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Severe acute respiratory syndrome coronavirus 2 (SARS‑CoV‑2) induced cytokine storm is the major cause of COVID-19 related deaths. Patients have been treated with drugs that work by inhibiting a specific protein partly responsible for the cytokines production. This approach provided very limited success, since there are multiple proteins involved in the complex cell signaling disease mechanisms. We targeted five proteins: Angiotensin II receptor type 1 (AT1R), A disintegrin and metalloprotease 17 (ADAM17), Nuclear Factor‑Kappa B (NF‑κB), Janus kinase 1 (JAK1) and Signal Transducer and Activator of Transcription 3 (STAT3), which are involved in the SARS‑CoV‑2 induced cytokine storm pathway. We developed machine-learning (ML) models for these five proteins, using known active inhibitors. After developing the model for each of these proteins, FDA-approved drugs were screened to find novel therapeutics for COVID‑19. We identified twenty drugs that are active for four proteins with predicted scores greater than 0.8 and eight drugs active for all five proteins with predicted scores over 0.85. Mitomycin C is the most active drug across all five proteins with an average prediction score of 0.886. For further validation of these results, we used the PyRx software to conduct protein-ligand docking experiments and calculated the binding affinity. The docking results support findings by the ML model. This research study predicted that several drugs can target multiple proteins simultaneously in cytokine storm-related pathway. These may be useful drugs to treat patients because these therapies can fight cytokine storm caused by the virus at multiple points of inhibition, leading to synergistically effective treatments.

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Background: Mutations in the receptor tyrosine kinase gene fibroblast growth factor receptor 2 (FGFR2) occur at a high frequency in endometrial cancer (EC) and have been linked to advanced and recurrent disease. However, little is known about how these mutations drive carcinogenesis. Methods: Differential transcriptomic analysis and two‐step quantitative real‐time PCR (qRT‐PCR) assays were applied to identify genes differentially expressed in two cohorts of EC patients carrying mutations in the FGFR2 gene as well as in EC cells harbouring mutations in the FGFR2. Candidate genes and target signalling pathways were investigated by qRT‐PCR assays, immunohistochemistry and bioinformatics analysis. The functional roles of differently regulated genes were analysed using in vitro and in vivo experiments, including 3D‐orthotypic co‐culture systems, cell proliferation and migration protocols, as well as colony and focus formation assays together with murine xenograft tumour models. The molecular mechanisms were examined using CRISPR/Cas9‐based loss‐of‐function and pharmacological approaches as well as luciferase reporter techniques, cell‐based ectodomain shedding assays and bioinformatics analysis. Results: We show that common FGFR2 mutations significantly enhance the sensitivity to FGF7‐mediated activation of a disintegrin and metalloprotease (ADAM)17 and subsequent transactivation of the epidermal growth factor receptor (EGFR). We further show that FGFR2 mutants trigger the activation of ADAM10‐mediated Notch signalling in an ADAM17‐dependent manner, highlighting for the first time an intimate cooperation between EGFR and Notch pathways in EC. Differential transcriptomic analysis in EC cells in a cohort of patients carrying mutations in the FGFR2 gene identified a strong association between FGFR2 mutations and increased expression of members of the Notch pathway and ErbB receptor family. Notably, FGFR2 mutants are not constitutively active but require FGF7 stimulation to reprogram Notch and EGFR pathway components, resulting in ADAM17‐dependent oncogenic growth. Conclusions: These findings highlight a pivotal role of ADAM17 in the pathogenesis of EC and provide a compelling rationale for targeting ADAM17 protease activity in FGFR2‐driven cancers. [ABSTRACT FROM AUTHOR]

Atherosclerosis is a chronic inflammatory disease that poses a huge economic burden due to its extremely poor prognosis. Therefore, it is necessary to explore potential mechanisms to improve the prevention and treatment of atherosclerosis. A disintegrin and metalloprotease 17 (ADAM17) is a cell membrane–bound protein that performs a range of functions through membrane protein shedding and intracellular signaling. ADAM17-mediated inflammation has been identified to be an important contributor to atherosclerosis; however, the specific relationship between its multiple regulatory roles and the pathogenesis of atherosclerosis remains unclear. Here, we reviewed the activation, function, and regulation of ADAM17, described in detail the role of ADAM17-mediated inflammatory damage in atherosclerosis, and discussed several controversial points. We hope that these insights into ADAM17 biology will lead to rational management of atherosclerosis. ADAM17 promotes vascular inflammation in endothelial cells, smooth muscle cells, and macrophages, and regulates the occurrence and development of atherosclerosis. [ABSTRACT FROM AUTHOR]

Background/Aims: High-mobility group box 1 (HMGB1) elicits inflammatory responses through interactions with the receptor for advanced glycation end products (RAGE) and toll-like receptor 4 (TLR4). We investigated how RAGE and TLR4 expressions are regulated after HMGB1 stimulation in cultured human aortic endothelial cells (HAECs). Methods: RAGE and TLR4 expressions were analyzed by Western blot analysis and immunofluorescence staining. A disintegrin and metalloprotease 17 (ADAM17) activity was measured using a fluorogenic substrate. Results: Upon treatment with HMGB1, both RAGE and TLR4 began to decrease in cell lysate and remained decreased up to 24 h. The decrease in cellular RAGE and TLR4 was accompanied by an increase of N-terminal fragment of RAGE and TLR4 in culture supernatant, indicating ectodomain shedding of the receptors. HMGB1 activated p38 mitogen-activated protein kinase (p38 MAPK) and ADAM17, while HMGB1-induced ADAM17 activation was inhibited by SB203580, a p38 MAPK inhibitor. HMGB1-induced ectodomain shedding of RAGE and TLR4 was prevented by siRNA depletion of ADAM17 as well as TAPI-2, an inhibitor of ADAM family, and SB203580. HMGB1 pretreatment abolished p38 MAPK activation in response to 2nd HMGB1 stimulation. In the cells depleted of ADAM17, HMGB1-induced p38 MAPK activation was prolonged. siRNA depletion of RAGE, but not TLR4, suppressed HMGB1-induced p38 MAPK activation. Conclusion: In response to HMGB1 stimulation, HAECs rapidly undergo ectodomain shedding of RAGE and TLR4, and thereby become insensitive to further HMGB1 stimulation. ADAM17, activated through RAGE-p38 MAPK pathway, is implicated in the ectodomain cleavage of the receptors.

Background/Aims: Lipopolysaccharide (LPS)-activated monocytes/macrophages develop endotoxin tolerance in part by reducing cell surface toll-like receptor 4 (TLR4) through cluster of differentiation 14 (CD14)-dependent endocytosis. In case of endothelial cells, CD14 is expressed in low copy numbers as compared with monocytes/macrophages. Thus, we explored how endothelial cells regulate TLR4 expression after LPS stimulation. Methods: Cultured human aortic endothelial cells (HAECs) were treated with LPS. TLR4 expression was analyzed by Western blot analysis and immunofluorescence staining. A disintegrin and metalloprotease 17 (ADAM17) activity was measured using a fluorescent substrate. Results: TLR4 in cell lysate began to decrease within 30 min of LPS treatment with a maximal reduction at 2 h, and it was accompanied by an increase of N-terminal fragment of TLR4 in culture supernatant, indicating ectodomain shedding of the receptor. LPS activated p38 mitogen-activated protein kinase (p38 MAPK) and ADAM17, while LPS-induced ADAM17 activation was inhibited by SB203580, a p38 MAPK inhibitor. LPS-induced ectodomain shedding of TLR4 was attenuated by siRNA depletion of ADAM17 as well as TAPI-2 (an inhibitor of ADAM family) and SB203580. LPS pretreatment resulted in a blunted response of p38 MAPK activation to further LPS stimulation. In the cells depleted of ADAM17, LPS-induced p38 MAPK activation was prolonged and LPS-induced intercellular adhesion molecule-1 expression was potentiated. Conclusion: HAECs respond to LPS by rapid shedding of the ectodomain of TLR4 and thereby reduce the responsiveness to subsequent LPS exposure. ADAM17, downstream of p38 MAPK, is implicated in the ectodomain cleavage of TLR4.

It is known that the renin-angiotensin system plays a major role in the pathophysiology of cardiovascular disease and renal injury. Within the renin-angiotensin system, angiotensin-converting enzyme 2 (ACE2) cleaves ANG II to generate ANG(1–7) peptide, which counteracts the adverse effects of ANG II accumulation. ACE2 can undergo cleavage or shedding to release the catalytically active ectodomain into the circulation by a disintegrin and metalloprotease (ADAM)17, also known as TNF-α-converting enzyme. ADAM17 is involved in many pathological processes such as cancer, inflammatory diseases, neurological diseases, cardiovascular diseases, atherosclerosis, diabetes, and hypertension. Clinical and experimental studies have shown that ADAM17 is involved in chronic kidney disease (CKD) with a proinflammatory and profibrotic role, suggesting that it could be an important mediator of CKD progression. ADAM17 inhibition attenuates fibrosis and inflammation, suggesting that its inhibition may be a possible new valuable therapeutic tool in fibrotic kidney disease treatment. In addition, in renal disease, some experimental studies have demonstrated that ADAM17 is differently expressed in the kidney. Thus, ADAM17 is highly expressed in distal renal tubules and increased in the whole kidney in diabetic models. In this article, we will review the role of ADAM17 under physiological and pathological conditions. We will mainly focus on the importance of ADAM17 in the context of CKD. [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • GKT137831 inhibits hypertensive cardiac remodelling in the rats subjected to AAC. • GKT137831 reduces the expression of proinflammatory cytokines IL-1, IL-6 and TNF-α. • GKT137831 mediated ADAM17-dependent proinflammatory cytokines-induced signalling pathways. Abstract NADPH oxidases (Noxs) 1/4 dual inhibitor GKT137831 prevents hypertensive cardiac remodelling in angiotensin II-infused transgenic mice with cardiomyocyte-specific human Nox4 (c-hNo x 4 Tg); however, further research is still required to determine the beneficial role of GKT137831 in hypertensive cardiac remodelling in other types of hypertensive models because this hypertensive model is insufficient to mimic the complicated pathological mechanisms of hypertension. A disintegrin and metalloprotease 17 (ADAM17) promotes the shedding of tumour necrosis factor α (TNF-α), TNF-α receptor, interleukin 1 receptor-II and interleukin 6 (IL-6) receptor from cells, thereby mediating the signalling pathways induced by corresponding proinflammatory cytokines. This study aimed to determine whether GKT137831 prevents hypertensive cardiac remodelling and its mechanisms of action in the rats with abdominal artery coarctation (AAC). The rats subjected to AAC were orally given GKT137831 for a consecutive period of 28 days. Echocardiography and histological analysis were performed to evaluate cardiac remodelling; and immunohistochemistry and real-time PCR were used to detect the expression of proinflammatory cytokines. GKT137831 significantly suppressed hypertensive cardiac remodelling in AAC-induced hypertensive rats. Concurrently, Nox1/4 dual inhibitor GKT137831 reduced the protein and mRNA levels of proinflammatory cytokines interleukin 1β (IL-1β), IL-6, and TNF-α in the left ventricle of AAC-induced hypertensive rats. Moreover, the treatment with GKT137831 markedly diminished the protein and mRNA levels of ADAM17 in the left ventricle of AAC-induced hypertensive rats. In summary, Nox1/4 dual inhibitor GKT137831 protects against hypertensive cardiac remodelling in AAC-induced hypertensive rats, and the inhibition of ADAM17-dependent proinflammatory cytokines-induced signalling pathways are related to its beneficial effect on hypertensive cardiac remodelling. [ABSTRACT FROM AUTHOR]

Background/Aims: High-mobility group box 1 (HMGB1) elicits inflammatory responses through interactions with the receptor for advanced glycation end products (RAGE) and toll-like receptor 4 (TLR4). We investigated how RAGE and TLR4 expressions are regulated after HMGB1 stimulation in cultured human aortic endothelial cells (HAECs). Methods: RAGE and TLR4 expressions were analyzed by Western blot analysis and immunofluorescence staining. A disintegrin and metalloprotease 17 (ADAM17) activity was measured using a fluorogenic substrate. Results: Upon treatment with HMGB1, both RAGE and TLR4 began to decrease in cell lysate and remained decreased up to 24 h. The decrease in cellular RAGE and TLR4 was accompanied by an increase of N-terminal fragment of RAGE and TLR4 in culture supernatant, indicating ectodomain shedding of the receptors. HMGB1 activated p38 mitogen-activated protein kinase (p38 MAPK) and ADAM17, while HMGB1-induced ADAM17 activation was inhibited by SB203580, a p38 MAPK inhibitor. HMGB1-induced ectodomain shedding of RAGE and TLR4 was prevented by siRNA depletion of ADAM17 as well as TAPI-2, an inhibitor of ADAM family, and SB203580. HMGB1 pretreatment abolished p38 MAPK activation in response to 2nd HMGB1 stimulation. In the cells depleted of ADAM17, HMGB1-induced p38 MAPK activation was prolonged. siRNA depletion of RAGE, but not TLR4, suppressed HMGB1-induced p38 MAPK activation. Conclusion: In response to HMGB1 stimulation, HAECs rapidly undergo ectodomain shedding of RAGE and TLR4, and thereby become insensitive to further HMGB1 stimulation. ADAM17, activated through RAGE-p38 MAPK pathway, is implicated in the ectodomain cleavage of the receptors. [ABSTRACT FROM AUTHOR]

Paeoniflorin (PF), the most abundant active ingredient of traditional Chinese herbal medicine Paeoniae Radix, has been recognized as a potential neuroprotectant due to its remarkable efficacy on mitigating cerebral infarction and preventing the neurodegenerative diseases. However, the precise mechanisms of PF remain incompletely understood. In this study, we first provided evidence for the protective effect of PF on hydrogen peroxide‐induced injury on mouse brain microvascular endothelial bEnd.3 cells, and for transactivation of the epidermal growth factor receptor (EGFR) signal induced by PF, suggesting that EGFR transactivation might be involved in the beneficial role of PF. Next, by detecting the phosphorylation of a disintegrin and metalloprotease 17 (ADAM17) at Thr 735 and performing loss‐of‐function experiments with the ADAM17 inhibitor and ADAM 17‐siRNA, we showed that PF‐induced transactivation of EGFR and downstream ERKs and AKT signaling pathways were dependent on ADAM17. Furthermore, PF‐induced phosphorylation of ADAM17 and the EGFR transactivation were inhibited by the inhibitors of adenosine A1 receptor (A1R) or Src kinase that were applied to cells prior to PF treatment, implying the involvement of A1R, and Src in the activation of ADAM17. Finally, PF reduced the cell surface level of TNF‐receptor 1 (TNFR1) and increased the content of soluble TNFR1 (sTNFR1) in the culture media, indicating that PF might enhance the shedding of sTNFR1. Taken together, we conclude that A1R and Src‐dependent activation of ADAM17 participates in PF‐induced EGFR transactivation and TNFR1 shedding on mouse brain microvascular endothelial cells, which may contributes to the neuroprotective effects of PF. In mouse brain microvascular endothelial cells, paeoniflorin (PF) induces activation of ADAM17, which participates in the PF‐induced epidermal growth factor receptor (EGFR) transactivation and TNFR1 shedding, contributing to the neuroprotective effects of PF. [ABSTRACT FROM AUTHOR]

In our previous study on hepatocellular carcinoma (HCC) susceptibility genes in chronic hepatitis patients, we identified the MHC class I polypeptide‐related sequence A (MICA). Natural killer cells eliminate various cancer cells, including HCC, by suppressing MICA shedding. Therefore, we investigated MICA sheddases and inhibitors for HCC immunotherapy. In this study, HepG2, PLC/PRF/5, and Hep3B were treated with the siRNA of a disintegrin and metalloproteases (ADAMs) and matrix metalloproteases to measure the concentration of soluble MICA (sMICA) by ELISA to detect the therapeutic target. Furthermore, an FDA‐approved drug library was tested for the enzymatic inhibition of the targeted enzyme in an in vitro drug screening assay system. ADAM17 knockdown reduced sMICA levels and increased membrane‐bound MICA (mMICA) expression in HCC cells. In an in vitro drug screen using an FDA‐approved drug library, lomofungin, an antifungal drug, was found to strongly decrease ADAM17 activity. In HCC cells, mMICA expression was induced and sMICA production was inhibited in a dose‐dependent manner. These effects were cancelled upon ADAM17 knockdown, suggesting that lomofungin targeted ADAM17. Analysis of lomofungin analogs revealed the responsible functional groups. In summary, we suggest lomofungin to be an attractive agent for the immunological control of HCC, via the suppression of ADAM17. What's new? Targeting immune escape mechanisms of cancer cells is an attractive therapeutic approach. Here the authors follow up on a previous observation identifying membrane‐bound major histocompatibility complex class I‐related chain A (MICA) as a natural killer cell ligand on cancer cells. They show that ADAM17, a disintegrin and metalloprotease, sheds MICA as a decoy in hepatocellular carcinoma as soluble MICA levels increase after knockdown of ADAM17 or treatment with lomofungin, an antifungal drug newly found to inhibit ADAM17 activity. These data establish lomofungin as a new drug candidate for immunotherapy in liver cancer. [ABSTRACT FROM AUTHOR]

Objective To study the effects of silencing ADAM 17 on the proliferation, migration, and invasion of human thyroid papillary carcinoma K1 cells. Methods K1 cells were divided into the control group, nonsense shRNA group, and ADAM17-shRNA group. Nonsense-shRNA and ADAM17-shRNA were transfected into K1 cells of the nonsense shRNA group and ADAM17-shRNA group by lentiviral transfection technology. The same amount of PBS was added into the control group. The expression levels of ADAM 17 were detected by RT-PCR and Western blotting. MTT assay and flow cytometry were used to assess the cell proliferation ability and the cell cycle. The cell migration and invasion abilities were detected by Transwell migration and invasion assay, respectively. Results Compared with the control group and nonsense-shRNA group, the mRNA and protein expression levels of ADAM 17 significantly decreased in the ADAM17-shRNA group(all P < 0. 05) ; the optical density(OD value) of the ADAM17-shRNA group at 24 , 48 , and 72 h was significantly lower that that of the control group and nonsense-shRNA group (all P < 0. 05) ; in the ADAM17-shRNA group, the cells arrested in G0/G, phase, and significant difference was found between the ADAM 17-shRNA group and the control group and nonsense-shRNA group (all P <0.05) ; compared with the control group and nonsense-shRNA group, the transmembrane cells decreased sharply in the ADAM 17-shRNA group (all P <0.05). Conclusion Silencing ADAM 17 can inhibit the proliferation, migration, and invasion of K1 cells. [ABSTRACT FROM AUTHOR]

Background/Aims: Lipopolysaccharide (LPS)-activated monocytes/macrophages develop endotoxin tolerance in part by reducing cell surface toll-like receptor 4 (TLR4) through cluster of differentiation 14 (CD14)-dependent endocytosis. In case of endothelial cells, CD14 is expressed in low copy numbers as compared with monocytes/macrophages. Thus, we explored how endothelial cells regulate TLR4 expression after LPS stimulation. Methods: Cultured human aortic endothelial cells (HAECs) were treated with LPS. TLR4 expression was analyzed by Western blot analysis and immunofluorescence staining. A disintegrin and metalloprotease 17 (ADAM17) activity was measured using a fluorescent substrate. Results: TLR4 in cell lysate began to decrease within 30 min of LPS treatment with a maximal reduction at 2 h, and it was accompanied by an increase of N-terminal fragment of TLR4 in culture supernatant, indicating ectodomain shedding of the receptor. LPS activated p38 mitogen-activated protein kinase (p38 MAPK) and ADAM17, while LPS-induced ADAM17 activation was inhibited by SB203580, a p38 MAPK inhibitor. LPS-induced ectodomain shedding of TLR4 was attenuated by siRNA depletion of ADAM17 as well as TAPI-2 (an inhibitor of ADAM family) and SB203580. LPS pretreatment resulted in a blunted response of p38 MAPK activation to further LPS stimulation. In the cells depleted of ADAM17, LPS-induced p38 MAPK activation was prolonged and LPS-induced intercellular adhesion molecule-1 expression was potentiated. Conclusion: HAECs respond to LPS by rapid shedding of the ectodomain of TLR4 and thereby reduce the responsiveness to subsequent LPS exposure. ADAM17, downstream of p38 MAPK, is implicated in the ectodomain cleavage of TLR4. [ABSTRACT FROM AUTHOR]

Objective To investigate the effect of a disintegrin and metalloprotease 17 ( ADAM17 )-shRNA on the proliferation of human breast cancer MCF-7 cells and to explore its possible mechanism .Methods MCF-7 cells were di-vided into the nonsense group , control group ( PBS) and transfection group .Four specific ADAM17-shRNAs ( ADAM17-shRNA-1219, ADAM17-shRNA-1508, ADAM17-shRNA-1134 and ADAM17-shRNA-297) were designed and synthesized for ADAM17 gene.MCF-7 cells were transfected by electroporation .The nonsense group was transfected by ADAM 17-shNC, the control group was added with PBS , and the transfection group was transfected by ADAM 17-shRNA-1219, AD-AM17-shRNA-1508, ADAM17-shRNA-1134 and ADAM17-shRNA-297, respectively.The expression of ADAM17 mRNA, the expression of ADAM17 protein, cell growth curve, proliferation activity and cell cycle of MCF-7 cells was respectively detected by real-time PCR, Western blotting, iCELLigence and flow cytometry .Results Four ADAM17-shRNA se-quences inhibited the expression of ADAM 17 gene, the shRNA1219 vector had the highest inhibitory efficiency , and com-pared with the nonsense group and control group , there was significant difference (all P<0.05).The expression level of ADAM17 mRNA and protein in the transfection group was significantly lower than that in the nonsense group and control group (all P<0.05).The cell proliferation activity and cell growth rate of the transfection group were significantly lower than those of the nonsense group and the control group (all P<0.05).The percentage of cells in S phase and G 2/M phase decreased in the transfection group , and most of the cells stayed in G0/G1 phase, which were significantly different from those in the control group and nonsense group (all P<0.05).Conclusion ADAM17-shRNA can inhibit the proliferation of human breast cancer MCF-7 cells, and the mechanism is related to its silencing effect on ADAM 17 gene [ABSTRACT FROM AUTHOR]

Objective To investigate the relationship between a disintegrin and metalloprotease 17 (ADAM17) and transforming growth factor-α(TGF-α) activation in prostate cancer cells DU145.Methods We cultured the prostate cancer DU145 cells. Then, the cells were divided into the control group, NC group, ADAM17-siRNA1 group and the combination group. ADAM17 siRNA was transfected into both the ADAM17-siRNA1 group and combination group.The NC group was transfected with non-DAM17 related sequence siRNA.After the combination group was transfected for 72 h, 20 g/L TGF-α was added. After 72-hour transfection, the cells of the control group, NC group and ADAM17-siRNA1 group were collected. Western blotting was used to detect the protein expression of ADAM17, epidermal growth factor receptor (EGFR) and pEGFR. ELISA was used to detect the level of TGF-αprotein. We collected the supernatant intervened for15 mins of TGF-α from the control group, ADAM17-siRNA1 group and the combination group, and Western blotting was used to detect the protein expression of EGFR and pEGFR. Results The protein expression of AD-AM17, pEGFR and TGF-α in the ADAM17-siRNA1 group was lower than that of the control group (all P<0.05).There was no significant difference in EGFR expression between the control group, NC group and ADAM17-siRNA1 group (P>0.05).The expression of pEGFR in the combination group was higher than that in the control group and the ADAM17-siRNA1 group (P<0.05).There was no significant difference in the EGFR expression between the control group, ADAM17-siRNA1 group and the combination group (P>0.05). Conclusion The expression of ADAM17 in prostate cancer DU145 cells is correlated with the activation of TGF-α and EG-FR, and ADAM17 activates pEGFR expression by activating TGF-α. [ABSTRACT FROM AUTHOR]

Tumour necrosis factor-α (TNF-α) converting enzyme (TACE), also termed a disintegrin and metalloprotease 17 (ADAM17), is involved in multiple cell signalling pathways. Through the secretion of epidermal growth factor receptor (EGFR) ligands, ADAM17 can activate the EGFR and is involved in various downstream signalling pathways. The present study aimed to investigate whether ADAM17-induced EGFR transactivation is involved in microglial cell survival following spinal cord injury (SCI). Reverse transcription quantitative polymerase chain reaction and western blot analysis revealed that ADAM17 was overexpressed in a mouse model following SCI. A 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay demonstrated that the viability of human microglia and oligodendrocytes were significantly reduced in a time- and dose-dependent manner following treatment with the ADAM17 antagonist, TNF protease inhibitor 2. Hoechst 33258 staining and flow cytometric analysis revealed that inhibiting ADAM17 increased the rate of cellular apoptosis in neuronal and glial cell cultures, which was accompanied by increased cleavage of caspase-3. Western blot analysis demonstrated that inhibiting ADAM17 resulted in a reduction in the phosphorylation of the EGFR signalling pathway components and thereby impaired functional recovery, inhibited cell viability and prompted microglial apoptosis following SCI. Pre-treatment with the EGFR inhibitor, AG1478, rescued the ADAM17-mediated proliferation of microglial cells. These data demonstrated that ADAM17 contributed to microglial cell survival, predominantly by EGFR signalling, following SCI. [ABSTRACT FROM AUTHOR]

A disintegrin and metalloprotease (ADAM) 17 has been implicated in the tumor progression of various types of solid tumor; however, little is known about its role in non-small cell lung carcinoma (NSCLC). The present study evaluated whether the downregulation of ADAM17 affects cell proliferation, the cell cycle, cell migration and cell invasion in NSCLC. A recombinant lentiviral small hairpin RNA (shRNA) expression vector carrying ADAM17 was constructed and then infected into A549 cells, a human NSCLC cell line. Cell proliferation, cell cycle progression, cell migration and cell invasion were determined following the downregulation of ADAM17 by siRNA. It was revealed that downregulation of ADAM17 expression using an RNA silencing approach in A549 tumor cells significantly suppressed cell proliferation and invasion in vitro, and tumor growth in vivo. These data suggested that ADAM17 is an important regulator of the tumorigenic properties of human NSCLC and may be used as a potential anticancer therapeutic target in NSCLC. [ABSTRACT FROM AUTHOR]

Background and Objective Endopeptidases, such as neutral endopeptidase ( NEP), endothelin-converting enzyme-1 ( ECE-1) and a disintegrin and metalloprotease 17 ( ADAM17), are believed to have various important roles in oral mucosal and epidermal tissue for the regulation of defensive biological responses in the oral cavity, and their expression and activity are influenced by various factors, including oral diseases. However, knowledge concerning these endopeptidases in the oral cavity has been minimal until now. This study focused on three metalloendopeptidases - NEP, ECE-1 and ADAM17 - in the oral buccal mucosal epithelium of patients with periodontal diseases and investigated the relationship between their gene-expression levels and periodontal disease. Material and Methods The levels of expression of NEP, ECE-1 and ADAM17 m RNAs in tissue samples collected from the oral buccal mucosal epithelium of 61 patients were investigated by relative quantification using real-time RT- PCR analysis. information on oral and systemic health was obtained from the clinical record of each patient. Results Among the three groups, classified based on the diagnosis of periodontal diseases (healthy/gingivitis, early periodontitis and moderate/advanced periodontitis), the relative expression level of NEP m RNA was significantly increased in the early periodontitis group and in the moderate/advanced periodontitis group compared with that in the healthy/gingivitis group. Moreover, the relative expression levels of ECE1 and ADAM17 m RNAs were significantly increased in the moderate/advanced periodontitis group compared with those in the healthy/gingivitis group. The correlation coefficients between the mean relative expression levels of NEP and ECE1 m RNAs, NEP and ADAM17 m RNAs, and ECE1 and ADAM17 m RNAs were r = 0.758, r = 0.707 and r = 0.934, respectively ( p < 0.001). Furthermore, among the oral-related factors, there was a significant correlation between the number of sites with probing pocket depths of more than 4 mm and of more than 6 mm and the relative expression levels of NEP, ECE1 and ADAM17 m RNAs. In stepwise logistic regression models, high relative expression levels of ECE1 and ADAM17 m RNAs were significantly associated with moderate/advanced periodontitis. Conclusion The present study suggests that the severity of periodontal disease may be associated with the expression of metalloendopeptidase genes, including NEP, ECE1 and ADAM17, in the buccal mucosal epithelium. [ABSTRACT FROM AUTHOR]

Jia HP, Look DC, Tan P, Shi L, Hickey M, Gakhar L, Chappell MC, Wohlford-Lenane C, McCray PB Jr. Ectodomain shedding of angiotensin converting enzyme 2 in human airway epithelia. Am J Physiol Lung Cell Mo! Physiol 297: L84-L96, 2009. First published May 1, 2009; doi: 10.11 52/ajplung.0007 1 .2009.-Angiotensin-converting enzyme 2 (ACE2) is a terminal carboxypeptidase and the receptor for the SARS and NL63 coronaviruses (CoV). Loss of ACE2 function is implicated in severe acute respiratory syndrome (SARS) pathogenesis, but little is known about ACE2 biogenesis and activity in the airways. We report that ACE2 is shed from human airway epithelia, a site of SARS-CoV infection. The regulation of ACE2 release was investigated in polarized human airway epithelia. Constitutive generation of soluble ACE2 was inhibited by DPC 333, implicating a disintegrin and metalloprotease 17 (ADAM 17). Phorbol ester, ionomycin, endotoxin, and IL-β and TNFz acutely induced ACE2 release, further supporting that ADAM17 and ADAM1O regulate ACE2 cleavage. Soluble ACE2 was enzymatically active and partially inhibited virus entry into target cells. We determined that the ACE2 cleavage site resides between amino acid 7 16 and the putative transmembrane domain starting at amino acid 741. To reveal structural determinants underlying ACE2 release, several mutant and chimeric ACE2 proteins were engineered. Neither the juxtamembrane stalk region, transmembrane domain, nor the cytosolic domain was needed for constitutive ACE2 release. Interestingly, a point mutation in the ACE2 ectodomain, L584A, markedly attenuated shedding. The resultant ACE2-L584A mutant trafficked to the cell membrane and facilitated SARS-CoV entry into target cells, suggesting that the ACE2 ectodomain regulates its release and that residue L584 might be part of a putative sheddase "recognition motif." Thus ACE2 must be cell associated to serve as a CoV receptor and soluble ACE2 might play a role in modifying inflammatory processes at the airway mucosal surface. [ABSTRACT FROM AUTHOR]

The transmembrane and multidomain neural cell adhesion molecule (NCAM) plays important functional roles in the developing and adult nervous system. NCAM is proteolytically processed and appears in soluble forms in the cerebrospinal fluid and in serum under normal and pathological conditions. In this report, we present evidence that the metalloprotease a disintegrin and a metalloprotease (ADAM)17/tumour necrosis factor α converting enzyme (TACE) cleaves the polysialylated as well as the non-polysialylated transmembrane isoforms of NCAM, whereas the glycophosphatidylinositol-linked isoform of NCAM is not proteolytically cleaved. A truncated, enzymatically inactive mutant of TACE did not result in release of the NCAM110 cleavage product. Proteolytic cleavage was enhanced by a calmodulin-specific inhibitor and the actin-destabilizing agents cytochalasin D and latrunculin B. In contrast, the microtubule-stabilizing agent colchicine or microtubule-destabilizing agent paclitaxel did not affect the release of the 110-kDa fragment of NCAM. Neurite outgrowth from cerebellar microexplants was inhibited in the presence of the metalloprotease inhibitor GM 6001 on substrate-coated NCAM, but not on poly-l-lysine. Upon transfection of hippocampal neurones with an enzymatically inactive mutant of TACE, NCAM-stimulated neurite outgrowth was inhibited without affecting neurite outgrowth on poly-l-lysine, showing that proteolytic processing of NCAM by the metalloprotease TACE is involved in NCAM-mediated neurite outgrowth. [ABSTRACT FROM AUTHOR]

Growth of the axial and appendicular skeleton depends on endochondral ossification, which is controlled by tightly regulated cell–cell interactions in the developing growth plates. Previous studies have uncovered an important role of a disintegrin and metalloprotease 17 (ADAM17) in the normal development of the mineralized zone of hypertrophic chondrocytes during endochondral ossification. ADAM17 regulates EGF-receptor signaling by cleaving EGFR-ligands such as TGFα from their membrane-anchored precursor. The activity of ADAM17 is controlled by two regulatory binding partners, the inactive Rhomboids 1 and 2 (iRhom1, 2), raising questions about their role in endochondral ossification. To address this question, we generated mice lacking iRhom2 (iR2−/−) with floxed alleles of iRhom1 that were specifically deleted in chondrocytes by Col2a1-Cre (iR1∆Ch). The resulting iR2−/−iR1∆Ch mice had retarded bone growth compared to iR2−/− mice, caused by a significantly expanded zone of hypertrophic mineralizing chondrocytes in the growth plate. Primary iR2−/−iR1∆Ch chondrocytes had strongly reduced shedding of TGFα and other ADAM17-dependent EGFR-ligands. The enlarged zone of mineralized hypertrophic chondrocytes in iR2−/−iR1∆Ch mice closely resembled the abnormal growth plate in A17∆Ch mice and was similar to growth plates in Tgfα−/− mice or mice with EGFR mutations. These data support a model in which iRhom1 and 2 regulate bone growth by controlling the ADAM17/TGFα/EGFR signaling axis during endochondral ossification. [ABSTRACT FROM AUTHOR]

The metalloproteinase ADAM17 (a disintegrin and metalloprotease 17) controls EGF receptor (EGFR) signaling by liberating EGFR ligands from their membrane anchor. Consequently, a patient lacking ADAM17 has skin and intestinal barrier defects that are likely caused by lack of EGFR signaling, and Adam17(-/-) mice die perinatally with open eyes, like Egfr(-/-) mice. A hallmark feature of ADAM17-dependent EGFR ligand shedding is that it can be rapidly and posttranslationally activated in a manner that requires its transmembrane domain but not its cytoplasmic domain. This suggests that ADAM17 is regulated by other integral membrane proteins, although much remains to be learned about the underlying mechanism. Recently, inactive Rhomboid 2 (iRhom2), which has seven transmembrane domains, emerged as a molecule that controls the maturation and function of ADAM17 in myeloid cells. However, iRhom2(-/-) mice appear normal, raising questions about how ADAM17 is regulated in other tissues. Here we report that iRhom1/2(-/-) double knockout mice resemble Adam17(-/-) and Egfr(-/-) mice in that they die perinatally with open eyes, misshapen heart valves, and growth plate defects. Mechanistically, we show lack of mature ADAM17 and strongly reduced EGFR phosphorylation in iRhom1/2(-/-) tissues. Finally, we demonstrate that iRhom1 is not essential for mouse development but regulates ADAM17 maturation in the brain, except in microglia, where ADAM17 is controlled by iRhom2. These results provide genetic, cell biological, and biochemical evidence that a principal function of iRhoms1/2 during mouse development is to regulate ADAM17-dependent EGFR signaling, suggesting that iRhoms1/2 could emerge as novel targets for treatment of ADAM17/EGFR-dependent pathologies.

A disintegrin and metalloprotease 17 (ADAM17) appears to be recognized as an important player in tissue destruction and also exacerbation of inflammation related with increased activities of angiogenesis in several pathological conditions. To examine the modulation of serum levels of ADAM17 and inflammatory cytokines in patients with rheumatoid arthritis (RA) in response to therapy of abatacept (ABT). Twenty four patients with RA were enrolled in our study. Serum was collected immediately prior to (baseline) and 24 weeks after starting ABT therapy. Serum levels of ADAM17 and cytokines/chemokine were quantified using enzyme-linked immunosorbent assay. ADAM17 level was markedly higher in RA patients than in healthy individuals. Positive correlation was observed between the baseline ADAM17 and CX3CL1 at baseline. There was a significant overall reduction of RA disease activity (Disease Activity Score 28) from 4.73 to 2.79 after 24 weeks after the ABT therapy. Furthermore, there was a significant reduction in serum level of ADAM17 in RA patients, and the patients achieved clinical responses, and also clinical remission had a significant decrease in ADAM17 level and also levels of tumor necrosis factor α, IL-6 and CX3CL1 after 24 weeks of ABT therapy. Our results suggest that the suppression of ADAM17 secretion and function seems to be a crucial therapeutic target in the treatment of ABT in patients with RA.