engineered a humanized mouse model of CHD4M202I (mouse CHD4M195I). Histological analysis, immunohistochemistry, flow cytometry, transmission electron microscopy, and echocardiography were used to analyze cardiac anatomy and function. Ex vivo culture, immunopurification coupled with mass spectrometry, transcriptional profiling, and chromatin immunoprecipitation were performed to deduce the mechanism of CHD4M195I-mediated ventricular wall defects. RESULTS: CHD4M195I/M195I mice developed biventricular hypertrabeculation and noncompaction and died at birth. Proliferation of cardiomyocytes was significantly increased in CHD4M195I hearts, and the excessive trabeculation was associated with accumulation of ECM (extracellular matrix) proteins and a reduction of ADAMTS1 (ADAM metallopeptidase with thrombospondin type 1 motif 1), an ECM protease. We rescued the hyperproliferation and hypertrabeculation defects in CHD4M195I hearts by administration of ADAMTS1. Mechanistically, the CHD4M195I protein showed augmented affinity to endocardial BRG1 (SWI/SNF-related, matrix-associated, actin-dependent regulator of chromatin, subfamily A, member 4). This enhanced affinity resulted in the failure of derepression of Adamts1 transcription such that ADAMTS1-mediated trabeculation termination was impaired. CONCLUSIONS: Our study reveals how a single mutation in the chromatin remodeler CHD4, in mice or humans, modulates ventricular chamber maturation and that cardiac defects associated with the missense mutation CHD4M195I can be attenuated by the administration of ADAMTS1.

The C-terminal domains of ADAMTS1 contain exosites involved in its proteoglycanase activity.

A disintegrin-like and metalloproteinase with thrombospondin type 1 motifs (ADAMTS1) is a protease involved in fertilization, cancer, cardiovascular development, and thoracic aneurysms. Proteoglycans such as versican and aggrecan have been identified as ADAMTS1 substrates, and Adamts1 ablation in mice typically results in versican accumulation; however, previous qualitative studies have suggested that ADAMTS1 proteoglycanase activity is weaker than that of other family members such as ADAMTS4 and ADAMTS5. Here, we investigated the functional determinants of ADAMTS1 proteoglycanase activity. We found that ADAMTS1 versicanase activity is approximately 1000-fold lower than ADAMTS5 and 50-fold lower than ADAMTS4 with a kinetic constant (kcat/Km) of 3.6 × 103 M-1 s-1 against full-length versican. Studies on domain-deletion variants identified the spacer and cysteine-rich domains as major determinants of ADAMTS1 versicanase activity. Additionally, we confirmed that these C-terminal domains are involved in the proteolysis of aggrecan as well as biglycan, a small leucine-rich proteoglycan. Glutamine scanning mutagenesis of exposed positively charged residues on the spacer domain loops and loop substitution with ADAMTS4 identified clusters of substrate-binding residues (exosites) in ß3-ß4 (R756Q/R759Q/R762Q), ß9-ß10 (residues 828-835), and ß6-ß7 (K795Q) loops. This study provides a mechanistic foundation for understanding the interactions between ADAMTS1 and its proteoglycan substrates and paves the way for development of selective exosite modulators of ADAMTS1 proteoglycanase activity.

Investigation roles of Adamts1 and Adamts5 in scleral fibroblasts under hypoxia and mice with form-deprived myopia.

Scleral hypoxia is considered a trigger in scleral remodeling-induced myopia. Identifying differentially expressed molecules within the sclera is essential for understanding the mechanism of myopia. We developed a scleral fibroblast hypoxia model and conducted RNA sequencing and bioinformatic analysis. RNA interference technology was then applied to knock down targeted genes with upregulated expression, followed by an analysis of COLLAGEN I protein level. Microarray data analysis showed that the expression of Adamts1 and Adamts5 were upregulated in fibroblasts under hypoxia (t-test, p < 0.05). Western blot analysis confirmed increased protein levels of ADAMTS1 and ADAMTS5, and a concurrent decrease in COLLAGEN I in hypoxic fibroblasts. The knockdown of either Adamts1 or Adamts5 in scleral fibroblasts under hypoxia resulted in an upregulation of COLLAGEN I. Moreover, a form-deprivation myopia (FDM) mouse model was established for validation. The sclera tissue from FDM mice exhibited increased levels of ADAMTS1 and ADAMTS5 protein and a decrease in COLLAGEN I, compared to controls. The study suggests that Adamts1 and Adamts5 may be involved in scleral remodeling induced by hypoxia and the development of myopia.

ADAMTS-1 has nuclear localization in cells with epithelial origin and leads to decreased cell migration.

In the study of tumorigenesis, the involvement of molecules within the extracellular matrix (ECM) is crucial. ADAMTSs (A Disintegrin and Metalloproteinase with Thrombospondin motifs), a group of secreted proteases known for their role in ECM remodeling, were primarily considered to be extracellular proteases. However, our research specifically detected ADAMTS-1, a member of this family, predominantly within the nucleus of mammary cells. Our main objective was to understand the mechanism of ADAMTS-1 translocation to the nucleus and its functional significance in this cellular compartment. Our investigation uncovered that nuclear ADAMTS-1 was present in cells exhibiting an epithelial phenotype, while cells of mesenchymal origin contained the protease in the cytoplasm. Moreover, disruption of ADAMTS-1 secretion, induced by Monensin treatment, resulted in its accumulation in the cytoplasm. Notably, our research indicated that alterations in the secretory pathways could influence the protease's compartmentalization. Additionally, experiments with conditioned medium from cells containing nuclear ADAMTS-1 demonstrated its internalization into the nucleus by HT-1080 cells and fibroblasts. Furthermore, heightened levels of ADAMTS-1 within the ECM reduced the migratory potential of mesenchymal cells. This highlights the potential significance of nuclear ADAMTS-1 as a critical component within the tumor microenvironment due to its functional activity in this specific cellular compartment.

The oncogenic ADAMTS1-VCAN-EGFR cyclic axis drives anoikis resistance and invasion in renal cell carcinoma.

BACKGROUND: Metastasis, the leading cause of renal cell carcinoma (RCC) mortality, involves cancer cells resisting anoikis and invading. Until now, the role of the matrix metalloproteinase (MMP)-related enzyme, A disintegrin and metalloprotease with thrombospondin motifs 1 (ADAMTS1), in RCC anoikis regulation remains unclear. METHODS: The clinical significance of ADAMTS1 and its associated molecules in patients with RCC was investigated using data from the Gene Expression Omnibus (GEO) and TCGA datasets. Human phosphoreceptor tyrosine kinase (RTK) array, luciferase reporter assays, immunoprecipitation (IP) assays, western blotting, and real-time reverse-transcription quantitative polymerase chain reaction (RT-qPCR) were used to elucidate the underlying mechanisms of ADAMTS1. Functional assays, including anoikis resistance assays, invasion assays, and a Zebrafish xenotransplantation model, were conducted to assess the roles of ADAMTS1 in conferring resistance to anoikis in RCC. RESULTS: This study found elevated ADAMTS1 transcripts in RCC tissues that were correlated with a poor prognosis. ADAMTS1 manipulation significantly affected cell anoikis through the mitochondrial pathway in RCC cells. Human receptor tyrosine kinase (RTK) array screening identified that epidermal growth factor receptor (EGFR) activation was responsible for ADAMTS1-induced anoikis resistance and invasion. Further investigations revealed that enzymatically active ADAMTS1-induced versican V1 (VCAN V1) proteolysis led to EGFR transactivation, which in turn, through positive feedback, regulated ADAMTS1. Additionally, ADAMTS1 can form a complex with p53 to influence EGFR signaling. In vivo, VCAN or EGFR knockdown reversed ADAMTS1-induced prometastatic characteristics of RCC. A clinical analysis revealed a positive correlation between ADAMTS1 and VCAN or the EGFR and patients with RCC with high ADAMTS1 and VCAN expression had the worst prognoses. CONCLUSIONS: Our results collectively uncover a novel cyclic axis involving ADAMTS1-VCAN-EGFR, which significantly contributes to RCC invasion and resistance to anoikis, thus presenting a promising therapeutic target for RCC metastasis.

ADAMTS1 as potential prognostic biomarker promotes malignant invasion of glioma.

BACKGROUND: Glioma is the most common intracranial malignancy in adults with a high degree of malignancy and poor prognosis, which is largely attributed to the existence of glioma stem cells (GSCs). Previous evidence indicated that the matrix metalloproteinase ADAMTS1 was implicated in the process of tumor invasion, but the involvement of ADAMTS1 in glioma malignant invasion remains poorly understood. METHODS: The expression and prognosis values of ADAMTS1 were investigated in patients with glioma based on ONCOMINE and GEPIA databases. ADAMTS1 expression of different malignancy grade tissues was determined by immunohistochemistry. The effects of ADAMTS1 on cell proliferation and invasion were determined by clone formation assay and Transwell migration assay. The animal experiment was performed in an intracranial orthotopic xenograft model by knockout of ADAMTS1. Stemness properties and Notch1-SOX2 pathway were examined in stable ADAMTS1 knockdown GSCs. RESULTS: The expression levels of ADAMTS1 were significantly higher in glioma tissues and significantly correlated with the grade of malignancy and prognosis of glioma. Elevated ADAMTS1 expression was associated with SOX2, N-cadherin and the resistance of chemoradiotherapy of glioma patients. ADAMTS1 knockout suppressed the intracranial orthotopic xenograft growth and prolonged the survival of xenograft mice in vivo. Mechanistically, we found a blockade of the migration and invasiveness of GSCs and the expression levels of Notch1 and SOX2 in absence of ADAMTS1. CONCLUSION: As a biomarker for prediction of prognosis, ADAMTS1 may affect the invasive phenotype of GSCs by regulating Notch1-SOX2 signaling pathway, thereby promoting the invasive growth of glioma.

ADAMTS-1 and syndecan-4 intersect in the regulation of cell migration and angiogenesis.

ADAMTS-1 is an extracellular protease with critical roles in organogenesis and angiogenesis. Here we demonstrate a functional convergence of ADAMTS-1 and the transmembrane heparan sulfate proteoglycan syndecan-4 in influencing adhesion, migration and angiogenesis. Knockdown of ADAMTS-1 in endothelial cells resulted in a parallel reduction in cell surface syndecan-4, attributable to increased matrix metalloproteinase-9 (MMP9) activity. Knockdown of either ADAMTS-1 or syndecan-4 increased cellular responses to vascular endothelial growth factor A isoform VEGFA164, and increased ex vivo aortic ring microvessel sprouting. On fibronectin, knockdown of either protein enhanced migration and promoted formation of long α5 integrin-containing fibrillar adhesions. However, integrin α5 null cells still showed increased migration in response to ADAMTS-1 and syndecan-4 siRNA treatment. Plating of naïve endothelial cells on cell-conditioned matrix from ADAMTS-1 and syndecan-4 knockdown cells demonstrated that the altered adhesive behaviour was matrix dependent, and this correlated with a lack of expression of fibulin-1: an extracellular matrix co-factor for ADAMTS-1 that is known to inhibit migration. These findings support the notion that ADAMTS-1 and syndecan-4 are functionally interconnected in regulating cell migration and angiogenesis, via collaboration with MMP9 and fibulin-1.This article has an associated First Person interview with the first author of the paper.

The role of serum ADAMTS-1 levels in Hyperemesis Gravidarum.

BACKGROUND: We aimed to investigate the levels of ADAMTS-1, which is secreted from the extracellular matrix during trophoblastic invasion in hyperemesis gravidarum (HEG). METHODS: In this cross-sectional study, we compared 45 HEG patients aged between 21 and 34 in terms of ADAMTS-1 levels with a control group consisting of 44 healthy pregnant women. The demographic characteristics and several laboratory parameters of the patients were recorded. Both groups were also compared in terms of ketonuria. We evaluated the correlation between ADAMTS-1 levels and ketonuria. RESULTS: The 2 groups were matched in terms of age, gestational age, gravidity, parity, and body mass index. Some inflammatory markers, such as neutrophil count, MPV, PDW, and PCT levels, were significantly higher in the HEG groups compared to the control group (all p < 0.05). However, mean MCV and serum TSH levels were statistically significantly lower in this group (both p < 0.001). ADAMTS-1 levels were 12.6 ± 1.4 ng/ml in the HEG group and 6.2 ± 1.6 ng/ml in the control group (p < 0.001). It was significantly and positively correlated with urine ketone, neutrophil count, and PDW, whereas negatively correlated with MCV and TSH value in the HEG group. ROC analysis showed that a threshold value of 11.275 ng/ml for ADAMTS-1 predicted HEG patients with a sensitivity of 60% and specificity of 95.5%. CONCLUSION: ADAMTS-1 serum levels are increased in HEG patients, and there is a positive correlation between ADAMTS-1 levels and ketonuria.

ADAMTS1 is regulated by the EP4 receptor in the zebrafish ovary.

Prostaglandins (PGs) are a class of fatty-acid derived hormones that are essential in ovulation of teleosts, but their exact role remains unknown. One putative target of PGs in ovulation is regulation of the expression of members of the A Disintegrin and Metalloproteinase with Thrombospondin motifs (ADAMTS) family, which are implicated in follicular rupture. This study investigated the regulation of ADAMTS, other proteases, and their inhibitors in response to treatment with PGE2 or PGF2α. Four members of the ADAMTS family, ADAMTS1, ADAMTS5, ADAMTS9, and ADAMTS16 were shown to be expressed in the ovary of zebrafish, but only adamts1 was upregulated in full-grown follicles following treatment with PGE2. Inhibitors of the PG receptors EP1 and EP2 had no effect on PGE2-stimulated adamts1 expression, while treatment of full-grown follicles with both PGE2 and GW627368x, an inhibitor of EP4 function, prevented the PGE2-induced increase in adamts1 expression. Treatment of full-grown follicles with the maturation-inducing hormone 17α,20ß-dihydroxy-4-pregnen-3-one (17,20ß-P) in vitro had no effect on the expression of adamts1 mRNA. These findings suggest that expression of ADAMTS1 in zebrafish ovarian follicles is regulated by the prostaglandin PGE2 via the EP4 series prostaglandin receptor.

Comprehensive bioinformatics analysis identifies lncRNA HCG22 as a migration inhibitor in esophageal squamous cell carcinoma.

Esophageal cancer is one of the most lethal malignancies worldwide, and esophageal squamous cell carcinoma (ESCC) is the dominant histological type. However, the long noncoding RNA (lncRNA) alterations in ESCC have not been elucidated to date. In this study, reliable databases from Gene Expression Omnibus (GEO), which analyzed lncRNA expression in ESCC tumor tissues and adjacent normal tissues were searched, and common differentially expressed lncRNAs and genes were analyzed. Next, cis- trans analysis was performed to predict the underlying relationships between altered lncRNAs and mRNAs, and the lncRNA-mRNA regulatory network was established. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses of altered lncRNA-related genes were performed. The promising lncRNA HCG22 was validated by quantitative polymerase chain reaction (qPCR), and clinicopathological data were collected to identify the relationship between lncRNA HCG22 expression level and clinical features. Finally, Transwell assays were performed to explore the biological functions of lncRNA HCG22 in ESCC cells. Two hundred forty-one lncRNAs and 835 mRNAs were observed to be remarkably altered between ESCC tumor tissues and adjacent normal tissues. The lncRNA-mRNA regulatory network showed the coexpression association between lncRNA HCG22 and SPINK7 and ADAMTS12. GO and KEGG analyses showed that HCG22 and ADAMTS12 had potential biological functions in the cell migration of ESCC. The downregulation of lncRNA HCG22 in ESCC tumor tissues was validated by qPCR, and the clinicopathological data showed a noticeable correlation between lncRNA HCG22 expression level and the ESCC differentiational degree and clinical TNM stage. Kaplan-Meier analysis showed that patients with ESCC having low lncRNA HCG22 expression in ESCC tissues had considerably shorter overall survival compared with patients with ESCC having high lncRNA HCG22 expression. Following Transwell assays confirmed the migratory role of lncRNA HCG22 in ESCC cells. In conclusion, lncRNA HCG22 was downregulated in ESCC tissues and can be a migration inhibitor of ESCC cells, and SPINK7 and ADAMTS12 are promising to be the regulatory targets of lncRNA HCG22.

Melatonin-triggered post-transcriptional and post-translational modifications of ADAMTS1 coordinately retard tumorigenesis and metastasis of renal cell carcinoma.

A disintegrin and metalloprotease with thrombospondin motifs (ADAMTS) family are widely implicated in tissue remodeling events manifested in cancer development. ADAMTS1, the most fully characterized ADAMTS, plays conflicting roles in different cancer types; however, the role of ADAMTS1 in renal cell carcinoma (RCC) remains unclear. Herein, we found that ADAMTS1 is highly expressed in RCC tissues compared to normal renal tissues, and its expression was correlated with an advanced stage and a poor prognosis of RCC patients. In vitro, we observed higher expression of ADAMTS1 in metastatic (m)RCC cells compared to primary cells, and manipulation of ADAMTS1 expression affected cell invasion and clonogenicity. Results from protease array showed that ADAMTS1 is modulated by melatonin through mechanisms independent of the MT1 receptor in mRCC cells, and overexpression of ADAMTS1 relieved the invasion/clonogenicity and growth/metastasis inhibition imposed by melatonin treatment in vitro and in an orthotopic xenograft model. The human microRNA (miR) OneArray showed that miR-181d and miR-let-7f were induced by melatonin and, respectively, targeted the 3'-UTR and non-3'-UTR of ADAMTS1 to suppress its expression and mRCC invasive ability. Clinically, RCC patients with high levels of miR-181d or miR-let-7f and a low level of ADAMTS1 had the most favorable prognoses. In addition, ubiquitin/proteasome-mediated degradation of ADAMTS1 can also be triggered by melatonin. Together, our study indicates that ADAMTS1 may be a useful biomarker for predicting RCC progression. The novel convergence between melatonin and ADAMTS1 post-transcriptional and post-translational regulation provides new insights into the role of melatonin-induced molecular regulation in suppressing RCC progression.

ADAMTS1 and HSPG2 mRNA levels in cumulus cells are related to human oocyte quality and controlled ovarian hyperstimulation outcomes.

PURPOSE: The study investigated potential correlations between the expression levels of ADAMTS1 and HSPG2 in cumulus cells (CCs) and controlled ovarian hyperstimulation (COH) outcomes. METHODS: RT-PCR was used to determine ADAMTS1 and HSPG2 mRNA levels in mice CCs at different timepoints (0, 4, 8, 12, and 16 h) after human chorionic gonadotropin (hCG) injection, and in CCs after RNAi treatment. Women with polycystic ovary syndrome (PCOS) (n = 45) and normal ovulatory controls (n = 103) undergoing IVF/ICSI were recruited. Relative ADAMTS1 and HSPG2 mRNA levels were measured by RT-PCR. Moreover, correlations of ADAMTS1 and HSPG2 levels with COH outcomes were analyzed. RESULTS: At different timepoints after hCG treatment, ADAMTS1 mRNA had the highest level at 12 h, whereas HSPG2 showed opposite profiles to ADAMTS1 with the lowest level at 12 h. HSPG2 expression was upregulated after ADAMTS1 RNAi treatment The PCOS group had higher HSPG2 and lower ADAMTS1 expression levels than controls. In normal ovulatory women (control group), a higher expression of ADAMTS1 and lower expression of HSPG2 were associated with more mature oocytes, transplantable embryos, and good quality embryos, whereas higher transplantable embryo rates and good quality embryo rates were obtained only with lower HSPG2 expression. ROC curves showed the co-measurement of ADAMTS1 and HSPG2 had a better predictive power than separate analyses. CONCLUSION: The dynamic profiles of ADAMTS1 and HSPG2 were inversely correlated in CCs. In PCOS and normal ovulatory patients, higher ADAMTS1 and lower HSPG2 expression levels in CCs were related to better COH outcomes.

ADAMTS1 Promotes Adhesion to Extracellular Matrix Proteins and Predicts Prognosis in Early Stage Breast Cancer Patients.

BACKGROUND/AIMS: Despite, several studies demonstrating pro-metastatic effects of the metalloproteinase ADAMTS1 in breast cancer, its role in facilitating the metastatic cascade remains unclear. To date there have been limited studies that have examined the expression of ADAMTS1 in primary and metastatic breast cancer tissues. METHODS: We assessed ADAMTS1 mRNA levels in publically available breast cancer sets and analysed ADAMTS1 protein levels by immunohistochemistry in breast tissue microarrays containing normal breast tissue (n=9), primary invasive ductal breast carcinomas (n=79) and metastatic lesions (n=58). To understand the underlying events influenced by ADAMTS1 and provide a mechanism by which tumors expressing this protease promote metastasis, we assessed the ability of PyMT/Adamts1+/+, PyMT/Adamts1+/- and PyMT/Adamts1-/- primary mammary cancer cells to adhere to matrigel and migrate or invade towards a chemoattractive environment. RESULTS: High ADAMTS1 expression was associated with reduced disease-free survival, distant metastasis free-survival and overall survival in breast cancer patients with node negative disease. Although ADAMTS1 expression was reduced in primary breast cancers compared to normal tissue and not elevated in metastatic lesions, high ADAMTS1 immunostaining was associated with a higher number of positive lymph nodes (p=0.006) and the presence of distant metastasis (p=0.023). Depletion of Adamts1 in mammary cancer cells impeded their adhesion to a biological matrix substratum and diminished cell migration but not invasion. CONCLUSION: The effects observed on cell adhesion and migration demonstrates a potential mechanism whereby ADAMTS1 promotes breast cancer metastasis.

Role of ADAMTS-5 in Aortic Dilatation and Extracellular Matrix Remodeling.

OBJECTIVE: Thoracic aortic aneurysm (TAA), a degenerative disease of the aortic wall, is accompanied by changes in the structure and composition of the aortic ECM (extracellular matrix). The ADAMTS (a disintegrin and metalloproteinase with thrombospondin motifs) family of proteases has recently been implicated in TAA formation. This study aimed to investigate the contribution of ADAMTS-5 to TAA development. APPROACH AND RESULTS: A model of aortic dilatation by AngII (angiotensin II) infusion was adopted in mice lacking the catalytic domain of ADAMTS-5 (Adamts5Δcat). Adamts5Δcat mice showed an attenuated rise in blood pressure while displaying increased dilatation of the ascending aorta (AsAo). Interestingly, a proteomic comparison of the aortic ECM from AngII-treated wild-type and Adamts5Δcat mice revealed versican as the most upregulated ECM protein in Adamts5Δcat mice. This was accompanied by a marked reduction of ADAMTS-specific versican cleavage products (versikine) and a decrease of LRP1 (low-density lipoprotein-related protein 1). Silencing LRP1 expression in human aortic smooth muscle cells reduced the expression of ADAMTS5, attenuated the generation of versikine, but increased soluble ADAMTS-1. A similar increase in ADAMTS-1 was observed in aortas of AngII-treated Adamts5Δcat mice but was not sufficient to maintain versican processing and prevent aortic dilatation. CONCLUSIONS: Our results support the emerging role of ADAMTS proteases in TAA. ADAMTS-5 rather than ADAMTS-1 is the key protease for versican regulation in murine aortas. Further studies are needed to define the ECM substrates of the different ADAMTS proteases and their contribution to TAA formation.

Pro-atherogenic proteoglycanase ADAMTS-1 is down-regulated by lauric acid through PI3K and JNK signaling pathways in THP-1 derived macrophages.

The prevalence of atherosclerosis has increased significantly in the recent years due to sedentary lifestyle and high-fat diet. However, the association between saturated fat intake and the increased risk for atherosclerotic cardiovascular diseases remains heavily debated. Lauric acid belongs to the saturated fatty acid group and its unique medium chain fatty acid properties are proven to be beneficial to humans in many ways. Thus, the aim of this project is to investigate the effect of lauric acid on the expression of a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS) genes-ADAMTS-1, ADAMTS-4, and ADAMTS-5-in macrophages. These genes encode for proteases that participate in the extracellular matrix remodeling and they play important roles in the vulnerability of atherosclerotic plaque. Here, we show that the treatment of 20 µM of lauric acid successfully reduced both transcriptional and translational expressions of these genes in THP-1 differentiated macrophages after 24-h incubation. Further cell signaling experiments using a panel of kinase inhibitors and phosphorylated antibodies proved that lauric acid down-regulated ADAMTS-1 by reducing the activation of PI3K and JNK at Tyr458 and Tyr185, respectively. Finally, JNK1 siRNA knockdown assay confirmed that ADAMTS-1 was regulated through JNK pathway, and lauric acid interfered with this pathway to down-regulate ADAMTS-1 expression. Although preliminary, this present study indicates that lauric acid has the potential to stabilize atherosclerotic plaque and may prevent thrombosis by interfering with the ADAMTS-1 expression through PI3K/JNK pathways.

ADAMTS-1 disrupts HGF/c-MET signaling and HGF-stimulated cellular processes in fibrosarcoma.

Extracellular matrix (ECM) serves as a reservoir for biologically active factors, such as growth factors and proteases that influence the tumor cell behavior. ADAMTS-1 (a disintegrin and metalloprotease with thrombospondin motifs) is a secreted protease that has the ability to modify the ECM during physiological and pathological processes. Here, we analyzed the role played by ADAMTS-1 regulating HGF and TGF-ß1 activities in the high-grade fibrosarcoma cell line (HT1080). We generated HT1080 and HEK293T cells overexpressing ADAMTS-1. HT1080 cells overexpressing ADAMTS-1 (HT1080-MPA) exhibited a significant decrease in cell proliferation and migration velocity, both in presence of HGF. We obtained similar results with ADAMTS-1-enriched conditioned medium from other cell type. However, ADAMTS-1 overexpression failed to affect TGF-ß1 activity associated with HT1080 cell proliferation and migration velocity. Immunoblotting showed that ADAMTS-1 overexpression disturbs c-Met activation upon HGF stimulation. Downstream ERK1/2 and FAK signaling pathways are also influenced by this protease. Additionally, ADAMTS-1 decreased the size of the fibrosarcospheres, both under normal conditions and in the presence of HGF. Likewise, in presence of HGF, ADAMTS-1 overexpression in HT1080 disrupted microtumors formation in vivo. These microtumors, including individual cells, presented characteristics of non-invasive lesions (rounded morphology). Our results suggest that ADAMTS-1 is involved in regulating HGF-related functions on fibrosarcoma cells. This protease may then represent an endogenous mechanism in controlling the bioavailability of different growth factors that have a direct influence on tumor cell behavior.

Abnormal expressions of ADAMTS-1, ADAMTS-9 and progesterone receptors are associated with lower oocyte maturation in women with polycystic ovary syndrome.

PURPOSE: ADAMTS-1 and 9 play a crucial role in the ovulation and their altered levels may play a role in the pathogenesis of polycystic ovary syndrome (PCOS). The aim of this study was to assess ADAMTS-1 and 9 expression and their correlation with the oocyte quality and maturity in the cumulus cells (CCs) of PCOS patients and normovulatory women during an IVF procedure. METHODS: Expression of ADAMTS-1 and 9 and progesterone receptors (PRs) in the CCs containing MII and germinal vesicle (GV) oocytes of 37 PCOS patients and 37 women with normal ovulatory function who underwent IVF treatment was evaluated using qRT-PCR. Moreover, correlation between ADAMTS-1 and 9 expression and oocyte quality were also investigated. RESULTS: mRNA expression levels of ADAMTS-1 and ADAMTS-9 were significantly reduced in the women with PCOS compared to the normovulatory women. ADAMTS-1 and ADAMTS-9 mRNA expression levels in the CCs showed a considerable correlation. Lower expression levels of ADAMTS-1 and ADAMTS-9 in PCOS patients were strongly correlated with diminished oocyte maturation. There was a remarkable association between ADAMTS-1 and ADAMTS-9 mRNA expression levels and oocyte quality. PRs (PRA and PRB) were dramatically decreased in PCOS patients when compared with the control group. CONCLUSIONS: The results of the present study indicated that ADAMTS-1 and ADAMTS-9 as well as PRs are downregulated in the human CCs in PCOS patients, which could be associated with impaired oocyte maturation and may result in a lower oocyte recovery and oocyte maturity rates, as well as lower fertilization rate.

Decreased ADAMTS-1, -9 and -20 levels in women with endometrial polyps: a possible link between extracellular matrix proteases and endometrial pathologies.

A disintegrin-like and metalloproteinase domain with thrombospondin-type 1 motifs (ADAMTS) protein superfamily includes 19 secreted metalloproteases. Proteolytic substrates of ADAMTS enzymes have been linked to female reproductive function. Herein, we aimed to investigate serum ADAMTS-1, -9 and -20 levels in women with and without endometrial polyps (EPs). The study group (n = 40) consisted of women who had hysteroscopically detected and histologically confirmed EPs whereas control group (n = 40) was recruited from those women without any endometrial pathology. Data recorded for every woman were as follows: age, body mass index, gravidity and parity, number of miscarriages, smoking status and serum ADAMTS-1, -9 and -20 levels. ADAMTS-1, -9 and -20 values were measured by commercially available ELISA kits. No statistically significant differences between the groups were observed in terms of demographics. There were also no statistically significant differences between the groups with regard to ADAMTS-1 and -20 levels, although both of them were lower in the study group. However, ADAMTS-9 was significantly lower in the study group compared to the controls (p = .010). The optimal cut off value of ADAMTS-9 in predicting EPs was found to be 163.2 ng/mL with 100% sensitivity and 35% specificity. In conclusion, ADAMTS-9 protein is decreased in women with EPs. Impact statement What is already known on this subject? Endometrial polyps (EPs) are common and are generally benign gynaecologic disorders. ADAMTS enzymes comprise a zinc metalloproteinase gene family that has roles in vascular biology, inflammation and especially in the control of the function and structure of the extracellular matrix (ECM). ECM plays an important role in the pathogenesis of myomas, adenomyosis and abnormal uterine bleeding, as well as EPs. There is an interest in these proteases, especially with regard to the physiology of ovulation and implantation. They are also associated with carcinogenesis and metastasis. One of the most feared consequences of EPs is the risk of malignancy. Therefore, it is important in gynaecology practice to diagnose these endometrial abnormalities. What do the results of this study add? This is the first study performed to investigate the relationship between some ADAMTS (-1, -9 and -20) proteases and uterine polyps. Our results demonstrate novel molecular mediators contributing to EPs physiopathology. What are the implications of these findings for clinical practice and/or further research? ADAMTS-9 is defined as a tumour suppressor gene in various malignancies. Decreased ADAMTS-9 protein, which is the product of this gene, may have a role in the pathogenesis of EPs. There is a need for further research that should be done with benign-malign EPs.

Prodigiosin Alleviates Pulmonary Fibrosis Through Inhibiting miRNA-410 and TGF-ß1/ADAMTS-1 Signaling Pathway.

BACKGROUND/AIMS: Pulmonary fibrosis is a common outcome of various interstitial lung diseases. Prodigiosin (PG) is a series of red pigment with methoxypyrrole ring. This studyinvestigates therole of prodigiosin in pulmonary fibrosis and its underlying mechanisms. METHODS: A pulmonary fibrosis rat model was established by intra-trachealinjection ofbleomycin A5. Rats were divided into 4 groups: Normal group, pulmonary fibrosis Model group, Prodigiosin treatment group and hydrocortisone treatmentgroup. HE and Masson staining were carried outto evaluate histopathological changes. The content of hydroxyproline in lung tissue was determined by alkaline hydrolysis. The expression of PICP and PIIINP was examined by ELISA. The mRNA expression of miR-410, TGF-ß1 and ADAMTS1 in lung homogenate were detected by RT-PCR. The bronchoalveolar lavage fluid (BALF) and lung tissues of rats were collected and analyzed. Human embryonic pulmonary fibroblast (HEPF) was used for study in vitro. A dual-luciferase reporter assay was conducted to examine the effect of miR-410 on ADAMTS1 expression. Cell transfection was conducted to inhibit miR-410. MTT assay was performed to investigate cell proliferation. The expressions of miR-410, TGF-ß1, ADAMTS1and other fibrosis related biomarkers (Col I, Col III, and α-SMA) wereexamined by RT-PCR and Western Blot. RESULTS: HE and Masson staining showed thickened alveolar septum, hyperplasticcapillaries, and large areas of collagen fiber deposition in pulmonary fibrosis model rats. Rats in prodigiosin and hydrocortisone treatment groups had alleviated symptoms. There was high hydroxyproline expression in model rats, whereas the expression of hydroxyproline reduced after prodigiosin or hydrocortisone treatments. RT-PCR results showed high miR-410,high TGF-ß1 and low ADAMTS1 in lung tissue of model rats. The expression of PICP and PIIINP werehigher in BALF of model group than in treatment groups. Prodigiosin and hydrocortisone treatment significantly reduced PICP and PIIINP content. RT-PCR and Western Blot analysis showed that prodigiosin inhibited expression of miR-410 and TGF-ß1, but up-regulated ADAMTS1 expression. MTT assay indicated that prodigiosin inhibited HEPF proliferation induced by miR-410 overexpression. CONCLUSION: Prodigiosin down-regulates the expression of miR-410 and TGF-ß1, up-regulates ADAMTS1, leading to decrease accumulation of fibrotic proteins. It could be used in alleviating pulmonary fibrosis.

Postnatal deficiency of ADAMTS1 ameliorates thoracic aortic aneurysm and dissection in mice.

NEW FINDINGS: What is the central question of this study? Thoracic aortic aneurysm and dissection (TAAD) is characterized by extracellular matrix remodelling and an inflammatory response. Evidence suggests that ADAMTS1 is closely associated with TAAD development, but whether it contributes to the pathophysiology of TAAD remains unknown. What is the main finding and its importance? We generated inducible postnatal ADAMTS1 knockout mice and found that ADAMTS1 deficiency attenuated ß-aminopropionitrile-dependent TAAD formation and rupture. Furthermore, ADAMTS1 deficiency suppressed neutrophil and macrophage infiltration by inhibiting inflammatory cytokine levels and macrophage migration during the early stage of ß-aminopropionitrile-induced TAAD. ADAMTS1 could be a new therapeutic target for TAAD. ABSTRACT: Thoracic aortic aneurysm and dissection (TAAD), as a life-threatening cardiovascular disease, is characterized by extracellular matrix remodelling and an inflammatory response. A disintegrin and metalloproteinase with thrombospondin motifs 1 (ADAMTS1) is an inflammation-related protein that is able to degrade extracellular matrix proteins in arteries. Herein, we investigated whether ADAMTS1 contributes to the pathophysiology of TAAD in mice. Using the mouse model of ß-aminopropionitrile (BAPN)-induced TAAD, we found that ADAMTS1 expression was upregulated beginning in the early stage of TAAD development and localized predominantly in the aortic adventitia. ADAMTS1-floxed mice and whole-body tamoxifen-inducible ADAMTS1 knockout mice (ADAMTS1flox/flox Ubc-CreERT2+ , ADAMTS1 KO) were generated to investigate the direct causal role of ADAMTS1 in TAAD development. The incidence and rupture rates of BAPN-induced TAAD in ADAMTS1 KO mice were significantly lower than those in ADAMTS1flox/flox mice (45.5 versus 81.8% and 18.2 versus 42.4%, respectively). Aortas from BAPN-treated ADAMTS1flox/flox mice displayed profound destruction of the elastic lamellae, abundant neutrophil and macrophage accumulation in the adventitia, obviously increased neutrophil proportions in peripheral blood and significantly increased expression of inflammatory factors in the early stage of TAAD induction, all of which were markedly suppressed in ADAMTS1 KO mice. Furthermore, ADAMTS1-deficient macrophages exhibited abrogated migration capacity both in vivo and in vitro. In conclusion, ADAMTS1 plays a crucial role in postnatal TAAD formation and rupture by regulating inflammatory responses, suggesting that ADAMTS1 might be a new therapeutic target for TAAD.