Regulation Of ADAM17 Research Articles

Chrysophanol Attenuates Liver Fibrosis via Downregulation of Disintegrin and Metalloprotease 17 in the Thioacetamide Mouse Model

It is unclear whether chrysophanol affects liver fibrosis via the regulation of ADAM17, a multifaceted enzyme involved in immune regulation, central nervous system function, and cancer development. This article underscores that the chrysophanol treatment reduced collagen deposition in thioacetamide-induced liver fibrosis mice while downregulating ADAM17.

Photoreceptors inhibit pathological retinal angiogenesis through transcriptional regulation of Adam17 via c-Fos

Pathological retinal angiogenesis profoundly impacts visual function in vascular eye diseases, such as retinopathy of prematurity (ROP) in preterm infants and age-related macular degeneration in the elderly. While the involvement of photoreceptors in these diseases is recognized, the underlying mechanisms remain unclear. This study delved into the pivotal role of photoreceptors in regulating abnormal retinal blood vessel growth using an oxygen-induced retinopathy (OIR) mouse model through the c-Fos/A disintegrin and metalloprotease 17 (Adam17) axis. Our findings revealed a significant induction of c-Fos expression in rod photoreceptors, and c-Fos depletion in these cells inhibited pathological neovascularization and reduced blood vessel leakage in the OIR mouse model. Mechanistically, c-Fos directly regulated the transcription of Adam17 a shedding protease responsible for the production of bioactive molecules involved in inflammation, angiogenesis, and cell adhesion and migration. Furthermore, we demonstrated the therapeutic potential by using an adeno-associated virus carrying a rod photoreceptor-specific short hairpin RNA against c-fos which effectively mitigated abnormal retinal blood vessel overgrowth, restored retinal thickness, and improved electroretinographic (ERG) responses. In conclusion, this study highlights the significance of photoreceptor c-Fos in ROP pathology, offering a novel perspective for the treatment of this disease.

Semaphorin 4B is an ADAM17-cleaved adipokine that inhibits adipocyte differentiation and thermogenesis

ObjectiveThe metalloprotease ADAM17 (also called TACE) plays fundamental roles in homeostasis by shedding key signaling molecules from the cell surface. Although its importance for the immune system and epithelial tissues is well-documented, little is known about the role of ADAM17 in metabolic homeostasis. The purpose of this study was to determine the impact of ADAM17 expression, specifically in adipose tissues, on metabolic homeostasis. MethodsWe used histopathology, molecular, proteomic, transcriptomic, in vivo integrative physiological and ex vivo biochemical approaches to determine the impact of adipose tissue-specific deletion of ADAM17 upon adipocyte and whole organism metabolic physiology. ResultsADAM17adipoq-creΔ/Δ mice exhibited a hypermetabolic phenotype characterized by elevated energy consumption and increased levels of adipocyte thermogenic gene expression. On a high fat diet, these mice were more thermogenic, while exhibiting elevated expression levels of genes associated with lipid oxidation and lipolysis. This hypermetabolic phenotype protected mutant mice from obesogenic challenge, limiting weight gain, hepatosteatosis and insulin resistance. Activation of beta-adrenoceptors by the neurotransmitter norepinephrine, a key regulator of adipocyte physiology, triggered the shedding of ADAM17 substrates, and regulated ADAM17 expression at the mRNA and protein levels, hence identifying a functional connection between thermogenic licensing and the regulation of ADAM17. Proteomic studies identified Semaphorin 4B (SEMA4B), as a novel ADAM17-shed adipokine, whose expression is regulated by physiological thermogenic cues, that acts to inhibit adipocyte differentiation and dampen thermogenic responses in adipocytes. Transcriptomic data showed that cleaved SEMA4B acts in an autocrine manner in brown adipocytes to repress the expression of genes involved in adipogenesis, thermogenesis, and lipid uptake, storage and catabolism. ConclusionsOur findings identify a novel ADAM17-dependent axis, regulated by beta-adrenoceptors and mediated by the ADAM17-cleaved form of SEMA4B, that modulates energy balance in adipocytes by inhibiting adipocyte differentiation, thermogenesis and lipid catabolism.

Potential Similarities in Sex Difference in Key Genes and Their Expression, Network, EQTL and Pathways between COVID-19 and Chronic Kidney Disease Based on Mouse Model.

COVID-19 and chronic kidney disease (CKD) share similarity in sex bias and key genes in the disease pathway of sex difference. We investigated the sex difference of molecular pathways of four key players of these two diseases using an existing large set of whole genome expression profiles from the kidneys of female and male mouse models. Our data show that there is little to no correlation at the whole genome expression level between female and male mice among these four genes. There are considerable sex differences among genes in upstream regulation, Ace2 complex interaction, and downstream pathways. Snap25 and Plcb4 may play important roles in the regulation of the expression level of Adam17, Tmprss2, and Cd146 in females. In males, Adh4 is a candidate gene for the regulation of Adam17, while Asl, Auts2, and Rabger1 are candidates for Tmprss2. Within the Ace2 complex, Cd146 directly influences the expression level of Adam17 and Ace2 in the female, while in the male Adam potentially has a stronger influence on Ace2 than that of Tmprss2. Among the top 100 most related genes, only one or two genes from four key genes and 11 from the control B-Actin were found to be the same between sexes. Among the top 10 sets of genes in the downstream pathway of Ace2, only two sets are the same between the sexes. We concluded that these known key genes and novel genes in CKD may play significant roles in the sex difference in the CKD and COVID-19 disease pathways.

Lentiviral CRISPR-guided RNA library screening identified Adam17 as an upstream negative regulator of Procr in mammary epithelium

BackgroundProtein C receptor (Procr) has recently been shown to mark resident adult stem cells in the mammary gland, vascular system, and pancreatic islets. More so, high Procr expression was also detected and used as indicator for subsets of triple-negative breast cancers (TNBCs). Previous study has revealed Procr as a target of Wnt/β-catenin signaling; however, direct upstream regulatory mechanism of Procr remains unknown. To comprehend the molecular role of Procr during physiology and pathology, elucidating the upstream effectors of Procr is necessary. Here, we provide a system for screening negative regulators of Procr, which could be adapted for broad molecular analysis on membrane proteins.ResultsWe established a screening system which combines CRISPR-Cas9 guided gene disruption with fluorescence activated cell sorting technique (FACS). CommaDβ (murine epithelial cells line) was used for the initial Procr upstream effector screening using lentiviral CRISPR-gRNA library. Shortlisted genes were further validated through individual lentiviral gRNA infection followed by Procr expression evaluation. Adam17 was identified as a specific negative inhibitor of Procr expression. In addition, MDA-MB-231 cells and Hs578T cells (human breast cancer cell lines) were used to verify the conserved regulation of ADAM17 over PROCR expression.ConclusionWe established an efficient CRISPR-Cas9/FACS screening system, which identifies the regulators of membrane proteins. Through this system, we identified Adam17 as the negative regulator of Procr membrane expression both in mammary epithelial cells and breast cancer cells.

Identification of Novel MicroRNAs as Promising Therapeutics for SARS-CoV-2 by Regulating the EGFR-ADAM17 Axis: An In Silico Analysis.

Cancer patients contracting SARS-CoV-2 encounter additional challenges due to inflammatory bursts and lymphopenia, which may aggravate breast cancer prognosis. In this in silico analysis, we identified the potential of miRNAs as new therapeutic targets to treat breast cancer patients infected with COVID-19 via the regulation of ADAM17 and EGFR expression microRNAs.

Differential Induction of the ADAM17 Regulators iRhom1 and 2 in Endothelial Cells.

Background: Endothelial function significantly depends on the proteolytic release of surface expressed signal molecules, their receptors and adhesion molecules via the metalloproteinase ADAM17. The pseudoproteases iRhom1 and 2 independently function as adapter proteins for ADAM17 and are essential for the maturation, trafficking, and activity regulation of ADAM17. Bioinformatic data confirmed that immune cells predominantly express iRhom2 while endothelial cells preferentially express iRhom1.Objective: Here, we investigate possible reasons for higher iRhom1 expression and potential inflammatory regulation of iRhom2 in endothelial cells and analyze the consequences for ADAM17 maturation and function.Methods: Primary endothelial cells were cultured in absence and presence of flow with and without inflammatory cytokines (TNFα and INFγ). Regulation of iRhoms was studied by qPCR, involved signaling pathways were studied with transcriptional inhibitors and consequences were analyzed by assessment of ADAM17 maturation, surface expression and cleavage of the ADAM17 substrate junctional adhesion molecule JAM-A.Results: Endothelial iRhom1 is profoundly upregulated by physiological shear stress. This is accompanied by a homeostatic phenotype driven by the transcription factor KLF2 which is, however, only partially responsible for regulation of iRhom1. By contrast, iRhom2 is most prominently upregulated by inflammatory cytokines. This correlates with an inflammatory phenotype driven by the transcription factors NFκB and AP-1 of which AP-1 is most relevant for iRhom2 regulation. Finally, shear stress exposure and inflammatory stimulation have independent and no synergistic effects on ADAM17 maturation, surface expression and JAM-A shedding.Conclusion: Conditions of shear stress and inflammation differentially upregulate iRhom1 and 2 in primary endothelial cells which then results in independent regulation of ADAM17.

IRhom2: An Emerging Adaptor Regulating Immunity and Disease.

The rhomboid family are evolutionary conserved intramembrane proteases. Their inactive members, iRhom in Drosophila melanogaster and iRhom1 and iRhom2 in mammals, lack the catalytic center and are hence labelled “inactive” rhomboid family members. In mammals, both iRhoms are involved in maturation and trafficking of the ubiquitous transmembrane protease a disintegrin and metalloprotease (ADAM) 17, which through cleaving many biologically active molecules has a critical role in tumor necrosis factor alpha (TNFα), epidermal growth factor receptor (EGFR), interleukin-6 (IL-6) and Notch signaling. Accordingly, with iRhom2 having a profound influence on ADAM17 activation and substrate specificity it regulates these signaling pathways. Moreover, iRhom2 has a role in the innate immune response to both RNA and DNA viruses and in regulation of keratin subtype expression in wound healing and cancer. Here we review the role of iRhom2 in immunity and disease, both dependent and independent of its regulation of ADAM17.

Elevated MicroRNA-326 Levels Regulate the IL-23/IL-23R/Th17 Cell Axis in Hashimoto's Thyroiditis by Targeting a Disintegrin and Metalloprotease 17.

Background: MicroRNAs (miRNAs) are a class of critical epigenetic regulators involved in several autoimmune diseases. Our previous study reported an miR-326-induced increase in T helper (Th) 17 cells in a mouse model of Hashimoto's thyroiditis (HT), but the pathogenic effect of miR-326 in HT patients has not been verified. The goal of the present study was to explore the pathogenic role of miR-326 and its underlying molecular mechanism in HT patients. Methods: A total of 58 HT patients and 55 normal controls were enrolled in this study. We examined whether Th17 cells and miR-326 were aberrantly altered in the peripheral blood mononuclear cells (PBMCs) of HT patients with flow cytometry and real-time polymerase chain reaction. Levels of membrane interleukin (IL)-23R (mIL-23R) were determined by flow cytometry and Western blot to explore the critical role of mIL-23R in the development of Th17 cells. Isolated CD3+ T cells were used to further investigate the ectodomain shedding of mIL-23R by a disintegrin and metalloprotease (ADAM17). Furthermore, miR-326 inhibitor and mimics were transfected into PBMCs derived from HT patients and healthy controls to verify the regulation of ADAM17 by miR-326. Results: We observed elevated miR-326 levels in the PBMCs of HT patients compared with those in the PBMCs of healthy controls. Consistent with IL-23-induced STAT3 overactivation, substantially more HT patient-derived PBMCs differentiated into Th17 cells under polarization culture conditions, which may, at least in part, have resulted from enhanced mIL-23R levels. Furthermore, ADAM17, an ectodomain sheddase of mIL-23R, was targeted and negatively regulated by miR-326. Inhibiting ADAM17 might attenuate the ectodomain shedding of mIL-23R. Conclusions: Our findings suggest that the effect of miR-326 on the IL-23/IL-23R/Th17 cell axis in HT patients might be partially due to the targeting of ADAM17.

ADAM-17 is a poor prognostic indicator for patients with hilar cholangiocarcinoma and is regulated by FoxM1

BackgroundA-disintegrin and metalloproteinases (ADAMs) are members of a family of multidomain transmembrane and secreted proteins. Specific ADAMs are upregulated in human cancers and correlated with tumor progression and poor outcome, but rarely studied in human hilar cholangiocarcinoma (HC). This study aimed to explore the expression profiles of ADAMs and their potential underlying mechanisms promoting cancer progression.MethodsmRNA expression of ADAM-9, − 10, − 11, − 12, − 15, − 17, − 28, and − 33 was analyzed in human hilar cholangiocarcinoma (HC) samples. Immunohistochemical (IHC) analysis was used to detect the expression of ADAM-10, − 17, − 28, and FoxM1 in HC. The regulation of ADAM-17 by FoxM1 and their functional study was investigated in vivo and in vitro.ResultsADAM-10, − 17, and − 28 were upregulated in tumors compared with matched non-cancerous tissues. IHC analysis revealed increased expression of ADAM-10, − 17, and − 28 in HC cells, and ADAM17 seems to be an independent prognostic factor. ADAM-17 is regulated by FoxM1. A decrease in the expression of ADAM-17 by silencing FoxM1 led to an inhibition of cell proliferation, tumor growth, and the production of tumor necrosis factor α. IHC analysis showed co-expression of FoxM1 and ADAM-17 in HC specimens.ConclusionsThe findings of the present study show an important role of the cross-talk among FoxM1, ADAM-17, and TNFa in HC development and progression.

Abstract 9: Involvement of Caveolin-1 in Vascular Remodeling and Inflammation Induced by Angiotensin II

We have recently reported that caveolin-1 (Cav1) enriched membrane microdomains in vascular smooth muscle cells (VSMC) mediate a metalloprotease ADAM17-dependent EGF receptor (EGFR) transactivation, which is linked to vascular remodeling induced by AngII. We have tested our hypothesis that Cav1, a major structural protein of caveolae, plays a critical role for development of vascular remodeling by AngII via regulation of ADAM17 and EGFR. Here, 8 week old male Cav1-/- and control Cav+/+ wild-type mice (WT) were infused with AngII (1 μg/kg/min) for 2 weeks to induce vascular remodeling and hypertension. Upon AngII infusion, histological assessments demonstrated medial hypertrophy and perivascular fibrosis of coronary and renal arteries in WT mice compared with saline-infused control mice. The AngII-infused WT mice also showed a phenotype of cardiac hypertrophy with increased HW/BW ratio (mg/g: 8.0±0.6 vs 5.7±0.7 p<0.01) compared with WT control. In contrast to AngII-infused WT mice, Cav1-/- mice with AngII showed attenuation of vascular remodeling but not cardiac hypertrophy ; HW/BW ratio (8.6±0.5 vs 6.4±0.2 p<0.05). Similar levels of AngII-induced hypertension were observed in both WT and Cav1-/- mice assessed by telemetry (MAP mmHg: 142±9 vs 154±20). In WT mice, Ang II enhanced ADAM17 expression and phospho-Tyr EGFR staining in heart and kidney vasculature. These events were attenuated in vessels from Cav1-/- mice infused with AngII. In addition, IHC analysis revealed less ER stress in heart and kidney vasculature of AngII-infused Cav1-/- mice compared with WT mice. Enhanced Cav1 and VCAM-1 expression were also observed in the aorta from AngII-infused WT mice but not in Cav1-/- aorta. These data suggest that Cav1 and presumably vascular caveolae play critical roles for vascular remodeling and inflammation via induction of ADAM17 and activation of EGFR independent of blood pressure or cardiac hypertrophy regulation.

Abstract 14133: Angiotensin II-induced Vascular Fibrosis was Prevented by Caveolin-1 Deficient Mice

Introduction: We have recently reported that caveolin-1 (Cav1) enriched membrane microdomains in vascular smooth muscle cells (VSMC) mediates a metalloprotease ADAM17-dependent EGF receptor (EGFR) transactivation, which is linked to vascular remodeling induced by AngII. Hypothesis: We have tested our hypothesis that Cav1, a major structural protein of caveolae, plays a critical role in AngII-mediated vascular remodeling via regulation of ADAM17 and EGFR. Methods: In vivo: 8 week old male Cav1-/- and control Cav+/+ wild-type mice (WT) were infused with AngII (1 μg/kg/min) for 2 weeks to induce vascular remodeling and hypertension. In vitro: Rat aortic VSMCs were cultured and stimulated with 100 nM AngII. Results: Upon AngII infusion, histological assessments demonstrated medial hypertrophy and perivascular fibrosis of coronary and renal arteries in WT mice compared to sham-operated mice. The AngII-infused WT mice also showed a phenotype of cardiac hypertrophy with increased HW/BW ratio (mg/g: 8.0±0.6 vs 5.7±0.7 p<0.01) compared with sham-operated WT control. In contrast, vascular remodeling but not cardiac hypertrophy were attenuated in Cav1-/- mice with AngII infusion; HW/BW ratio (8.6±0.5 vs 6.4±0.2 p<0.05) compared to sham-operated mice. However, AngII induced similar levels of hypertension in both WT and Cav1-/- mice as assessed by telemetry (MAP mmHg: 142±9 vs 154±20). AngII infusion in WT mice enhanced ADAM17 and phospho-Tyr EGFR staining in heart and kidney vasculature, whereas AngII-infused Cav1-/- mice showed diminished ADAM17 and phospho-Tyr EGFR staining within the vasculature. In addition, IHC analyses revealed reduced vascular ER stress in heart and kidney samples of AngII-infused Cav1-/- mice compared to WT mice. In cultured rat VSMC, Cav1 silencing attenuated induction and activation of ADAM17 by AngII. Conclusions: These data suggest that Cav1, and presumably caveolae microdomains, plays a critical role in vascular fibrosis via vascular induction of ADAM17 and activation of EGFR independent of blood pressure or cardiac hypertrophy regulation.

Abstract P108: Caveolin-1 is Required for Vascular Remodeling Induced by Angiotensin Ii

We have recently reported that caveolin-1 (Cav1) enriched membrane microdomains in vascular smooth muscle cells (VSMC) mediates a metalloprotease ADAM17-dependent EGF receptor (EGFR) transactivation, which is linked to vascular remodeling induced by AngII. We have tested our hypothesis that Cav1, a major structural protein of caveolae, plays a critical role in AngII-mediated vascular remodeling via regulation of ADAM17 and EGFR. 8 week old male Cav1-/- and control Cav+/+ wild-type mice (WT) were infused with AngII (1 μg/kg/min) for 2 weeks to induce vascular remodeling and hypertension. Upon AngII infusion, histological assessments demonstrated medial hypertrophy and perivascular fibrosis of coronary and renal arteries in WT mice compared to sham-operated mice. The AngII-infused WT mice also showed a phenotype of cardiac hypertrophy with increased HW/BW ratio (mg/g: 8.0±0.6 vs 5.7±0.7 p<0.01) compared with sham-operated WT control. In contrast, vascular remodeling but not cardiac hypertrophy were attenuated in Cav1-/- mice with AngII infusion; HW/BW ratio (8.6±0.5 vs 6.4±0.2 p<0.05) compared to sham-operated mice. However, AngII induced similar levels of hypertension in both WT and Cav1-/- mice as assessed by telemetry (MAP mmHg: 142±9 vs 154±20). AngII infusion in WT mice enhanced ADAM17 and phospho-Tyr EGFR staining in heart and kidney vasculature, whereas AngII-infused Cav1-/- mice showed diminished ADAM17 and phospho-Tyr EGFR staining within the vasculature. In addition, IHC analyses revealed reduced vascular ER stress in heart and kidney samples of AngII-infused Cav1-/- mice compared to WT mice. Expression of Cav1 was predominantly observed within the endothelium and was enhanced upon AngII infusion in WT mice. These data suggest that Cav1, and presumably endothelial caveolae microdomains, plays a critical role in vascular remodeling via vascular induction of ADAM17 and activation of EGFR independent of blood pressure or cardiac hypertrophy regulation.

Estrogen upregulates MICA/B expression in human non-small cell lung cancer through the regulation of ADAM17

Estrogen is involved in promoting lung cancer cell division and metastasis. MICA and MICB function as ligands for NKG2D, an important immunoreceptor expressed on natural killer (NK) cells. However, whether estrogen regulates MICA/B expression and affects tumor immune escape remains unknown. In this study, we measured the mRNA levels of MICA, MICB and ADAM17in non-small cell lung cancer (NSCLC) cell lines treated with estrogen. Surface expression of MICA/B on LTEP-a2 and A549 was detected using flow cytometry. We demonstrate that both mRNA and secretory protein levels of MICA/B in lung adenocarcinoma cell lines were upregulated by estradiol. Estradiol enhanced the expression of ADAM17, which was associated with the secretion of MICA/B. This secretion of MICA/B downregulated the NKG2D receptor on the surface of NK92 cells and impaired the cytotoxic activity of NK cells. Estradiol enhanced the expression of ADAM17, which was associated with the secretion of MICA/B. Furthermore, a significant correlation between the concentration of estradiol and the expression of MICA was found in tumor tissues of NSCLC patients. Therefore, we conclude that estrogen can regulate the expression and secretion of MICA/B through ADAM17, which helps lung cancer cells escape NKG2D-mediated immune surveillance.

IRHOM2-dependent regulation of ADAM17 in cutaneous disease and epidermal barrier function

iRHOM2 is a highly conserved, catalytically inactive member of the Rhomboid family, which has recently been shown to regulate the maturation of the multi-substrate ectodomain sheddase enzyme ADAM17 (TACE) in macrophages. Dominant iRHOM2 mutations are the cause of the inherited cutaneous and oesophageal cancer-susceptibility syndrome tylosis with oesophageal cancer (TOC), suggesting a role for this protein in epithelial cells. Here, using tissues derived from TOC patients, we demonstrate that TOC-associated mutations in iRHOM2 cause an increase in the maturation and activity of ADAM17 in epidermal keratinocytes, resulting in significantly upregulated shedding of ADAM17 substrates, including EGF-family growth factors and pro-inflammatory cytokines. This activity is accompanied by increased EGFR activity, increased desmosome processing and the presence of immature epidermal desmosomes, upregulated epidermal transglutaminase activity and heightened resistance to Staphylococcal infection in TOC keratinocytes. Many of these features are consistent with the presence of a constitutive wound-healing-like phenotype in TOC epidermis, which may shed light on a novel pathway in skin repair, regeneration and inflammation.

Abstract 47: Caveolin-1 is Critical for Abdominal Aortic Aneurysm Induced by Angiotensin II

Abdominal aortic aneurysm (AAA) is a significant cause of mortality for adults aged >60 years. Accumulating evidence suggests a role of angiotensin II (Ang II) in abdominal aortic aneurysm (AAA) formation. However, the Ang II-sensitive proximal signaling events primarily responsible for AAA formation remain unclear. We recently reported that caveolin-1 (Cav1) enriched membrane microdomains in vascular smooth muscle cells (VSMC) mediate a metalloprotease ADAM17-dependent EGF receptor (EGFR) transactivation, which is linked to vascular remodeling induced by Ang II. Given that ADAM17 expression is one of the key features in AAA, we have tested our hypothesis that Cav1, a major structural protein of caveolae, plays a critical role for development of AAA by Ang II via regulation of ADAM17. 8 week old male Cav1-/- and the control C57Bl/6 wild-type mice (WT) were co-infused with Ang II (1 μg/kg/min) and β-aminopropionitrile (BAPN: 150mg/kg/day) for 4 weeks to induce AAA. In WT with the co-infusion, 58% (14/24) were dead due to aortic rupture/dissection. All surviving WT with co-infusion had AAA with max diameter (mm) of 2.6±0.18 vs 0.93±0.09 with saline infusion (p<0.01). In contrast, we found that Cav1-/- with co-infusion did not die or develop AAA. The max diameter (mm) of AAA in Cav1-/- with co-infusion was 1.0±0.04 vs 1.1±0.06 with saline infusion (n=7). In contrast, both WT and Cav1-/- with the co-infusion developed hypertension assessed by telemetry (MAP mmHg: 151±5 vs 161±7). We found an increased expression of ADAM17 by IHC and qPCR, and enhanced phosphorylation of EGFR by IHC in WT abdominal aortae with aneurysms. These events were markedly attenuated in Cav1-/- aorta with co-infusion (ADAM17/18S mRNAx10,000 = 3.08±0.71 vs 0.97±0.42 p<0.05, n=4). Furthermore, Cav1-/- aortae showed less ER and oxidative stress compared to WT aortae assessed by IHC. In addition, Cav1 silencing induced by adenovirus encoding Cav1 targeting siRNA embedded miRNA in cultured vascular smooth muscle cells prevented Ang II-induced ADAM17 induction and activation. In conclusion, Cav1 and presumably vascular caveolae microdomains appear to play a critical role in the formation of AAA by Ang II via regulation of the ADAM17/EGFR signaling and subsequent ER/oxidative stress.

The unfolded protein response controls induction and activation of ADAM17/TACE by severe hypoxia and ER stress

The family of ADAM (a disintegrin and metalloproteinase) proteins has been implicated in tumor initiation and progression. ADAM17/tumor necrosis factor-α (TNFα)-converting enzyme (TACE) has been initially recognized to release TNFα as well as its receptors (TNFRs) from the membrane. ADAM17, TNFα and TNFR have been found upregulated in cancer patients, although the underlying mechanisms remain largely unknown. As hypoxia is a hallmark of cancer that can lead to severe stress conditions accumulating in endoplasmic reticulum (ER) stress and the unfolded protein response (UPR), we investigated the role of these stress conditions in the regulation of ADAM17 and release of TNFR1.We found that severe hypoxia induced ADAM17 expression and activity. Although hypoxia-inducible factor 1α (HIF1α) was important to maintain basal ADAM17 mRNA levels during moderate hypoxia, it was not sufficient to induce ADAM17 levels under severe hypoxia. Instead, we found that ADAM17 induction by severe hypoxia can be mimicked by ER stressors such as Thapsigargin and occurs as a consequence of the activation of the PERK/eIF2α/ATF4 and activating transcription factor 6 (ATF6) arms of UPR in several tumor cell lines. ADAM17 expression was also increased in xenografts displaying ER stress because of treatment with the vascular endothelial growth factor (VEGF) inhibitory antibody Bevacizumab. Additionally, severe hypoxia and ER stress activated ADAM17 and ectodomain shedding of TNFR1 involving mitogen-activated protein (MAP) kinases and reactive oxygen species (ROS). Collectively, these results show that ADAM17 is a novel UPR-regulated gene in response to severe hypoxia and ER stress, which is actively involved in the release of TNFR1 under these conditions. These data provide a novel link between severe hypoxic stress conditions and inflammation in the tumor environment.

Caveolin-1 negatively regulates a metalloprotease-dependent epidermal growth factor receptor transactivation by angiotensin II

A metalloprotease, ADAM17, mediates the generation of mature ligands for the epidermal growth factor receptor (EGFR). This is the key signaling step by which angiotensin II (AngII) induces EGFR transactivation leading to hypertrophy and migration of vascular smooth muscle cells (VSMCs). However, the regulatory mechanism of ADAM17 activity remains largely unclear. Here we hypothesized that caveolin-1 (Cav1), the major structural protein of a caveolae, a membrane microdomain, is involved in the regulation of ADAM17. In cultured VSMCs, infection of adenovirus encoding Cav1 markedly inhibited AngII-induced EGFR ligand shedding, EGFR transactivation, ERK activation, hypertrophy and migration, but not intracellular Ca2+ elevation. Methyl-β-cyclodextrin and filipin, reagents that disrupt raft structure, both stimulated an EGFR ligand shedding and EGFR transactivation in VSMCs. In addition, non-detergent sucrose gradient membrane fractionations revealed that ADAM17 cofractionated with Cav1 in lipid rafts. These results suggest that lipid rafts and perhaps caveolae provide a negative regulatory environment for EGFR transactivation linked to vascular remodeling induced by AngII. These novel findings may provide important information to target cardiovascular diseases under the enhanced renin angiotensin system.

Thiol isomerases negatively regulate the cellular shedding activity of ADAM17

ADAM17 (where ADAM is 'a disintegrin and metalloproteinase') can rapidly modulate cell-surface signalling events by the proteolytic release of soluble forms of proligands for cellular receptors. Many regulatory pathways affect the ADAM17 sheddase activity, but the mechanisms for the activation are still not clear. We have utilized a cell-based ADAM17 assay to show that thiol isomerases, specifically PDI (protein disulfide isomerase), could be responsible for maintaining ADAM17 in an inactive form. Down-regulation of thiol isomerases, by changes in the redox environment (for instance as elicited by phorbol ester modulation of mitochondrial reactive oxygen species) markedly enhanced ADAM17 activation. On the basis of ELISA binding studies with novel fragment antibodies against ADAM17 we propose that isomerization of the disulfide bonds in ADAM17, and the subsequent conformational changes, form the basis for the modulation of ADAM17 activity. The shuffling of disulfide bond patterns in ADAMs has been suggested by a number of recent adamalysin crystal structures, with distinct disulfide bond patterns altering the relative orientations of the domains. Such a mechanism is rapid and reversible, and the role of thiol isomerases should be investigated further as a potential factor in the redox regulation of ADAM17.

Differential expression and regulation of ADAM17 and TIMP3 in acute inflamed intestinal epithelia

The acute phase of Crohn's disease (CD) is characterized by a large afflux of polymorphonuclear leukocytes (PMNL) into the mucosa and by the release of TNF-alpha. Conversion of inactive TNF-alpha into an active form requires the cleavage of a transmembrane TNF-alpha precursor by the TNF-alpha-converting enzyme (ADAM17), a protease mainly regulated by the tissue inhibitor of metalloproteinase 3 (TIMP3). The aim of the present study was to investigate in an in vitro model of PMNL transepithelial migration and in the intestinal mucosa of patients with CD the expression and regulation of ADAM17 and TIMP3 in intestinal epithelial cells (IEC). ADAM17 and TIMP3 expression was analyzed by Western blotting, RT-PCR, confocal microscopy, and immunohistochemistry by using the T84 model and digestive biopsies. ADAM17 expression in IEC was increased at a posttranscriptional level during the early phase (from 2 to 4 h) of PMNL transepithelial migration whereas TIMP3 was only increased 24 h later. TNF-alpha induced an early upregulation of ADAM17 in T84 cells, whereas PMNL adhesion, H(2)O(2), or epithelial tight junction opening alone did not affect the amount of ADAM17. Immunohistochemistry of intestinal biopsies revealed that strong expression of ADAM17 was associated with a high activity of CD. In contrast, TIMP3 was very poorly expressed in these biopsies. ADAM17 and TIMP3 profiling did not correlated with the NOD2/CARD15 status. The ADAM17 activity was higher both in the early phase of PMNL transepithelial migration and in active CD. These results showed early posttranscriptional upregulation of ADAM17 in IEC linked to PMNL transepithelial migration and a high activity of CD.