The Increased Burden of Rare Variants in Four Matrix Metalloproteinase-Related Genes in Childhood Glaucoma Suggests a Complex Genetic Inheritance of the Disease.

Childhood glaucoma encompasses congenital and juvenile primary glaucoma, which are heterogeneous, uncommon, and irreversible optic neuropathies leading to visual impairment with a poorly understood genetic basis. Our goal was to identify gene variants associated with these glaucoma types by assessing the mutational burden in 76 matrix metalloproteinase-related genes. We studied 101 childhood glaucoma patients with no identified monogenic alterations using next-generation sequencing. Gene expression was assessed through immunohistochemistry. Functional analysis of selected gene variants was conducted in cultured cells and in zebrafish. Patients presented a higher proportion of rare variants in four metalloproteinase-related genes, including CPAMD8 and ADAMTSL4, compared to controls. ADAMTSL4 protein expression was observed in the anterior segment of both the adult human and zebrafish larvae's eye, including tissues associated with glaucoma. In HEK-293T cells, expression of four ADAMTSL4 variants identified in this study showed that two variants (p.Arg774Trp and p.Arg98Trp) accumulated intracellularly, inducing endoplasmic reticulum stress. Additionally, overexpressing these ADAMTSL4 variants in zebrafish embryos confirmed partial loss-of-function effects for p.Ser719Leu and p.Arg1083His. Double heterozygous functional suppression of adamtsl4 and cpamd8 zebrafish orthologs resulted in reduced volume of both the anterior eye chamber and lens within the chamber, supporting a genetic interaction between these genes. Our findings suggest that accumulation of partial functional defects in matrix metalloproteinase-related genes may contribute to increased susceptibility to early-onset glaucoma and provide further evidence supporting the notion of a complex genetic inheritance pattern underlying the disease.

ADAMTSL2 is a potential prognostic biomarker and immunotherapeutic target for colorectal cancer: Bioinformatic analysis and experimental verification.

The ADAMTS Like 2 (ADAMTSL2) mutation has been identified to be associated with different human genetic diseases. The role of ADAMTSL2 is unclear in colorectal cancer (CRC). The study investigated the expression of ADAMTSL2 in both pan cancer and CRC, using data from The Cancer Genome Atlas (TCGA) database to assess its diagnostic value. The study examined the correlation between ADAMTSL2 expression levels and clinical characteristics, as well as prognosis in CRC. The study explored potential regulatory networks involving ADAMTSL2, including its association with immune infiltration, immune checkpoint genes, tumor mutational burden (TMB) / microsatellite instability (MSI), tumor stemness index (mRNAsi), and drug sensitivity in CRC. ADAMTSL2 expression was validated using GSE71187 and quantitative real-time PCR (qRT-PCR). ADAMTSL2 was aberrantly expressed in pan cancer and CRC. An increased level of ADAMTSL2 expression in patients with CRC was significantly associated with the pathologic N stage (p < 0.001), pathologic stage (p < 0.001), age (p < 0.001), histological type (p < 0.001), and neoplasm type (p = 0.001). The high expression of ADAMTSL2 in patients with CRC was found to be significantly associated with a poorer overall survival (OS) (HR: 1.67; 95% CI: 1.18-2.38; p = 0.004), progression-free survival (PFS) (HR: 1.55; 95% CI: 1.14-2.11; p = 0.005) and disease-specific survival (DSS) (HR: 1.83; 95% CI: 1.16-2.89; p = 0.010). The expression of ADAMTSL2 in patients with CRC (p = 0.009) was identified as an independent prognostic determinant. ADAMTSL2 was associated with extracellular matrix receptor (ECM-receptor) interaction, transforming growth factor ß (TGF-ß) signaling pathway, and more. ADAMTSL2 expression was correlated with immune infiltration, immune checkpoint genes, TMB / MSI and mRNAsi in CRC. ADAMTSL2 expression was significantly and negatively correlated with 1-BET-762, Trametinib, and WZ3105 in CRC. ADAMTSL2 was significantly upregulated in CRC cell lines. The high expression of ADAMTSL2 is significantly correlated with lower OS and immune infiltration of CRC. ADAMTSL2 may be a potential prognostic biomarker and immunotherapeutic target for CRC patients.

Multi-Omics Pan-Cancer Analysis of Procollagen N-Propeptidase Gene Family of ADAMTS as Novel Biomarkers to Associate with Prognosis, Tumor Immune Microenvironment, Signaling Pathways, and Drug Sensitivities.

BACKGROUND: The extracellular matrix (ECM) modeling induced by the metalloproteinases is a vital characteristic for tumor progression. Previous studies mainly focus on the functions of two subgroups of metalloproteinases: matrix metalloproteinases (MMPs) and a disintegrin and metalloproteases (ADAMs) in tumors. The roles of another important group: the ADAMs with thrombospondin motifs (ADAMTS) remain unclear. This study aimed to perform a pan-cancer analysis of procollagen N-propeptidase subgroup of ADAMTS (PNPSA). METHODS: We systematically analyzed expression landscape, genomic variations, prognostic value, and cell expression clusters of PNPSA in pan-cancer based on the multiple integrated open databases. Besides, we also analyzed the impacts of expressions and genomic variations of PNPSA members on tumor immune microenvironment (TIME) and immune-related molecules in pan-cancer based on the immune-related open databases. The Gene Set Variation Analysis (GSVA) was performed to evaluate the associations of the whole PNPSA with prognosis, tumor indicators, TIME, and drug sensitivities. Meanwhile, the Kyoto Encyclopedia of Genes and Genomes (KEGG) was performed to reveal related signaling pathways. Finally, immunohistochemical staining was used to validate the differential analysis results. RESULTS: We found a dual prognostic role of PNPSA members in pan-cancer and they were significantly correlated with TIME and immune-related molecules. Interestingly, the copy number variations (CNVs) of all PNPSA members were revealed to be negatively correlated with NK cell infiltration in most cancers. Single-cell sequencing analysis reveals expressions of PNPSA gene family members on some specific tumor and immune cells in addition to the fibroblasts. The GSVA score was found to have some predictive value for survival status in Brain Lower Grade Glioma (LGG), Mesothelioma (MESO), and Uveal Melanoma (UVM) and to be significantly correlated with tumorigenesis-related pathways such as PI3K-Akt, AGE-RAGE, etc. The GSVA score also shows some predictive value for chemotherapy and immunotherapy efficacy in some tumors. CONCLUSIONS: PNPSA was correlated with tumor development and might be potential tumor biomarker and therapeutic target.

Lung Inflammatory Phenotype in Mice Deficient in Fibulin-2 and ADAMTS-12.

Interaction between extracellular matrix (ECM) components plays an important role in the regulation of cellular behavior and hence in tissue function. Consequently, characterization of new interactions within ECM opens the possibility of studying not only the functional but also the pathological consequences derived from those interactions. We have previously described the interaction between fibulin2 and ADAMTS-12 in vitro and the effects of that interaction using cellular models of cancer. Now, we generate a mouse deficient in both ECM components and evaluate functional consequences of their absence using different cancer and inflammation murine models. The main findings indicate that mice deficient in both fibulin2 and ADAMTS12 markedly increase the development of lung tumors following intraperitoneal urethane injections. Moreover, inflammatory phenotype is exacerbated in the lung after LPS treatment as can be inferred from the accumulation of active immune cells in lung parenchyma. Overall, our results suggest that protective effects in cancer or inflammation shown by fibulin2 and ADAMTS12 as interactive partners in vitro are also shown in a more realistic in vivo context.

HSFAS mediates fibroblast proliferation, migration, trans-differentiation and apoptosis in hypertrophic scars via interacting with ADAMTS8.

Hypertrophic scar (HS) is one of the most common sequelae of patients, especially after burns and trauma. The roles of regulatory long noncoding RNAs (lncRNAs) in mediating HS remain underexplored. Human hypertrophic scar-derived fibroblasts (HSFBs) have been shown to exert more potent promoting effects on extracellular matrix (ECM) accumulation than normal skin-derived fibroblasts (NSFBs) and are associated with enhanced HS formation. The purpose of this study is to search for lncRNAs enriched in HSFBs and investigate their roles and mechanisms. LncRNA MSTRG.59347.16 is one of the most highly expressed lncRNAs in HS detected by lncRNA-seq and qRT-PCR and named as hypertrophic scar fibroblast-associated lncRNA (HSFAS). HSFAS overexpression significantly induces fibroblast proliferation, migration, and myofibroblast trans-differentiation and inhibits apoptosis in HSFBs, while knockdown of HSFAS results in augmented apoptosis and attenuated proliferation, migration, and myofibroblast trans-differentiation of HSFBs. Mechanistically, HSFAS suppresses the expression of A disintegrin and metalloproteinase with thrombospondin motifs 8 (ADAMTS8). ADAMTS8 knockdown rescues downregulated HSFAS-mediated fibroblast proliferation, migration, myofibroblast trans-differentiation and apoptosis. Thus, our findings uncover a previously unknown lncRNA-dependent regulatory pathway for fibroblast function. Targeted intervention in the HSFAS-ADAMTS8 pathway is a potential therapy for HS.

Extracellular Matrix Disorganization Caused by ADAMTS16 Deficiency Leads to Bicuspid Aortic Valve With Raphe Formation.

BACKGROUND: A better understanding of the molecular mechanism of aortic valve development and bicuspid aortic valve (BAV) formation would significantly improve and optimize the therapeutic strategy for BAV treatment. Over the past decade, the genes involved in aortic valve development and BAV formation have been increasingly recognized. On the other hand, ADAMTS (a disintegrin and metalloproteinase with thrombospondin motifs) gene family members have been reported to be able to modulate cardiovascular development and diseases. The present study aimed to further investigate the roles of ADAMTS family members in aortic valve development and BAV formation. METHODS: Morpholino-based ADAMTS family gene-targeted screening for zebrafish heart outflow tract phenotypes combined with DNA sequencing in a 304 cohort BAV patient registry study was initially carried out to identify potentially related genes. Both ADAMTS gene-specific fluorescence in situ hybridization assay and genetic tracing experiments were performed to evaluate the expression pattern in the aortic valve. Accordingly, related genetic mouse models (both knockout and knockin) were generated using the CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/clustered regularly interspaced short palindromic repeat-associated 9) method to further study the roles of ADAMTS family genes. The lineage-tracing technique was used again to evaluate how the cellular activity of specific progenitor cells was regulated by ADAMTS genes. Bulk RNA sequencing was used to investigate the signaling pathways involved. Inducible pluripotent stem cells derived from both BAV patients and genetic mouse tissue were used to study the molecular mechanism of ADAMTS. Immunohistochemistry was performed to examine the phenotype of cardiac valve anomalies, especially in the extracellular matrix components. RESULTS: ADAMTS genes targeting and phenotype screening in zebrafish and targeted DNA sequencing on a cohort of patients with BAV identified ADAMTS16 (a disintegrin and metalloproteinase with thrombospondin motifs 16) as a BAV-causing gene and found the ADAMTS16 p. H357Q variant in an inherited BAV family. Both in situ hybridization and genetic tracing studies described a unique spatiotemporal pattern of ADAMTS16 expression during aortic valve development. Adamts16+/- and Adamts16+/H355Q mouse models both exhibited a right coronary cusp-noncoronary cusp fusion-type BAV phenotype, with progressive aortic valve thickening associated with raphe formation (fusion of the commissure). Further, ADAMTS16 deficiency in Tie2 lineage cells recapitulated the BAV phenotype. This was confirmed in lineage-tracing mouse models in which Adamts16 deficiency affected endothelial and second heart field cells, not the neural crest cells. Accordingly, the changes were mainly detected in the noncoronary and right coronary leaflets. Bulk RNA sequencing using inducible pluripotent stem cells-derived endothelial cells and genetic mouse embryonic heart tissue unveiled enhanced FAK (focal adhesion kinase) signaling, which was accompanied by elevated fibronectin levels. Both in vitro inducible pluripotent stem cells-derived endothelial cells culture and ex vivo embryonic outflow tract explant studies validated the altered FAK signaling. CONCLUSIONS: Our present study identified a novel BAV-causing ADAMTS16 p. H357Q variant. ADAMTS16 deficiency led to BAV formation.

Opposing roles for ADAMTS2 and ADAMTS14 in myofibroblast differentiation and function.

Crosstalk between cancer and stellate cells is pivotal in pancreatic cancer, resulting in differentiation of stellate cells into myofibroblasts that drives tumour progression. To assess cooperative mechanisms in a 3D context, we generated chimeric spheroids using human and mouse cancer and stellate cells. Species-specific deconvolution of bulk-RNA sequencing data revealed cell type-specific transcriptomes underpinning invasion. This dataset highlighted stellate-specific expression of transcripts encoding the collagen-processing enzymes ADAMTS2 and ADAMTS14. Strikingly, loss of ADAMTS2 reduced, while loss of ADAMTS14 promoted, myofibroblast differentiation and invasion independently of their primary role in collagen-processing. Functional and proteomic analysis demonstrated that these two enzymes regulate myofibroblast differentiation through opposing roles in the regulation of transforming growth factor ß availability, acting on the protease-specific substrates, Serpin E2 and fibulin 2, for ADAMTS2 and ADAMTS14, respectively. Showcasing a broader complexity for these enzymes, we uncovered a novel regulatory axis governing malignant behaviour of the pancreatic cancer stroma. © 2023 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Expression and Purification of Recombinant ADAMTS8.

ADAMTS8 (A Disintegrin-like and Metalloproteinase with Thrombospondin motifs 8) is a secreted zinc-dependent metalloproteinase whose expression is downregulated in a variety of solid tumors. Xenografts expressing high levels of ADAMTS8 have a poor capacity to invade and migrate in nude mice. While this data highlights a beneficial, anti-cancerogenic role of ADAMTS8, the mechanism behind this activity is still not fully elucidated. So far, the only reported substrate for ADAMTS8 is osteopontin (OPN), an extracellular matrix protein widely implicated in multiple steps of cancer progression, albeit, similar to other ADAMTS family members, it is very likely that ADAMTS8 cleaves a variety of substrates. The availability of purified ADAMTS8 may enlighten the biological role of this metalloproteinase.Here we describe methods for expression and purification of recombinant ADAMTS8 in HEK293T cells as well as a convenient assay to test ADAMTS8 proteolytic activity using OPN as a substrate.

ADAMTS12 is a stromal modulator in chronic liver disease.

Adamalysins, a family of metalloproteinases containing a disintegrin and metalloproteinases (ADAMs) and ADAM with thrombospondin motifs (ADAMTSs), belong to the matrisome and play important roles in various biological and pathological processes, such as development, immunity and cancer. Using a liver cancer dataset from the International Cancer Genome Consortium, we developed an extensive in silico screening that identified a cluster of adamalysins co-expressed in livers from patients with hepatocellular carcinoma (HCC). Within this cluster, ADAMTS12 expression was highly associated with recurrence risk and poorly differentiated HCC signatures. We showed that ADAMTS12 was expressed in the stromal cells of the tumor and adjacent fibrotic tissues of HCC patients, and more specifically in activated stellate cells. Using a mouse model of carbon tetrachloride-induced liver injury, we showed that Adamts12 was strongly and transiently expressed after a 24 h acute treatment, and that fibrosis was exacerbated in Adamts12-null mice submitted to carbon tetrachloride-induced chronic liver injury. Using the HSC-derived LX-2 cell line, we showed that silencing of ADAMTS12 resulted in profound changes of the gene expression program. In particular, genes previously reported to be induced upon HSC activation, such as PAI-1, were mostly down-regulated following ADAMTS12 knock-down. The phenotype of these cells was changed to a less differentiated state, showing an altered actin network and decreased nuclear spreading. These phenotypic changes, together with the down-regulation of PAI-1, were offset by TGF-ß treatment. The present study thus identifies ADAMTS12 as a modulator of HSC differentiation, and a new player in chronic liver disease.

ADAMTS12 promotes oxaliplatin chemoresistance and angiogenesis in gastric cancer through VEGF upregulation.

BACKGROUND: While ADAMTS12 (A disintegrin and metalloproteinase with thrombospondin motifs 12) has been established as an important regulator of gastrointestinal tumor development and angiogenic activity, the precise mechanistic functions of ADAMTS12 have yet to be fully clarified in gastric cancer (GC). Accordingly, this study was developed to explore the molecular functions of ADAMTS12 in GC and to examine its utility as a biomarker associated with chemoresistance and prognostic outcomes in this cancer type. METHODS: The ability of ADAMTS12 to modulate the proliferative, migratory, invasive, chemoresistant, and tube formation activity of tumor cells was assessed in vivo and in vitro through gain- and loss-of-function approaches. Correlations between ADAMTS12, CD31, and VEGF expression levels in GC patient tumor tissue samples from individuals that did and did not undergo neoadjuvant chemotherapy (NAC) treatment were analyzed via immunohistochemical staining. RESULTS: These analyses revealed the ability of ADAMTS12 to promote in vivo and in vitro cellular proliferative and angiogenic activity, promoting the activation of ERK and the consequent upregulation of VEGF, thereby inducing angiogenesis and decreasing GC cell oxaliplatin sensitivity. A positive correlation between ADAMTS12 levels and both the expression of VEGF as well as the density of microvessels was observed in GC patient tumor tissues. Moreover, those GC patients exhibiting higher intratumoral ADAMTS12 expression exhibited worse responses to NAC treatment and worse overall survival outcomes. CONCLUSIONS: These findings suggest that ADAMTS12 can modulate signaling via the MAPK/VEGF axis in GC cells to enhance tumor cell resistance to oxaliplatin treatment under hypoxic and normoxic conditions. Elevated ADAMTS12 levels can additionally predict vascular abnormalities, worse survival outcomes, and chemoresistance in patients with GC.

Expression of placental growth factor and a disintegrin and metalloprotease with a thrombospondin type motifs 1-4-8 during the three trimesters of rat pregnancy at the maternal-fetal interface.

The functional roles of the a disintegrin and metalloprotease with a thrombospondin type motifs (ADAMTS) gene family in reproductive physiology, reproductive organs developments and adult reproductive health are still under investigation. The expression of the anti-angiogenic proteases ADAMTS-1, ADAMTS-4 and ADAMTS-8 in placental angiogenesis at various stages of pregnancy also remains unclear. The purpose of this study was therefore to determine the localization and expression of the ADAMTS-1, ADAMTS-4 and ADAMTS-8 proteins during the three stages of pregnancy in rats. Maternal-fetal tissue samples were collected on Days 5, 12 and 19 of each trimester, corresponding to the first, second and third trimesters. The expression of placental growth factor (PlGF) and ADAMTS-1, ADAMTS-4 and ADAMTS-8 at the maternal-fetal interface was examined using immunohistochemistry and western blot at three distinct phases of pregnancy. ADAMTS-1, ADAMTS-4 and ADAMTS-8 were detected in all three trimesters of pregnancy. The relative amount of PIGF increased in the first trimester and decreased significantly in the third trimester (p < 0.05). The expression of ADAMTS-1 and ADAMTS-4 was significantly higher in the second (p < 0.05) and third trimesters (p < 0.01) compared to the first trimester. However, no statistically significant change was observed in ADAMTS-8 expression between trimesters. The ADAMTS exhibiting the highest expression during the first trimester was ADAMTS8. These findings indicate that the expression of ADAMTS-1, ADAMTS-4 and ADAMTS-8 in the three different stages of rat pregnancy may be involved in the modulation of decidualization, morphogenesis and angiogenesis. Periodic changes in ADAMTS expression are thought to be regulated by gonadal steroids.

Epigenetic regulatory mechanism of ADAMTS12 expression in osteoarthritis.

BACKGROUND: Osteoarthritis (OA) is a degenerative joint disease with lacking effective prevention targets. A disintegrin and metalloproteinase with thrombospondin motifs 12 (ADAMTS12) is a member of the ADAMTS family and is upregulated in OA pathologic tissues with no fully understood molecular mechanisms. METHODS: The anterior cruciate ligament transection (ACL-T) method was used to establish rat OA models, and interleukin-1 beta (IL-1ß) was administered to induce rat chondrocyte inflammation. Cartilage damage was analyzed via hematoxylin-eosin, Periodic Acid-Schiff, safranin O-fast green, Osteoarthritis Research Society International score, and micro-computed tomography assays. Chondrocyte apoptosis was detected by flow cytometry and TdT dUTP nick-end labeling. Signal transducer and activator of transcription 1 (STAT1), ADAMTS12, and methyltransferase-like 3 (METTL3) levels were detected by immunohistochemistry, quantitative polymerase chain reaction (qPCR), western blot, or immunofluorescence assay. The binding ability was confirmed by chromatin immunoprecipitation-qPCR, electromobility shift assay, dual-luciferase reporter, or RNA immunoprecipitation (RIP) assay. The methylation level of STAT1 was analyzed by MeRIP-qPCR assay. STAT1 stability was investigated by actinomycin D assay. RESULTS: The STAT1 and ADAMTS12 expressions were significantly increased in the human and rat samples of cartilage injury, as well as in IL-1ß-treated rat chondrocytes. STAT1 is bound to the promoter region of ADAMTS12 to activate its transcription. METTL3/ Insulin-like growth factor 2 mRNA-binding protein 2 (IGF2BP2) mediated N6-methyladenosine modification of STAT1 promoted STAT1 mRNA stability, resulting in increased expression. ADAMTS12 expression was reduced and the IL-1ß-induced inflammatory chondrocyte injury was attenuated by silencing METTL3. Additionally, knocking down METTL3 in ACL-T-produced OA rats reduced ADAMTS12 expression in their cartilage tissues, thereby alleviating cartilage damage. CONCLUSION: METTL3/IGF2BP2 axis increases STAT1 stability and expression to promote OA progression by up-regulating ADAMTS12 expression.

ADAMTS12 promotes migration and epithelial-mesenchymal transition and predicts poor prognosis for pancreatic cancer.

BACKGROUND: ADAMTS (a disintegrin and metalloproteinase with thrombospondin-like motifs) family, a group of extracellular multifunctional enzymes, has been proven to play a pivotal role in the tumor. In pancreatic cancer, the role and mechanism of this family remain unclear. The present study aimed to figure out the hub gene of ADAMTSs and explore the exact roles in the prognosis and biological functions in pancreatic ductal adenocarcinoma (PDAC). METHODS: We used several databases to analyze the ADAMTS family and then screen out the hub genes. The expression of ADAMTS12 in 106 pairs of PDAC tumors and adjacent normal tissues was examined by immunohistochemistry, and its correlations with clinical parameters were further analyzed. The impacts of ADAMTS12 on the migration of PDAC cells were predicted by gene set enrichment analysis and confirmed by transwell assays. The potential impacts of ADAMTS12 on the epithelial-mesenchymal transition (EMT) were identified by database analysis and experimental proof of real-time quantitative polymerase chain reaction (qPCR) and Western blotting. RESULTS: Our study found that ADAMTS12 was a crucial gene in PDAC, and it was highly expressed in tumor tissues when compared to that in the adjacent tissues. ADATMS12 had predictive value of a poor prognosis for PDAC. The elevation of ADAMTS12 was parallel to the progression of PDAC. Inhibition of ADAMTS12 suppressed the migration of PDAC cells and interfered with the process of EMT. CONCLUSIONS: ADAMTS12 is a crucial member of ADAMTSs in PDAC and a predictor of poor prognosis. Additionally, based on its impacts on migration and metastasis in PDAC and the relationship with EMT, ADAMTS12 plays a role of an oncogene in PDAC and may be a promising target for treatment.

Downregulation of ADAMTS3 Suppresses Stemness and Tumorigenicity in Glioma Stem Cell.

AIMS: Glioblastoma multiforme (GBM) is the most aggressive type of human brain tumor, with a poor prognosis and a median overall survival of fewer than 15 months. Glioma stem cells (GSCs) have recently been identified as a key player in tumor initiation and therapeutic resistance in GBM. ADAMTS family of metalloproteinases is known to cleave a wide range of extracellular matrix substrates and has been linked to tissue remodeling events in tumor development. Here, we investigate that ADAMTS3 regulates GSC proliferation and self-renewal activities, and tumorigenesis in orthotopic xenograft models. METHODS: ADAMTS3 mRNA expression levels in normal human astrocyte (NHA), glioma, and GSCs cell lines were compared. After knockdown of ADAMTS3, alamarBlue assay, in vitro limiting dilution, and orthotopic xenograft assays were performed. To investigate the tumor-associated roles of ADAMTS3, several statistical assays were conducted using publicly available datasets. RESULTS: ADAMTS3 level was remarkably higher in GSCs than in NHA, glioma cell lines, and their matched differentiated tumor cells. Interestingly, knockdown of ADAMTS3 disrupted GSC's proliferation, self-renewal activity, and tumor formation in vivo. Furthermore, ADAMTS3 could be used as an independent predictor of malignancy progression in GBM. CONCLUSION: We identified ADAMTS3 as a potential therapeutic target for GBM.

Curcumin Inhibits Proliferation of Renal Cell Carcinoma in vitro and in vivo by Regulating miR-148/ADAMTS18 through Suppressing Autophagy.

OBJECTIVE: To explore the effect of curcumin on the proliferation of renal cell carcinoma and analyze its regulation mechanism. METHODS: In RCC cell lines of A498 and 786-O, the effects of curcumin (2.5, 5, 10 µ mo/L) on the proliferation were analyzed by Annexin V+PI staining. Besides, A498 was inoculated into nude mice to establish tumorigenic models, and the model mice were treated with different concentrations of curcumin (100, 200, and 400 mg/kg), once daily for 30 days. Then the tumor diameter was measured, the tumor cells were observed by hematoxylin-eosin staining, and the protein expressions of miR-148 and ADAMTS18 were detected by immunohistochemistry. In vitro, after transfection of miR-148 mimics, miR-148 inhibitor or si-ADAMTS18 in cell lines, the expression of ADAMTS18 was examined by Western blotting and the cell survival rate was analyzed using MTT. Subsequently, Western blot analysis was again used to examine the autophagy phenomenon by measuring the relative expression level of LC3-II/LC3-I; autophagy-associated genes, including those of Beclin-1 and ATG5, were also examined when miR-148 was silenced in both cell lines with curcumin treatment. RESULTS: Curcumin could inhibit the proliferation of RCC in cell lines and nude mice. The expression of miR-148 and ADAMTS18 was upregulated after curcumin treatment both in vitro and in vivo (P<0.05). The cell survival rate was dramatically declined upon miR-148 or ADAMTS18 upregulated. However, si-ADAMTS18 treatment or miR-148 inhibitor reversed these results, that is, both of them promoted the cell survival rate. CONCLUSION: Curcumin can inhibit the proliferation of renal cell carcinoma by regulating the miR-148/ ADAMTS18 axis through the suppression of autophagy in vitro and in vivo. There may exist a positive feedback loop between miR-148 and ADAMTS18 gene in RCC.

The Prognostic Value of ADAMTS8 and Its Role as a Tumor Suppressor in Breast Cancer.

A disintegrin-like and metalloprotease with therombospondin type1 motif 8 (ADAMTS8) plays an important role in many malignancies. However, the clinical and biological significance of ADAMTS8 in breast cancer remain unknown. In this study, the clinical data from 1066 breast cancer patients were analyzed by The Cancer Genome Atlas (TCGA) database, and were analyzed using the correlation between ADAMTS8 expression and the clinicopathological features and prognoses. The CCK-8 assay, clone formation assay, flow cytometry and Transwell assay were used to characterize the effects of ADAMTS8 on proliferation, migration and invasion of breast cancer cells. Gene set enrichment analysis (GSEA) and western blotting were used to identify the potential molecular mechanism on how ADAMTS8 exert its biological function. ADAMTS8 overexpression correlated longer overall survival (OS) and progression-free survival (PFS). ADAMTS8 was considered as an independent prognostic factor for OS. ADAMTS8 overexpression inhibited breast cancer cell proliferation, migration and invasion in vitro, and induced G2/M cell cycle arrest. ADAMTS8 was also involved in cell cycle regulation and was associated with the EGFR/Akt signaling pathway. ADAMTS8 knockdown showed the reverse effect. Together, the results showed that ADAMTS8 functioned as a tumor suppressor gene (TGS) and could be a prognostic biomarker for breast cancer.

The Novel Protein ADAMTS16 Promotes Gastric Carcinogenesis by Targeting IFI27 through the NF-κb Signaling Pathway.

A disintegrin and metalloproteinase with thrombospondin motifs 16 (ADAMTS16) has been reported to be involved in the pathogenesis of solid cancers. However, its role in gastric cancer (GC) is unclear. In this study, the role of ADAMTS16 in gastric cancer was investigated. The effects of ADAMTS16 on cell migration, invasion, and proliferation were investigated by functional experiments in vivo and in vitro. Downstream signal pathways of ADAMTS16 were confirmed by using bioinformatics analysis, co-immunoprecipitation, and immunofluorescence. Meanwhile, bioinformatics analysis, qRT-PCR, western blot, and dual-luciferase reporter gene analysis assays were used to identify ADAMTS16 targets. The expression of ADAMTS16 in GC was analyzed in public datasets. The expression of ADAMTS16 and its correlations with the clinical characteristics of GC were investigated by immunohistochemistry. Ectopic ADAMTS16 expression significantly promoted tumor cell migration, invasion, and growth. Bioinformatics analysis and western blot showed that ADAMTS16 upregulated the IFI27 protein through the NF-κb pathway, which was confirmed by immunofluorescence and western blot. Dual-luciferase reporter gene analysis identified a binding site between P65 and IFI27 that may be directly involved in the transcriptional regulation of IFI27. IFI27 knockdown reversed the promoting effect of ADAMTS16 on cell invasion, migration, and proliferation indicating that ADAMTS16 acts on GC cells by targeting the NF-κb/IFI27 axis. ADAMTS16 was associated with poor prognosis in clinical characteristics. ADAMTS16 promotes cell migration, invasion, and proliferation by targeting IFI27 through the NF-κB pathway and is a potential progressive and survival biomarker of GC.

Differential Expression and Localization of ADAMTS Proteinases in Proliferative Diabetic Retinopathy.

We analyzed the expression of ADAMTS proteinases ADAMTS-1, -2, -4, -5 and -13; their activating enzyme MMP-15; and the degradation products of proteoglycan substrates versican and biglycan in an ocular microenvironment of proliferative diabetic retinopathy (PDR) patients. Vitreous samples from PDR and nondiabetic patients, epiretinal fibrovascular membranes from PDR patients, rat retinas, retinal Müller glial cells and human retinal microvascular endothelial cells (HRMECs) were studied. The levels of ADAMTS proteinases and MMP-15 were increased in the vitreous from PDR patients. Both full-length and cleaved activation/degradation fragments of ADAMTS proteinases were identified. The amounts of versican and biglycan cleavage products were increased in vitreous from PDR patients. ADAMTS proteinases and MMP-15 were localized in endothelial cells, monocytes/macrophages and myofibroblasts in PDR membranes, and ADAMTS-4 was expressed in the highest number of stromal cells. The angiogenic activity of PDR membranes correlated significantly with levels of ADAMTS-1 and -4 cellular expression. ADAMTS proteinases and MMP-15 were expressed in rat retinas. ADAMTS-1 and -5 and MMP-15 levels were increased in diabetic rat retinas. HRMECs and Müller cells constitutively expressed ADAMTS proteinases but not MMP-15. The inhibition of NF-κB significantly attenuated the TNF-α-and-VEGF-induced upregulation of ADAMTS-1 and -4 in a culture medium of HRMECs and Müller cells. In conclusion, ADAMTS proteinases, MMP-15 and versican and biglycan cleavage products were increased in the ocular microenvironment of patients with PDR.

Contribution of MicroRNA-27b-3p to Synovial Fibrotic Responses in Knee Osteoarthritis.

OBJECTIVE: Synovial fibrosis contributes to osteoarthritis (OA) pathology, but the underlying mechanisms remain unknown. We have observed increased microRNA-27b-3p (miR-27b-3p) levels in synovial fluid of patients with late-stage radiographic knee OA. Here, we investigated the contribution of miR-27b-3p to synovial fibrosis in patients with severe knee OA and in a mouse model of knee OA. METHODS: We stained synovium sections obtained from patients with radiographic knee OA scored according to the Kellgren/Lawrence scale and mice that underwent destabilization of the medial meniscus (DMM) for miR-27b-3p using in situ hybridization. We examined the effects of intraarticular injection of miR-27b-3p mimic into naive mouse knee joints and intraarticular injection of a miR-27b-3p inhibitor into mouse knee joints after DMM. We performed transfection with miR-27b-3p mimic and miR-27b-3p inhibitor in human OA fibroblast-like synoviocytes (FLS) using reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR) array, RNA sequencing, RT-qPCR, Western blotting, immunofluorescence, and migration assays. RESULTS: We observed increased miR-27b-3p expression in the synovium from patients with knee OA and in mice with DMM-induced arthritis. Injection of the miR-27b-3p mimic in mouse knee joints induced a synovial fibrosis-like phenotype, increased synovitis scores, and increased COL1A1 and α-smooth muscle actin (α-SMA) expression. In the mouse model of DMM-induced arthritis, injection of the miR-27b-3p inhibitor decreased α-SMA but did not change COL1A1 expression levels or synovitis scores. Transfection with the miR-27b-3p mimic in human OA FLS induced profibrotic responses, including increased migration and expression of key extracellular matrix (ECM) genes, but transfection with the miR-27b-3p inhibitor had the opposite effects. RNA sequencing identified a PPARG/ADAMTS8 signaling axis regulated by miR-27b-3p in OA FLS. Human OA FLS transfected with miR-27b-3p mimic and then treated with the PPARG agonist rosiglitazone or with ADAMTS8 small interfering RNA exhibited altered expression of select ECM genes. CONCLUSION: Our findings demonstrate that miR-27b-3p has a key role in ECM regulation associated with synovial fibrosis during OA.

The Mechanism and Role of ADAMTS Protein Family in Osteoarthritis.

Osteoarthritis (OA) is a principal cause of aches and disability worldwide. It is characterized by the inflammation of the bone leading to degeneration and loss of cartilage function. Factors, including diet, age, and obesity, impact and/or lead to osteoarthritis. In the past few years, OA has received considerable scholarly attention owing to its increasing prevalence, resulting in a cumbersome burden. At present, most of the interventions only relieve short-term symptoms, and some treatments and drugs can aggravate the disease in the long run. There is a pressing need to address the safety problems due to osteoarthritis. A disintegrin-like and metalloprotease domain with thrombospondin type 1 repeats (ADAMTS) metalloproteinase is a kind of secretory zinc endopeptidase, comprising 19 kinds of zinc endopeptidases. ADAMTS has been implicated in several human diseases, including OA. For example, aggrecanases, ADAMTS-4 and ADAMTS-5, participate in the cleavage of aggrecan in the extracellular matrix (ECM); ADAMTS-7 and ADAMTS-12 participate in the fission of Cartilage Oligomeric Matrix Protein (COMP) into COMP lyase, and ADAMTS-2, ADAMTS-3, and ADAMTS-14 promote the formation of collagen fibers. In this article, we principally review the role of ADAMTS metalloproteinases in osteoarthritis. From three different dimensions, we explain how ADAMTS participates in all the following aspects of osteoarthritis: ECM, cartilage degeneration, and synovial inflammation. Thus, ADAMTS may be a potential therapeutic target in osteoarthritis, and this article may render a theoretical basis for the study of new therapeutic methods for osteoarthritis.