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A disintegrin and metallopeptidase with thrombospondin motifs (ADAMTSs), which is frequently dysregulated in cancers and is involved in carcinogenesis and cancer progression. The present study identified that ADAMTS8 expression is downregulated in esophageal squamous cell carcinoma (ESCC) tissues when compared with nontumor tissue. The expression of ADAMTS8 is closely associated with clinical stage and lymph node metastasis in patients with ESCC. Furthermore, functional studies have shown that ADAMTS8 overexpression could reduce abilities of proliferation, migration, and invasion and promote apoptosis of ESCC cells. Meanwhile, monocyte chemotactic protein-1 and interleukin-6 are markedly deregulated by ADAMTS8 overexpression. Consistently, in vivo data showed that ADAMTS8 overexpression led to a reduction in tumor growth. These results indicate that altering ADAMTS8 expression could modify the outcomes of ESCC by inhibiting cell proliferation and invasion, while promoting the apoptosis of ECSS cells. Thus, ADAMTS8 represents a potential therapeutic target for ESCC therapy.

Introduction A disintegrin and metallopeptidase with thrombospondin motifs (ADAMTSs), whose expression is dysregulated in various cancers, is implicated in cancer development. Herein, we aimed to investigate the functional role of ADAMTS8 in breast cancer (BC) and explore the underlying mechanisms. Methods The protein expression of ADAMTS8 in BC cell lines and tumor tissues from BC patients was quantified by Western blot. ADAMTS8 overexpression was induced by transfection with pEZ-M90-ADAMTS8 plasmid using lipofectamine 2000. To generate ADAMTS8 stable knockdown cells, MDA-MB-231 cells were transfected with psi-H1-ADAMTS8siRNA plasmids. Cell counting kit-8 (CCK-8) assay, wound-healing assay, transwell assay and flow cytometry assay were employed to analyze the effects of ADAMTS8 on the proliferation, migration, invasion and apoptosis of BC cells. Chemosensitivity also was assessed using CCK-8 assay. The expressions of β-catenin, MMP-7 and c-Myc were measured by Western blot. Results Our results showed that ADAMTS8 expression was significantly lower in BC tissues than that in adjacent non-tumor tissues. Overexpression of ADAMTS8 in MDA-MB-453 cells could inhibit the cell proliferation, migration and invasion and promote apoptosis. ADAMTS8 knockdown displayed the reverse effect in MDA-MB-231 cells. Consistently, in vivo data showed that ADAMTS8 overexpression led to a reduction in tumor growth. In addition, chemosensitivity testing in MDA-MB-453 cells transfected with pEZ-M90-ADAMTS8 plasmid indicated that cisplatin inhibited cell growth dramatically. Furthermore, attenuated β-catenin, MMP-7 and c-Myc level was detected after ADAMTS8 overexpression. Conclusion These results indicate that increased ADAMTS8 expression could modify the progression of BC by inhibiting cell proliferation and invasion while promoting the apoptosis of BC cells. Thus, ADAMTS8 represents a potential therapeutic target for BC therapy.

Myocardial infarction or pressure overload leads to cardiac fibrosis, the leading cause of heart failure. ADAMTS8 (A disintegrin and metalloproteinase with thrombospondin motifs 8) has been reported to be involved in many fibrosis-related diseases. However, the specific role of ADAMTS8 in cardiac fibrosis caused by myocardial infarction or pressure overload is yet unclear. The present study aimed to explore the function of ADAMTS8 in cardiac fibrosis and its underlying mechanism. ADAMTS8 expression was significantly increased in patients with dilated cardiomyopathy; its expression myocardial infarction and TAC rat models was also increased, accompanied by increased expression of α-SMA and Collagen1. Adenovirus-mediated overexpression of ADAMTS8 through cardiac in situ injection aggravated cardiac fibrosis and impaired cardiac function in the myocardial infarction rat model. Furthermore, in vitro studies revealed that ADAMTS8 promoted the activation of cardiac fibroblasts; ADAMTS8 acted as a paracrine mediator allowing for cardiomyocytes and fibroblasts to communicate indirectly. Our findings showed that ADAMTS8 could damage the mitochondrial function of cardiac fibroblasts and then activate the PI3K-Akt pathway and MAPK pathways, promoting up-regulation of YAP expression, with EGFR upstream of this pathway. This study systematically revealed the pro-fibrosis effect of ADAMTS8 in cardiac fibrosis and explored its potential role as a therapeutic target for the treatment of cardiac fibrosis and heart failure.

Rheumatoid arthritis (RA), chronic, autoimmune causing joint stiffness, pain, fatigue and functional disorders is a disease. The aim of this study is to investigate the presence of ADAMTS-8 expression in cells in synovial fluid in rheumatoid arthritis patients. Patients with active RA were aspirated with arthro synthesis from the knee joint. Blood samples were taken from the patients at the same time, plasma samples were separated. Patients with sinovial fluid treated between 2019-2020 year in the Gazi Yasargil Hospital Physical Therapy and Rehabilitation Clinic were involved in the study that was consulted with knee pain and difficulty in performing their daily activities. Informed consent form was obtained from all of the patients (10 RA and 10 patients). CRP, RF and ESH results were statistically evaluated in patients with Control and Rheumatoid arthritis. CRP, RF and ESH results were statistically evaluated in patients with Control and Rheumatoid arthritis. In patient samples with Rheumatoid arthritis, degeneration in synovial cells was observed in some nuclei in the form of pycnosis and apoptotic. ADAMTS-8 expression in the form of aggregates in synovial cells was found to be positive. It was observed that rheumatoid arthritis increased apoptotic change in synovial cells. ADAMTS8 in synovial cells in rheumatoid arthritis cases may cause a significant increase in apoptosis levels by suppressing some signal pathways, and predicts that apoptosis induction may play a role in the inhibition of ADAMTS8-mediated synovial cell proliferation.

Introduction A disintegrin and metallopeptidase with thrombospondin motifs (ADAMTSs), whose expression is dysregulated in various cancers, is implicated in cancer development. Herein, we aimed to investigate the functional role of ADAMTS8 in breast cancer (BC) and explore the underlying mechanisms. Methods The protein expression of ADAMTS8 in BC cell lines and tumor tissues from BC patients was quantified by Western blot. ADAMTS8 overexpression was induced by transfection with pEZ-M90-ADAMTS8 plasmid using lipofectamine 2000. To generate ADAMTS8 stable knockdown cells, MDA-MB-231 cells were transfected with psi-H1-ADAMTS8siRNA plasmids. Cell counting kit-8 (CCK-8) assay, wound-healing assay, transwell assay and flow cytometry assay were employed to analyze the effects of ADAMTS8 on the proliferation, migration, invasion and apoptosis of BC cells. Chemosensitivity also was assessed using CCK-8 assay. The expressions of β-catenin, MMP-7 and c-Myc were measured by Western blot. Results Our results showed that ADAMTS8 expression was significantly lower in BC tissues than that in adjacent non-tumor tissues. Overexpression of ADAMTS8 in MDA-MB-453 cells could inhibit the cell proliferation, migration and invasion and promote apoptosis. ADAMTS8 knockdown displayed the reverse effect in MDA-MB-231 cells. Consistently, in vivo data showed that ADAMTS8 overexpression led to a reduction in tumor growth. In addition, chemosensitivity testing in MDA-MB-453 cells transfected with pEZ-M90-ADAMTS8 plasmid indicated that cisplatin inhibited cell growth dramatically. Furthermore, attenuated β-catenin, MMP-7 and c-Myc level was detected after ADAMTS8 overexpression. Conclusion These results indicate that increased ADAMTS8 expression could modify the progression of BC by inhibiting cell proliferation and invasion while promoting the apoptosis of BC cells. Thus, ADAMTS8 represents a potential therapeutic target for BC therapy.

BACKGROUND The aim of this study was to investigate the correlations between ADAMTSs expression and breast invasive ductal carcinoma (IDC), and to offer a theoretical basis for novel treatment methods for IDC patients. MATERIAL AND METHODS Non-proliferative catheter of breast fibroadenoma (FA) and IDC were used as the normal control and experimental group, respectively. Immunohistochemical (IHC) staining and Western blot (WB) analysis was used to assess protein expression levels of ADAMTS8, ADAMTS18, and ADAMTS20 in both FA and IDC tissues. The results of IHC, the relationship between the protein expression and the tumor molecular classification, and clinical pathological parameters were all evaluated. RESULTS IHC and WB results showed that the expression of ADAMTS8/18 in IDC samples was higher than in FA samples, while the expression of ADAMTS20 in IDC samples was lower than that in FA samples. According to the results of WB, the level of ADAMTS8 was higher in the HER2+ group than in the HER2- group and FA group. The expression of ADAMTS18 in the HR+ (including ER+ and PR+) group was significantly higher than in the HR- group and FA group. The expression of ADAMTS18 protein was also higher in the Ki67+ group than in the Ki67- group. ADAMTS20 was higher in HER2+ IDC compared with the basal subtype of IDC. CONCLUSIONS ADAMTS8/18/20 levels were not significantly correlated to the molecular subtype of IDC. ADAMTS18/20 was significantly associated with histological grade of IDC. ADAMTS8 may predict poor prognosis results of IDC patients.

// Dan Zhou 1, 2 , Weiwei Tang 1 , Xinli Liu 1 , Han-Xiang An 1 and Yun Zhang 1, 2 1 Department of Medical Oncology, The First Affiliated Hospital of Xiamen University, Xiamen, Fujian, China 2 Xiamen Institute of Rare Earth Materials, Chinese Academy of Sciences and Key Laboratory of Design and Assembly of Functional Nanostructures, Fujian Provincial Key Laboratory of Nanomaterials, Fujian Institute of Research on the Structure of Matter, Chinese Academy of Sciences, Xiamen, Fujian, China Correspondence to: Yun Zhang, email: zhangy@fjirsm.ac.cn Han-Xiang An, email: anhanxiang@xmu.edu.cn Keywords: lung cancer, biomarker, cell-free RNA, HJURP, ADAMTS8 Received: November 30, 2016 Accepted: March 08, 2017 Published: March 30, 2017 ABSTRACT Lung cancer (LC) remains associated with significant mortality worldwide. The lack of reliable noninvasive biomarkers and targeted therapies contributes to poor survival rate. Herein, we initially took advantage of the public microarray data from Oncomine database to filter messenger RNAs (mRNAs) as potential biomarkers. Subsequently, clinical validation was applied to identify candidate noninvasive biomarkers in plasma from patients with LC. Through comprehensive analysis of transcriptional expression profiles across 12 studies, top 6 over- and underexpressed mRNAs were generated. Then, a pair of matched plasma samples from LC patient and normal control was detected by RT-PCR, and three genes with positive bands were selected for further validation. Finally, qPCR was conducted to further assess values of the three identified genes. We displayed with high confidence that two cell-free mRNAs (HJURP and ADAMTS8) were expressed at significantly levels compared to normal controls. Receiver-operating characteristic (ROC) curves on the diagnostic efficacy of plasma HJURP and ADAMTS8 mRNAs in LC diagnosis showed that the area under the ROC (AUC) was 0.6960 and 0.6877; sensitivity was 66.0% and 83.7%; specificity was 78.6% and 71.4%, respectively. Combined ROC analyses using these two biomarkers revealed an elevated AUC of 0.75. Furthermore, the higher HJURP level could be associated with early-stage LC while lower ADAMTS8 level could be correlated with non-small cell lung cancer. Collectively, circulating HJURP and ADAMTS8 mRNAs are promising noninvasive biomarkers for LC diagnosis. Our integrative strategy provides new insights into novel noninvasive biomarker identification for other types of cancer.

The aim of this study was to investigate the serum levels of the A disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS) 5 and 8 in patients diagnosed with endometrial cancer. Our study included 41 patients diagnosed with endometrial cancer. The control group consisted of 41 patients diagnosed with benign endometrial pathology. The serum samples were centrifuged and stored at -80 °C. The serum levels of ADAMTS were significantly higher (

Objective: Alzheimer’s disease (AD) is a progressive neurodegenerative disease that mostly affects the elderly population. Recent studies performed in AD highlight the pathophysiological relevance of disintegrin and metalloproteinase with thrombospondin type 1like motifs (ADAMTS) genes and their products, namely hypoxia inducible factor-1 (HIF-1) and thrombospondin-1 (TSP1). Thus, the aim of this study was to describe and identify the distribution, characteristics, and any changes in the expression and immunoreactivity for HIF-1, TSP-1, and ADAMTS1 and 8 in AD brains. Materials and Methods: Nine patients who were autopsied in the Council of Forensic Medicine, Bursa Morgue Department in 2013, were selected. All patients were sent for autopsy to the Morgue Department within 8 h after death. At the autopsy, tissue samples of the organs were obtained for histopathological examination for determining the cause of death. Among these, two patients were clinically diagnosed with AD. Results: Immunohistochemical staining was performed, and the staining intensity/extensity was evaluated using a semiquantitative scoring system. Median distribution (extensity) scores of the immunohistochemical staining were estimated as 2 for HIF-1, 0.67 for TSP-1, 3.11 for ADAMTS1, and 2.78 for ADAMTS8. Intensity scores were estimated as 1.22 for HIF-1, 0.56 for TSP-1, 3 for ADAMTS1, and 2.11 for ADAMTS8. Conclusion: Our study suggests that ADAMTS1 and ADAMTS8 expressions are not specific for AD. To understand and provide definitive data on all aspects of metalloproteinases, extracellular matrix proteins, and transcriptional factor effects to AD, further studies are needed, where other metalloproteinases and related molecules/enzymes should be studied.

Xuetao Zhao,1 Congrong Yang,2 Jianhua Wu,3 Yuemin Nan4 1Department of Blood Transfusion, The Fourth Hospital of Hebei Medical University, Shijiazhuang, People’s Republic of China; 2Department of Radiation Oncology, The Fourth Hospital of Hebei Medical University, Shijiazhuang, People’s Republic of China; 3Animal Center, The Fourth Hospital of Hebei Medical University, Shijiazhuang, People’s Republic of China; 4Department of Traditional and Western Medical Hepatology, Third Hospital of Hebei Medical University, Shijiazhuang, People’s Republic of China Introduction: Hepatocellular carcinoma (HCC) is one of the most common malignant tumors of the digestive system. A disintegrin and metallopeptidase with thrombospondin motif (ADAMTS) has been identified as a secreted metalloproteinase that participates in the inhibition of tumor cell growth and invasion. The aims of the present study were to investigate the clinical significance of ADAMTS8 in patients with HCC and to determine the effect of ADAMTS8 on HCC cell biological activity in vitro and in vivo.Methods: The tumor tissues and matched adjacent non-tumor tissues were collected from 61 patients with HCC, and ADAMTS8 expression was detected with immunohistochemistry. Flow cytometry and MTT assays were used to assess cell apoptosis and cell viability, respectively, and ERK, p-ERK, Stat3, p-Stat3, Akt, and p-Akt protein expressions were measured by Western blot.Results: The results showed that ADAMTS8 expression was significantly lower in HCC tissues than that in adjacent non-tumor tissues. Moreover, ADAMTS8 expression was inversely associated with clinical stages and metastasis in patients with HCC. Furthermore, we found that transfection with exogenous ADAMTS8 inhibited proliferation and migration and induced apoptosis in HepG2 cells. In the in vivo study, tumor growth of upregulated HepG2 cells in nude mice was significantly slower. Moreover, decreased ERK activity was detected after transfection with ADAMTS8.Conclusion: These results indicate that low ADAMTS8 expression is a predictor of a poor prognosis in patients with HCC and that ADAMTS8 plays an important role in regulating HCC growth, invasion, and apoptosis by modulating the ERK signaling pathway. ADAMTS8 maybe a new target in HCC treatment. Keywords: hepatocellular carcinoma, ADAMT8, apoptosis, invasion and metastasis, signaling pathway

OBJECTIVE The aim is to investigate the expression status of ADAMTS1,8, and 18 proteases in gastric cancer (GC) and lymphatic metastasis. BACKGROUND A disintegrin and metalloprotease with thrombospondin motifs (ADAMTS) is a new protease family, and has important biological functions such as hemostasis, extracellular matrix remodeling and regulation of angiogenesis. METHODS The immunostaining status of ADAMTS1,8, and 18 were investigated in formalin-fixed paraffin-embedded samples of 25 patients who underwent curative resection for GC. RESULTS The immune reactivity scores (IRS) of ADAMTS1, 8, and 18 were significantly higher in the cancerous gastric tissue in comparison to non-cancerous gastric tissue (p < 0.001). In addition, IRS scores of these three ADAMTS proteases were higher in the metastatic lymph nodes compared with healthy lymph nodes (p < 0.001). The expression status of the three ADAMTSs in cancerous gastric tissue was correlated with stage of tumor. Additionally, ADAMTS1 expression and ADAMTS8 expression in cancerous gastric tissue were found to correlate with grade and tumor size, respectively. CONCLUSION This study showed that ADAMTS1, 8, and 18 are highly expressed in GC and its nodal metastases, suggesting important roles of these proteases in carcinogenesis and lymphatic metastasis. The findings from the present study indicate that these proteases may be promising candidates for novel and alternative treatments in GC (Tab. 3, Fig. 3, Ref. 27).

et al., [Objective]: The purpose of this study was to determine the global patterns of aberrant DNA methylation in thyroid cancer. [Research Design and Methods]: We have used DNA methylation arrays to determine, for the first time, the genome-wide promoter methylation status of papillary, follicular, medullary, and anaplastic thyroid tumors. [Results]: We identified 262 and 352 hypermethylated and 13 and 21 hypomethylated genes in differentiated papillary and follicular tumors, respectively. Interestingly, the other tumor types analyzed displayed more hypomethylated genes (280 in anaplastic and 393 in medullary tumors) than aberrantly hypermethylated genes (86 in anaplastic and 131 in medullary tumors). Among the genes indentified, we show that 4 potential tumor suppressor genes (ADAMTS8, HOXB4, ZIC1, and KISS1R) and 4 potential oncogenes (INSL4, DPPA2, TCL1B, and NOTCH4) are frequently regulated by aberrant methylation in primary thyroid tumors. In addition, we show that aberrant promoter hypomethylation- associated overexpression of MAP17 might promote tumor growth in thyroid cancer. [Conclusions]: Thyroid cancer subtypes present differential promoter methylation signatures, and nondifferentiated subtypes are characterized by aberrant promoter hypomethylation rather than hypermethylation. Additional studies are needed to determine the potential clinical interest of the tumor subtype-specific DNA methylation signatures described herein and the role of aberrant promoter hypomethylation in nondifferentiated thyroid tumors. Copyright © 2013 by The Endocrine Society., This work has been financially supported by Fundación ASTURCOR(to S.R.R); the FIS/FEDER (PI11/01728 to A.F.F. and PI11/02795 to E.D.-A.); the ISCIII (CP11/00131 to A.F.F.); the FIS PI11/01359 to M.R.; the Spanish Ministry of Health (PS09/02454 and PI12/01080 to M.F.F.); the Spanish National Research Council (CSIC; 200820I172 to M.F.F.); Instituto Universitario de Oncología del Principado de Asturias (to C.F. and G.F.B.); Ramon Areces Foundation (to M.F.F. and G.F.B.); Fundacion Cientifica de la AECC (to R.G.U.); RD12/0036/0013, BFU-2010-16025, RD12/0036/0030 (to P.S.), RD12/0036/0015 and PI11/00929 (to C.S.), and S2011/BMD-2328 TIRONET (to P.S. and M.R.). Tumor samples were obtained with the support of Red de Biobancos ISCIII (RD09/0076/0059). V.M. is supported by a fellowship of “la Caixa”/Spanish National Cancer Research Center international PhD Program. The IUOPA is supported by the Obra Social Cajastur, Spain.

Background: ADAMTS1 and ADAMTS8 are proteases involved in extracellular matrix proteolysis and antiangiogenesis, but little is known about their expression and function in cerebral ischemia. We investigated the changes in ADAMTS1 and ADAMTS8 in a rat model of permanent middle cerebral artery occlusion (pMCAO). The expressions of glyseraldehyde-3-phosphate dehydrogenase (GAPDH), β-actin, cyclophilin, and RPL13A were examined in order to validate the appropriate housekeeping genes for a long duration after inducing cerebral ischemia. Methods: Male Sprague–Dawley rats were subjected to pMCAO, and ischemic penumbra was collected at 2, 24 h, 3, 7, and 21 days after inducing ischemia, ADAMTS1, ADAMTS8, and the four housekeeping genes were quantified using real-time polymerase chain reaction (PCR). Results: The expression of β-actin increased up to 21 days, and that of GAPDH decreased at 24 h after pMCAO, with no statistically significant changes in RPL13A and cyclophilin being detected. ADAMTS1 mRNA increased at 2 h after pMCAO, peaked at 24 h, and remained at a high level until 21 days. The expression of ADAMTS8 mRNA decreased at 2 and 24 h after pMCAO, followed by a slight increase at 3 days, and then decreased again at 7 days. Conclusion: The results suggest that RPL13A and cyclophilin are two appropriate housekeeping genes for the rat pMCAO model up to 21 days. ADAMTS1 mRNA levels increased, but ADAMTS8 decreased after pMCAO. Our data provide new insight into the mechanism of brain ischemia and self-repair after injury.

Contains fulltext : 50313.pdf (Publisher’s version ) (Closed access) We recently undertook expression profiling of all 19 human ADAMTS metalloproteinases (a disintegrin and metalloproteinase with thrombospondin motifs) in malignant and non-neoplastic breast tissue and showed that 11 of the ADAMTS genes are dysregulated in breast carcinoma. We identified a subgroup of ADAMTSs, based on functional and amino acid sequence similarity (ADAMTS1, 4, 5, 8 and 15), to be the focus of further study in breast carcinoma. Further RNA expression analysis by real-time PCR on a different cohort of 229 patients with breast cancer has identified ADAMTS8 as a predictor of poor overall survival (OS) (hazard ratio (HR) = 2.20, 95% C.I. = 1.29-3.74, p = 0.004) and confirmed ADAMTS15 as a predictor of prolonged relapse-free survival (RFS) (HR = 0.54, 95% C.I. = 0.32-0.89, p = 0.016). We explored the differences in survival of the 4 groups that could be categorized based on the expression levels of ADAMTS8 and ADAMTS15. For both RFS and OS, the group with high ADAMTS8 and low ADAMTS15 expression had a particularly poor prognosis. This group had a 3-fold higher chance of recurrence (HR = 3.03, 95% C.I. = 1.49-6.15, p = 0.001) and a greater than 5-fold higher chance of death (HR = 5.40, 95% C.I. = 2.16-13.5, p < 0.001) than the most favorable prognostic group. This prediction of poor prognosis by ADAMTS8 and ADAMTS15 expression was found to be independent of other classical clinicopathological factors. Results observed in FVB-PyMT mice, a robust transgenic model of highly metastatic breast carcinoma, fitted the expectation that relatively high expression levels of ADAMTS8 together with low expression levels of ADAMTS15 seen in human breast carcinoma are associated with a poor clinical outcome. In summary, ADAMTS8 and ADAMTS15 have emerged as novel predictors of survival in patients with breast carcinoma.

A disintegrins and metalloproteinases with thrombospondin motifs (ADAMTS) family members have been reported dysregulated in various cancers. Through refining a loss of heterozygosity locus at 11q25 by array-CGH, we identified ADAMTS8 as a novel candidate tumor suppressor gene. Although ADAMTS8 downregulation has been reported in several tumors, its biologic function and underlying mechanism remain largely unknown. Here, we found that ADAMTS8 is broadly expressed in normal tissues but frequently downregulated or silenced by promoter methylation in common carcinoma cell lines, including nasopharyngeal, esophageal squamous cell, gastric, and colorectal carcinomas. Pharmacologic or genetic demethylation restored ADAMTS8 expression, indicating that promoter methylation mediates its silencing. Aberrant methylation of ADAMTS8 was also detected in several types of primary tumors but rarely in normal tissues. Further functional studies showed that restoring ADAMTS8 expression suppressed tumor cell clonogenicity through inducing apoptosis. ADAMTS8 as a secreted protease inhibited epidermal growth factor receptor (EGFR) signaling along with decreased levels of phosphorylated MEK and ERK. We further found that ADAMTS8 disrupted actin stress fiber organization and inhibited tumor cell motility. Thus, our data demonstrate that ADAMTS8 metalloprotease acts as a functional tumor suppressor through antagonizing EGFR–MEK–ERK signaling, in addition to its previously reported anti-angiogenesis function, and is frequently methylated in common tumors. Implications: This study uncovers the tumor suppressive function of ADAMTS8, one of the ADAMTS family members, and its frequent methylation in certain tumors could be developed as a potential biomarker. Mol Cancer Res; 12(2); 228–38. ©2013 AACR.

Introduction ADAMTS4 and ADAMTS8 are proteases involved in ECM proteolysis and antiangiogenesis, but little is known about their expression and function in myocardial infarction (MI). We examined ADAMTS4 and ADAMTS8 expression in a rat MI model by quantitative real-time polymerase chain reaction (qPCR) and enzyme linked immunosorbent assay (ELISA). The expressions of glyseraldehyde-3-phosphate dehydrogenase (GAPDH), beta-actin (ACTB), acidic ribosomal phosphoprotein P0 (ARBP), and ribosomal protein L13A (RPL13A) were examined in order to validate the appropriate housekeeping genes after MI. Methods Male Wistar rats were subjected to MI, and infarcted myocardial tissue was collected at 3, 6, 12, 24 h, 3, 7, 14 and 21 days after MI. ADAMTS4, ADAMTS8, and the four housekeeping genes were quantified using qPCR and the expression stability of the four housekeeping genes was investigated using GeNorm software. The protein levels of ADAMTS4 were detected using ELISA kits. Results The M values of GAPDH, ACTB, ARBP and RPL13A were 0.721, 1.2, 0.812 and 0.812 respectively. GAPDH and ARBP were ranked the most stable genes. ADAMTS4 mRNA increased at 3 h after MI, peaked at 6 h, then decreased rapidly. ADAMTS8 mRNA increased at 6 h, peaked at 24 h, remained high at 3 d, then decreased gradually. The protein levels of ADAMTS4 were significantly increased at 6 h, 12 h, 24 h and 3 d after MI. Conclusion The results suggest that GAPDH and ARBP are two appropriate housekeeping genes for the rat MI model. Both ADAMTS4 and ADAMTS8 mRNA levels and ADAMTS4 protein level increased, but they exhibited different expression profiles.

Purpose: Head and neck squamous cell carcinomas (HNSCC) are characterized by high morbidity and mortality, largely due to the high invasive and metastatic potential of these tumors, high recurrence rates, and low treatment responses. Proteinases have been implicated in several aspects of tumor growth and metastasis in a broad range of tumors including HNSCC. Experimental Design: Comprehensive expression profiling of proteinases [matrix metalloproteinases (MMPs), A disintegrin and metalloproteinase (ADAMs), and ADAMs with thrombospondin motif (ADAMTSs)] and their inhibitors [tissue inhibitor of metalloproteinases (TIMPs)] was done using quantitative real-time reverse transcription-PCR analysis of a large cohort of tissue samples representing the tumor (n = 83), the invasive margin (n = 41), and the adjacent tissue (n = 41) from 83 HNSCC patients, along with normal tissue controls (n = 13), as well as cell lines established from tumors of 34 HNSCC patients. Results: The results show specifically elevated gene expression of several proteinases, including MMP1, MMP3, MMP10, and MMP13 within tumor tissue and peritumoral adjacent tissue. In addition, the results identify several novel HNSCC-associated proteinases, including ADAM8, ADAM9, ADAM17, ADAM28, ADAMTS1, ADAMTS8, and ADAMTS15. There were also significant differences in proteinase expression based on clinical parameters, i.e., tumor location, grade, and local invasion. MMP13 expression was significantly higher in large (>4 cm) locally invasive tumors (P < 0.05). MMP9 expression was significantly decreased in tumors with regional metastasis, whereas increased expression of ADAM8 was noted in the metastatic tumors (P < 0.001 for both). Conclusions: These findings suggest the HNSCC degradome as a valuable source of diagnostic, predictive, and prognostic molecular markers for these malignant tumors. Clin Cancer Res; 16(7); 2022–35. ©2010 AACR.

Background A disintegrin and metalloprotease 33 (ADAM33) polymorphism is strongly associated with asthma and bronchial hyperresponsiveness. Although considered to be a mesenchymal cell–specific gene, recent reports have suggested epithelial expression of ADAM33 in patients with severe asthma. Objectives Because dysregulated expression of ADAM33 can contribute to disease pathogenesis, we characterized the mechanism or mechanisms that control its transcription and investigated ADAM33 expression in bronchial biopsy specimens and brushings from healthy and asthmatic subjects. Methods The ADAM33 promoter and CpG island methylation were analyzed by using bioinformatics, luciferase reporters, and bisulfite sequencing of genomic DNA. Epithelial-mesenchymal transition was induced by using TGF-β1. ADAM33 mRNA was scrutinized in bronchial biopsy specimens and brushings by using reverse transcriptase–quantitative polymerase chain reaction, melt-curve analysis, and direct sequencing. Results The predicted ADAM33 promoter (−550 to +87) had promoter transcriptional activity. Bisulfite sequencing showed that the predicted promoter CpG island (−362 to +80) was hypermethylated in epithelial cells but hypomethylated in ADAM33-expressing fibroblasts. Treatment of epithelial cells with 5-aza-deoxycytidine caused demethylation of the CpG island and induced ADAM33 expression. In contrast, phenotypic transformation of epithelial cells through a TGF-β–induced epithelial-mesenchymal transition was insufficient to induce ADAM33 expression. ADAM33 mRNA was confirmed in bronchial biopsy specimens, but no validated signal was detected in bronchial brushings from healthy or asthmatic subjects. Conclusion The ADAM33 gene contains a regulatory CpG island within its promoter, the methylation status of which tightly controls its expression in a cell type–specific manner. ADAM33 repression is a stable feature of airway epithelial cells, irrespective of disease.

Background We hypothesized that cleavage of the extracellular matrix (ECM) proteoglycans versican and aggrecan by ADAMTS (a disintegrin and metalloprotease with thrombospondin motifs) proteases, which contributes to stress-induced ECM-reorganization in atherogenesis and osteoarthritis, also play a role in heart failure development. Objectives The primary objective was to identify alterations in expression of ADAMTS versicanases and aggrecanases during development of heart failure, while evaluation of the effects of in vivo modulation of relevant changes in ADAMTS activity constituted the secondary objective. Methods Myocardial levels of versican, aggrecan, and their ADAMTS cleaving proteases were examined in Wistar rats six weeks after aortic banding (AB), and versican and selected ADAMTS versicanases were further analyzed in neonatal cardiomyocytes (NCM) and cardiac fibroblasts (NFB) after stimulation by inflammatory mediators. Based on the initial findings, ADAMTS4 was selected the most promising therapeutic target. Thus, rats with AB were treated with pentosan polysulfate (PPS), a polysaccharide with known ADAMTS4-inhibitory properties, and effects on versican fragmentation, left ventricular function and geometry were evaluated. Results We discovered that myocardial mRNA and protein levels of ADAMTS1 and -4, and mRNA levels of versican, aggrecan, and ADAMTS8 increased after AB, and TNF-α and IL-1β synergistically increased mRNA of versican and ADAMTS4 in NCM and NFB and secretion of ADAMTS4 from NCM. Furthermore, PPS-treatment improved systolic function, demonstrated by an improved fractional shortening (vehicle 48±3% versus PPS 60±1%, p

We have studied two related proteins that contain a repeated amino acid motif homologous to the anti-angiogenic type 1 repeats of thrombospondin-1 (TSP1). Complete sequence analysis revealed no other similarities with TSP1, but identified unique signal sequences, as well as metalloprotease and disintegrin-like domains in the NH(2) termini. We named these proteins METH-1 and METH-2 due to the novel combination of metalloprotease and thrombospondin domains. Overall amino acid sequence identity between METH-1 and METH-2 is 51. 7%, yet transcript distribution revealed non-overlapping patterns of expression in tissues and cultured cell lines. To characterize these proteins functionally, we isolated full-length cDNAs, produced recombinant protein, and generated antisera to the recombinant proteins. Both METH-1 and METH-2 represent single copy genes, which encode secreted and proteolytically processed proteins. METH proteins suppressed fibroblast growth factor-2-induced vascularization in the cornea pocket assay and inhibited vascular endothelial growth factor-induced angiogenesis in the chorioallantoic membrane assay. Suppression of vessel growth in both assays was considerably greater than that mediated by either thrombospondin-1 or endostatin on a molar basis. Consistent with an endothelial specific response, METH-1 and METH-2 were shown to inhibit endothelial cell proliferation, but not fibroblast or smooth muscle growth. We propose that METH-1 and METH-2 represent a new family of proteins with metalloprotease, disintegrin, and thrombospondin domains. The distinct distribution of each gene product suggests that each has evolved distinct regulatory mechanisms that potentially allow for fine control of activity during distinct physiological and pathological states.

Background and aims: Loss of chromosome 13q regions is a frequent event in both esophageal squamous cell carcinoma (ESCC) and nasopharyngeal carcinoma (NPC). By microcell-mediated chromosome transfer, chromosome 13 was demonstrated to play a significant tumor suppressive role in both ESCC and NPC in our laboratory. THSD1 is a novel candidate tumor suppressor gene (TSG) previously identified by microarray differential gene expression profiling. It encodes a transmembrane molecule containing a thrombospondin type 1 repeat (TSR), which may be involved in cell adhesion and angiogenesis. Proteins possessing the TSR, such as thrombospondin-1 (Tsp1), ADAM metallopeptidase with thrombospondin type 1 motif, 1 (ADAMTS1) and ADAMTS8, regulate angiogenesis. We previously showed that the mechanisms for THSD1 silencing in ESCC involved loss of heterozygosity and promoter hypermethylation. Our preliminary data indicated transfection of wild type THSD1 into an ESCC cell line, SLMT-1, resulted in significant reduction of colony-forming ability, hence providing functional evidence for its growth suppressive activity. The current study aims to examine if THSD1 plays a significant tumor suppressive role in the nude mouse tumorigenicity assay and to elucidate the mechanism(s) involved in tumor suppression. Methods: Transfection by lipofection of both variants of THSD1 into the ESCC SLMT-1 and NPC HONE1-2 cell lines was used to study the effect of THSD1 in tumor suppression and anti-angiogenesis. Results: Quantitative RT-PCR detected down-regulation of THSD1 expression in 36.4% (16/44) primary ESCC and 54.5% (12/22) NPC tissues. Nude mouse tumorigenicity assays showed tumor suppression in THSD1 over-expressing clones in both ESCC and NPC. Flow cytometry analysis showed over-expression of THSD1 resulted in endoreplication. Tube formation assay with HUVEC is underway to examine the angiogenic role of THSD1. Conclusions: THSD1 is a promising candidate TSG in both ESCC and NPC. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3061.