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1.
Biochim Biophys Acta Mol Basis Dis ; 1866(3): 165627, 2020 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-31785407

RESUMO

Macrophages play an important role in aldosterone-induced myocardial fibrosis, in which the first key steps are macrophage recruitment and infiltration. We hypothesized that IL-6 may be a key mediator of aldosterone-induced macrophage recruitment and infiltration. To test this hypothesis, we designed cell studies with a human monocytic cell line THP-1 that with monocyte/macrophage functions to explore the signaling pathway of aldosterone-induced macrophage infiltration, and further investigated the phenomenon and consequent pathway in aldosterone-infused mice studies. The results showed that aldosterone induced the expression of IL-6 via mineralocorticoid receptors, and enhanced THP-1 cell migration and infiltration. Further experiments using a protease array and siRNA revealed that expressions of MMP-1 and MMP-9 were associated with aldosterone-induced macrophage infiltration. In addition, aldosterone-induced MMP-1 and MMP-9 expressions were mediated via cyclooxygenase-II and prostaglandin E2/EP-2 and EP-4 receptors. In aldosterone-infused mice, mRNA expressions of MMP-1, MMP-9 and COX-2 in peripheral blood monocytic cells were significantly increased. Moreover, the number of mouse macrophage-restricted F4/80 protein-positive cells in the myocardium was significantly higher in the aldosterone-infused mice compared with control mice. The increase in F4/80-positive cells in the myocardium was suppressed in the aldosterone-infused mice with the aldosterone antagonist eplerenone or anti-IL-6 antibody treatment. In conclusion, interleukin-6 played an important role in aldosterone-induced macrophage recruitment and infiltration in the myocardium.

2.
Cell Mol Life Sci ; 77(2): 331-350, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31209506

RESUMO

Disintegrin and metalloproteinases (ADAMs) 10 and 17 can release the extracellular part of a variety of membrane-bound proteins via ectodomain shedding important for many biological functions. So far, substrate identification focused exclusively on membrane-anchored ADAM10 and ADAM17. However, besides known shedding of ADAM10, we identified ADAM8 as a protease capable of releasing the ADAM17 ectodomain. Therefore, we investigated whether the soluble ectodomains of ADAM10/17 (sADAM10/17) exhibit an altered substrate spectrum compared to their membrane-bound counterparts. A mass spectrometry-based N-terminomics approach identified 134 protein cleavage events in total and 45 common substrates for sADAM10/17 within the secretome of murine cardiomyocytes. Analysis of these cleavage sites confirmed previously identified amino acid preferences. Further in vitro studies verified fibronectin, cystatin C, sN-cadherin, PCPE-1 as well as sAPP as direct substrates of sADAM10 and/or sADAM17. Overall, we present the first degradome study for sADAM10/17, thereby introducing a new mode of proteolytic activity within the protease web.


Assuntos
Proteína ADAM10/metabolismo , Proteína ADAM17/metabolismo , Secretases da Proteína Precursora do Amiloide/metabolismo , Proteínas de Membrana/metabolismo , Metaloproteases/metabolismo , Aminoácidos/metabolismo , Animais , Linhagem Celular , Células HEK293 , Humanos , Camundongos , Miócitos Cardíacos/metabolismo
3.
Gut ; 2019 Nov 04.
Artigo em Inglês | MEDLINE | ID: mdl-31685519

RESUMO

OBJECTIVE: Cancer-associated fibroblasts (CAFs) influence the tumour microenvironment and tumour growth. However, the role of CAFs in colorectal cancer (CRC) development is incompletely understood. DESIGN: We quantified phosphorylation of STAT3 (pSTAT3) expression in CAFs of human colon cancer tissue using a tissue microarray (TMA) of 375 patients, immunofluorescence staining and digital pathology. To investigate the functional role of CAFs in CRC, we took advantage of two murine models of colorectal neoplasia and advanced imaging technologies. In loss-of-function and gain-of-function experiments, using genetically modified mice with collagen type VI (COLVI)-specific signal transducer and activator of transcription 3 (STAT3) targeting, we evaluated STAT3 signalling in fibroblasts during colorectal tumour development. We performed a comparative gene expression profiling by whole genome RNA-sequencing of fibroblast subpopulations (COLVI+ vs COLVI-) on STAT3 activation (IL-6 vs IL-11). RESULTS: The analysis of pSTAT3 expression in CAFs of human TMAs revealed a negative correlation of increased stromal pSTAT3 expression with the survival of colon cancer patients. In the loss-of-function and gain-of-function approach, we found a critical role of STAT3 activation in fibroblasts in driving colorectal tumourigenesis in vivo. With different imaging technologies, we detected an expansion of activated fibroblasts in colorectal neoplasias. Comparative gene expression profiling of fibroblast subpopulations on STAT3 activation revealed the regulation of transcriptional patterns associated with angiogenesis. Finally, the blockade of proangiogenic signalling significantly reduced colorectal tumour growth in mice with constitutive STAT3 activation in COLVI+ fibroblasts. CONCLUSION: Altogether our work demonstrates a critical role of STAT3 activation in CAFs in CRC development.

4.
Cancers (Basel) ; 11(11)2019 Nov 05.
Artigo em Inglês | MEDLINE | ID: mdl-31694340

RESUMO

All ligands of the epidermal growth factor receptor (EGF-R) are transmembrane proteins, which need to be proteolytically cleaved in order to be systemically active. The major protease responsible for this cleavage is the membrane metalloprotease ADAM17, which also has been implicated in cleavage of TNFα and interleukin-6 (IL-6) receptor. It has been recently shown that in the absence of ADAM17, the main protease for EGF-R ligand processing, colon cancer formation is largely abrogated. Intriguingly, colon cancer formation depends on EGF-R activity on myeloid cells rather than on intestinal epithelial cells. A major activity of EGF-R on myeloid cells is the stimulation of IL-6 synthesis. Subsequently, IL-6 together with the ADAM17 shed soluble IL-6 receptor acts on intestinal epithelial cells via IL-6 trans-signaling to induce colon cancer formation, which can be blocked by the inhibitor of IL-6 trans-signaling, sgp130Fc. Blockade of IL-6 trans-signaling therefore offers a new therapeutic window downstream of the EGF-R for the treatment of colon cancer and possibly of other EGF-R related neoplastic diseases.

5.
Cells ; 8(10)2019 Oct 09.
Artigo em Inglês | MEDLINE | ID: mdl-31601007

RESUMO

Fibrosis in the liver is mainly associated with the activation of hepatic stellate cells (HSCs). Both activation and clearance of HSCs can be mediated by ligand-receptor interactions. Members of the a disintegrin and metalloprotease (ADAM) family are involved in the proteolytic release of membrane-bound ligands and receptor ectodomains and the remodelling of the extracellular matrix. ADAM proteases are therefore major regulators of intercellular signalling pathways. In the present review we discuss how ADAM proteases modulate pro- and anti-fibrotic processes and how ADAM proteases might be harnessed therapeutically in the future.

6.
Blood ; 134(23): 2092-2106, 2019 Dec 05.
Artigo em Inglês | MEDLINE | ID: mdl-31578204

RESUMO

Graft-versus-host disease (GVHD) after allogeneic stem cell transplantation (alloSCT) is characterized by interleukin-6 (IL-6) dysregulation. IL-6 can mediate effects via various pathways, including classical, trans, and cluster signaling. Given the recent availability of agents that differentially inhibit these discrete signaling cascades, understanding the source and signaling and cellular targets of this cytokine is paramount to inform the design of clinical studies. Here we demonstrate that IL-6 secretion from recipient dendritic cells (DCs) initiates the systemic dysregulation of this cytokine. Inhibition of DC-driven classical signaling after targeted IL-6 receptor (IL-6R) deletion in T cells eliminated pathogenic donor Th17/Th22 cell differentiation and resulted in long-term survival. After engraftment, donor DCs assume the same role, maintaining classical IL-6 signaling-dependent GVHD responses. Surprisingly, cluster signaling was not active after transplantation, whereas inhibition of trans signaling with soluble gp130Fc promoted severe, chronic cutaneous GVHD. The latter was a result of exaggerated polyfunctional Th22-cell expansion that was reversed by IL-22 deletion or IL-6R inhibition. Importantly, inhibition of IL-6 classical signaling did not impair the graft-versus-leukemia effect. Together, these data highlight IL-6 classical signaling and downstream Th17/Th22 differentiation as important therapeutic targets after alloSCT.

7.
Nature ; 574(7776): 63-68, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31554967

RESUMO

The gp130 receptor cytokines IL-6 and CNTF improve metabolic homeostasis but have limited therapeutic use for the treatment of type 2 diabetes. Accordingly, we engineered the gp130 ligand IC7Fc, in which one gp130-binding site is removed from IL-6 and replaced with the LIF-receptor-binding site from CNTF, fused with the Fc domain of immunoglobulin G, creating a cytokine with CNTF-like, but IL-6-receptor-dependent, signalling. Here we show that IC7Fc improves glucose tolerance and hyperglycaemia and prevents weight gain and liver steatosis in mice. In addition, IC7Fc either increases, or prevents the loss of, skeletal muscle mass by activation of the transcriptional regulator YAP1. In human-cell-based assays, and in non-human primates, IC7Fc treatment results in no signs of inflammation or immunogenicity. Thus, IC7Fc is a realistic next-generation biological agent for the treatment of type 2 diabetes and muscle atrophy, disorders that are currently pandemic.


Assuntos
Receptor gp130 de Citocina/metabolismo , Citocinas/síntese química , Citocinas/uso terapêutico , Diabetes Mellitus Tipo 2/tratamento farmacológico , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Ligação Competitiva , Citocinas/química , Diabetes Mellitus Tipo 2/metabolismo , Desenho de Drogas , Fígado Gorduroso/prevenção & controle , Teste de Tolerância a Glucose , Humanos , Hiperglicemia/tratamento farmacológico , Hiperglicemia/metabolismo , Incretinas/metabolismo , Interleucina-6/antagonistas & inibidores , Interleucina-6/metabolismo , Masculino , Camundongos , Músculo Esquelético/efeitos dos fármacos , Obesidade/metabolismo , Pâncreas/metabolismo , Fosfoproteínas/metabolismo , Engenharia de Proteínas , Receptores de Interleucina-6/metabolismo , Transdução de Sinais , Fatores de Transcrição , Ganho de Peso/efeitos dos fármacos
8.
Brain Behav Immun ; 82: 145-159, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31401302

RESUMO

Alzheimer's disease (AD) is the most commonly diagnosed dementia but its underlying pathological mechanisms still unclear. Neuroinflammation and secretion of cytokines such as interleukin-6 (IL-6) accompany the main hallmarks of the disease: amyloid plaques and neurofibrillary tangles. In this study, we analyzed the role of IL-6 trans-signaling in two mouse models of AD, Tg2576 and 3xTg-AD mice. The inhibition of IL-6 trans-signaling partially rescued the AD-induced mortality in females of both models. Before amyloid plaques deposition, it reversed AD-induced changes in exploration and anxiety (but did not affect locomotion) in Tg2576 female mice. However, after plaque deposition the only behavioral trait affected by the inhibition of IL-6 trans-signaling was locomotion. Results in the Morris water maze suggest that cognitive flexibility was reduced by the blocking of the IL-6 trans-signaling in young and old Tg2576 female mice. The inhibition of IL-6 trans-signaling also decreased amyloid plaque burden in cortex and hippocampus, and Aß40 and Aß42 levels in the cortex, of Tg2576 female mice. The aforementioned changes might be correlated with changes in blood vessels and matrix structure and organization rather than changes in neuroinflammation. 3xTgAD mice showed a very mild phenotype regarding amyloid cascade, but results were in accordance with those of Tg2576 mice. These results strongly suggest that the inhibition of the IL-6 trans-signaling could represent a powerful therapeutic target in AD.

9.
Front Aging Neurosci ; 11: 182, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31396076

RESUMO

These days, the important role of retinoids in adult brain functionality and homeostasis is well accepted and has been proven by genomic as well as non-genomic mechanisms. In the healthy brain, numerous biological processes, e.g., cell proliferation, neurogenesis, dendritic spine formation as well as modulation of the immune system, have been attributed to retinoid signaling. This, together with the finding that retinoid metabolism is impaired in Alzheimer's disease (AD), led to preclinical and early clinical testing of natural and synthetic retinoids as innovative pharmaceuticals with multifactorial properties. Acitretin, an aromatic retinoid, was found to exert an anti-amyloidogenic effect in mouse models for AD as well as in human patients by stimulating the alpha-secretase ADAM10. The lipophilic drug was already demonstrated to easily pass the blood brain barrier after i.p. administration and evoked increased nest building capability in the 5xFAD mouse model. Additionally, we analyzed the immune-modulatory capacity of acitretin via a multiplex array in the 5xFAD mouse model and evaluated some of our findings in human CSF derived from a pilot study using acitretin. Although several serum analytes did not display changes, Interleukin-6 (IL-6) was found to be significantly increased in both-mouse and human neural material. This demonstrates that acitretin exerts an immune stimulatory effect-besides the alpha-secretase induction-which could impact the alleviation of learning and memory disabilities observed in the mouse model.

10.
Cancers (Basel) ; 11(9)2019 Aug 21.
Artigo em Inglês | MEDLINE | ID: mdl-31438559

RESUMO

Lung cancer is the leading cause of cancer-related mortality, which histologically is classified into small-cell lung cancer (SCLC) and non-small cell lung cancer (NSCLC). NSCLC accounts for approximately 85% of all lung cancer diagnoses, with the majority of patients presenting with lung adenocarcinoma (LAC). KRAS mutations are a major driver of LAC, and are closely related to cigarette smoking, unlike mutations in the epidermal growth factor receptor (EGFR) which arise in never-smokers. Although the past two decades have seen fundamental progress in the treatment and diagnosis of NSCLC, NSCLC still is predominantly diagnosed at an advanced stage when therapeutic interventions are mostly palliative. A disintegrin and metalloproteinase 17 (ADAM17), also known as tumour necrosis factor-α (TNFα)-converting enzyme (TACE), is responsible for the protease-driven shedding of more than 70 membrane-tethered cytokines, growth factors and cell surface receptors. Among these, the soluble interleukin-6 receptor (sIL-6R), which drives pro-inflammatory and pro-tumourigenic IL-6 trans-signaling, along with several EGFR family ligands, are the best characterised. This large repertoire of substrates processed by ADAM17 places it as a pivotal orchestrator of a myriad of physiological and pathological processes associated with the initiation and/or progression of cancer, such as cell proliferation, survival, regeneration, differentiation and inflammation. In this review, we discuss recent research implicating ADAM17 as a key player in the development of LAC, and highlight the potential of ADAM17 inhibition as a promising therapeutic strategy to tackle this deadly malignancy.

11.
JCI Insight ; 4(15)2019 08 08.
Artigo em Inglês | MEDLINE | ID: mdl-31391340

RESUMO

Aberrant activity of the glycoprotein 130 130/JAK/STAT3 (gp130/JAK/STAT3) signaling axis is a recurrent event in inflammation and cancer. In particular, it is associated with a wide range of hematological malignancies, including multiple myeloma and leukemia. Novel targeted therapies have only been successful for some subtypes of these malignancies, underlining the need for developing robust mouse models to better dissect the role of this pathway in specific tumorigenic processes. Here, we investigated the role of selective gp130/JAK/STAT3 activation by generating a conditional mouse model. This model targeted constitutively active, cell-autonomous gp130 activity to B cells, as well as to the entire hematopoietic system. We found that regardless of the timing of activation in B cells, constitutively active gp130 signaling resulted in the formation specifically of mature B cell lymphomas and plasma cell disorders with full penetrance, only with different latencies, where infiltrating CD138+ cells were a dominant feature in every tumor. Furthermore, constitutively active gp130 signaling in all adult hematopoietic cells also led to the development specifically of largely mature, aggressive B cell cancers, again with a high penetrance of CD138+ tumors. Importantly, gp130 activity abrogated the differentiation block induced by a B cell-targeted Myc transgene and resulted in a complete penetrance of the gp130-associated, CD138+, mature B cell lymphoma phenotype. Thus, gp130 signaling selectively provides a strong growth and differentiation advantage for mature B cells and directs lymphomagenesis specifically toward terminally differentiated B cell cancers.

12.
FASEB J ; 33(11): 11925-11940, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31381863

RESUMO

Meprin ß is a membrane-bound metalloprotease involved in extracellular matrix assembly and inflammatory processes in health and disease. A disintegrin and metalloproteinase (ADAM)10 and ADAM17 are physiologic relevant sheddases of inactive promeprin ß, which influences its substrate repertoire and subsequent biologic functions. Proteomic analysis also revealed several ADAMs as putative meprin ß substrates. Here, we demonstrate specific N-terminal processing of ADAM9, 10, and 17 by meprin ß and identify cleavage sites within their prodomains. Because ADAM prodomains can act as specific inhibitors, we postulate a role for meprin ß in the regulation of ADAM activities. Indeed, prodomain cleavage by meprin ß caused increased ADAM protease activities, as observed by peptide-based cleavage assays and demonstrated by increased ectodomain shedding activity. Direct interaction of meprin ß and ADAM proteases could be shown by immunofluorescence microscopy and immunoprecipitation experiments. As demonstrated by a bacterial activator of meprin ß and additional measurement of TNF-α shedding on bone marrow-derived macrophages, meprin ß/ADAM protease interactions likely influence inflammatory conditions. Thus, we identified a novel proteolytic pathway of meprin ß with ADAM proteases to control protease activities at the cell surface as part of the protease web.-Wichert, R., Scharfenberg, F., Colmorgen, C., Koudelka, T., Schwarz, J., Wetzel, S., Potempa, B., Potempa, J., Bartsch, J. W., Sagi, I., Tholey, A., Saftig, P., Rose-John, S., Becker-Pauly, C. Meprin ß induces activities of A disintegrin and metalloproteinases 9, 10, and 17 by specific prodomain cleavage.

13.
Neuroscience ; 414: 280-296, 2019 08 21.
Artigo em Inglês | MEDLINE | ID: mdl-31301368

RESUMO

The role of the pro-inflammatory cytokine interleukin-6 (IL-6) in the etiology of stress-induced synaptic plasticity is yet unknown. We took advantage of a genetically modified mouse (TG) in which IL-6 trans-signaling via the soluble IL-6 receptor was blocked, to determine the role of IL-6 trans-signaling in the effects of a Social Defeat protocol (SD) on synaptic function of the medial prefrontal cortex (mPFC). Synaptic function in stress-sensitive (S) and stress-resilient (R) animals was studied in a mPFC slice preparation with whole-cell patch-clamp recording. SD altered numerous synaptic properties of the mPFC: R WT (but not TG) displayed a decreased ratio between N methyl-D-aspartate receptor (NMDAR-) dependent and amino propionic acid receptor (AMPAR-) dependent-current (INMDA/IAMPA), while S WT animals (but not TG) showed a reduced ratio between AMPA and γ-amino-butyric acid receptor type A (GABAAR)-dependent currents (IAMPA/IGABA). Also, SD induced an increase in the frequency but a decrease in the amplitude of excitatory action-potential dependent PSCs (sEPSCs), both in an IL-6 dependent manner, as well as a generalized (S/R-independent) decrease in the frequency of action potential independent (miniature) excitatory (IL-6 dependent) as well as inhibitory (IL-6 independent) postsynaptic current frequency. Interestingly, corner preference (measuring the intensity of social defeat) correlated positively with INMDA/IAMPA and eEPSC frequency and negatively with IAMPA/IGABA. Our results suggest that SD induces behaviorally-relevant synaptic rearrangement in mPFC circuits, part of which is IL-6 dependent. In particular, IL-6 is necessary to produce synaptic plasticity leading to stress resilience in some individuals, but to stress sensitivity in others.

14.
Carcinogenesis ; 2019 Jun 27.
Artigo em Inglês | MEDLINE | ID: mdl-31257400

RESUMO

Lung cancer is the leading cause of cancer-related mortality, with most cases attributed to tobacco smoking, in which nicotine-derived nitrosamine ketone (NNK) is the most potent lung carcinogen. The ADAM17 protease is responsible for the ectodomain shedding of many pro-tumourigenic cytokines, growth factors and receptors, and therefore is an attractive target in cancer. However, the role of ADAM17 in promoting tobacco smoke carcinogen-induced lung carcinogenesis is unknown. The hypomorphic Adam17ex/ex mice - characterized by reduced global ADAM17 expression - were backcrossed onto the NNK-sensitive pseudo-A/J background. CRISPR-driven and inhibitor-based (GW280264X, and ADAM17 prodomain) ADAM17 targeting was employed in the human lung adenocarcinoma cell lines A549 and NCI-H23. Human lung cancer biopsies were also used for analyses. The Adam17ex/ex mice displayed marked protection against NNK-induced lung adenocarcinoma. Specifically, the number and size of lung lesions in NNK-treated pseudo-A/J Adam17ex/ex mice were significantly reduced compared to wild-type littermate controls. This was associated with lower proliferative index throughout the lung epithelium. ADAM17 targeting in A549 and NCI-H23 cells led to reduced proliferative and colony-forming capacities. Notably, among select ADAM17 substrates, ADAM17 deficiency abrogated shedding of the soluble IL-6 receptor (sIL-6R), which coincided with the blockade of sIL-6R-mediated trans-signaling via ERK MAPK cascade. Furthermore, NNK upregulated phosphorylation of p38 MAPK, whose pharmacological inhibition suppressed ADAM17 threonine phosphorylation. Importantly, ADAM17 threonine phosphorylation was significantly upregulated in human lung adenocarcinoma with smoking history compared to their cancer-free controls. Our study identifies the ADAM17/sIL-6R/ERK MAPK axis as a candidate therapeutic strategy against tobacco smoke associated lung carcinogenesis.

15.
Int J Mol Sci ; 20(15)2019 Jul 26.
Artigo em Inglês | MEDLINE | ID: mdl-31357561

RESUMO

Interleukin-11 (IL-11) has been associated with inflammatory conditions, bone homeostasis, hematopoiesis, and fertility. So far, these functions have been linked to classical IL-11 signaling via the membrane bound receptor (IL-11R). However, a signaling cascade via the soluble IL-11R (sIL-11R), generated by proteolytic cleavage, can also be induced. This process is called IL-11 trans-signaling. A disintegrin and metalloprotease 10 (ADAM10) and neutrophil elastase were described as ectodomain sheddases of the IL-11R, thereby inducing trans-signaling. Furthermore, previous studies employing approaches for the stimulation and inhibition of endogenous ADAM-proteases indicated that ADAM10, but not ADAM17, can cleave the IL-11R. Herein, we show that several metalloproteases, namely ADAM9, ADAM10, ADAM17, meprin ß, and membrane-type 1 matrix metalloprotease/matrix metalloprotease-14 (MT1-MMP/MMP-14) when overexpressed are able to shed the IL-11R. All sIL-11R ectodomains were biologically active and capable of inducing signal transducer and activator of transcription 3 (STAT3) phosphorylation in target cells. The difference observed for ADAM10/17 specificity compared to previous studies can be explained by the different approaches used, such as stimulation of protease activity or making use of cells with genetically deleted enzymes.


Assuntos
Proteínas ADAM/metabolismo , Metaloproteinase 14 da Matriz/metabolismo , Receptores de Interleucina-11/metabolismo , Tiopronina/metabolismo , Proteínas ADAM/química , Humanos , Metaloproteinase 14 da Matriz/química , Fosforilação , Proteólise , Receptores de Interleucina-11/química , Receptores de Interleucina-6/metabolismo , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais , Relação Estrutura-Atividade
16.
Am J Physiol Endocrinol Metab ; 317(2): E411-E420, 2019 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-31237452

RESUMO

Serum levels of interleukin-6 (IL-6) are increased in patients with type 2 diabetes (T2D). IL-6 exerts its pleiotropic effects via the IL-6 α-receptor (IL-6R), which exists in membrane-bound and soluble (sIL-6R) forms and activates cells via the ß-receptor glycoprotein 130 (gp130). The nonsynonymous single-nucleotide polymorphism (SNP) rs2228145 (Asp358Ala) within the IL6R locus is associated with T2D. The aim of this study was to determine whether sIL-6R in combination with soluble gp130 (sgp130) is able to form an IL-6-neutralizing buffer in healthy subjects and whether this is disturbed in T2D. We found that sIL-6R-sgp130 indeed forms an IL-6-neutralizing buffer in the serum of healthy humans, whose capacity is controlled by the SNP of the IL-6R. Circulating sIL-6R-sgp130 levels were lower in T2D subjects (P < 0.001), whereas IL-6 was high and inversely correlated with sIL-6R (r = -0.57, P < 0.001), indicating a severe disturbance of the buffer. This phenomenon is also observed in sex- and age-matched patients with both T2D and atherosclerosis but not in patients with atherosclerosis alone. In conclusion, sIL-6R and sgp130 serum levels were significantly lower in T2D patients compared with healthy subjects or atherosclerosis patients, although IL-6 levels were high. These data suggest that disturbance of the protective buffer may be closely associated with T2D pathophysiology.

17.
Int J Mol Sci ; 20(9)2019 May 04.
Artigo em Inglês | MEDLINE | ID: mdl-31060243

RESUMO

Colorectal cancer is one of the most commonly diagnosed malignancies in the Western world and is associated with elevated expression and activity of epidermal growth factor receptors (EGF-R). The metalloproteinase ADAM17 is involved in EGF-R activation by processing EGF-R ligands from membrane-bound pro-ligands. Underlining the link between colon cancer and ADAM17, genetic intestinal cancer models in ADAM17-deficient mice show a reduced tumor burden. In this study, we characterize point mutations within the ADAM17 gene found in the tissue of colon cancer patients. In order to shed light on the role of ADAM17 in cancer development, as well as into the mechanisms that regulate maturation and cellular trafficking of ADAM17, we here perform overexpression studies of four ADAM17 variants located in the pro-, membrane-proximal- and cytoplasmic-domain of the ADAM17 protein in ADAM10/17-deficient HEK cells. Interestingly, we found a cancer-associated point mutation within the pro-domain of ADAM17 (R177C) to be most impaired in its proteolytic activity and trafficking to the cell membrane. By comparing this variant to an ADAM17 construct lacking the entire pro-domain, we discovered similar functional limitations and propose a crucial role of the pro-domain for ADAM17 maturation, cellular trafficking and thus proteolytic activity.


Assuntos
Proteína ADAM17/genética , Proteína ADAM17/metabolismo , Neoplasias do Colo/genética , Neoplasias do Colo/metabolismo , Mutação , Proteína ADAM17/química , Animais , Biomarcadores , Neoplasias do Colo/patologia , Suscetibilidade a Doenças , Humanos , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Transporte Proteico , Proteólise , Relação Estrutura-Atividade
18.
Mol Ther Nucleic Acids ; 16: 419-433, 2019 Jun 07.
Artigo em Inglês | MEDLINE | ID: mdl-31026677

RESUMO

Interleukin-6 (IL-6)-type cytokines share the common receptor glycoprotein 130 (gp130), which activates a signaling cascade involving Janus kinases (JAKs) and signal transducer and activator of transcription (STAT) transcription factors. IL-6 and/or its signaling pathway is often deregulated in diseases, such as chronic liver diseases and cancer. Thus, the identification of compounds inhibiting this pathway is of interest for future targeted therapies. We established novel cellular screening systems based on a STAT-responsive reporter gene (Cypridina luciferase). Of a library containing 538 microRNA (miRNA) mimics, several miRNAs affected hyper-IL-6-induced luciferase activities. When focusing on candidate miRNAs specifically targeting 3' UTRs of signaling molecules of this pathway, we identified, e.g., miR-3677-5p as a novel miRNA affecting protein expression of both STAT3 and JAK1, whereas miR-16-1-3p, miR-4473, and miR-520f-3p reduced gp130 surface expression. Interestingly, combination treatment with 2 or 3 miRNAs targeting gp130 or different signaling molecules of the pathway did not increase the inhibitory effects on phospho-STAT3 levels and STAT3 target gene expression compared to treatment with single mimics. Taken together, we identified a set of miRNAs of potential therapeutic value for cancer and inflammatory diseases, which directly target the expression of molecules within the IL-6-signaling pathway and can dampen inflammatory signal transduction.

19.
Brain Behav Immun ; 80: 129-145, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-30851378

RESUMO

A disintegrin and metalloproteinase 17 (ADAM17) is the major sheddase involved in the cleavage of a plethora of cytokines, cytokine receptors and growth factors, thereby playing a substantial role in inflammatory and regenerative processes after central nervous system trauma. By making use of a hypomorphic ADAM17 knockin mouse model as well as pharmacological ADAM10/ADAM17 inhibitors, we showed that ADAM17-deficiency or inhibition significantly increases clearance of apoptotic cells, promotes axon growth and improves functional recovery after spinal cord injury (SCI) in mice. Microglia-specific ADAM17-knockout (ADAM17flox+/+-Cx3Cr1 Cre+/-) mice also showed improved functional recovery similar to hypomorphic ADAM17 mice. In contrast, endothelial-specific (ADAM17flox+/+-Cdh5Pacs Cre+/-) and macrophage-specific (ADAM17flox+/+-LysM Cre+/-) ADAM17-knockout mice or bone marrow chimera with transplanted ADAM17-deficient macrophages, displayed no functional improvement compared to wild type mice. These data indicate that ADAM17 expression on microglia cells (and not on macrophages or endothelial cells) plays a detrimental role in inflammation and functional recovery after SCI.

20.
EMBO Mol Med ; 11(4)2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30833304

RESUMO

Oncogenic KRAS mutations are major drivers of lung adenocarcinoma (LAC), yet the direct therapeutic targeting of KRAS has been problematic. Here, we reveal an obligate requirement by oncogenic KRAS for the ADAM17 protease in LAC In genetically engineered and xenograft (human cell line and patient-derived) Kras G12D-driven LAC models, the specific blockade of ADAM17, including with a non-toxic prodomain inhibitor, suppressed tumor burden by reducing cellular proliferation. The pro-tumorigenic activity of ADAM17 was dependent upon its threonine phosphorylation by p38 MAPK, along with the preferential shedding of the ADAM17 substrate, IL-6R, to release soluble IL-6R that drives IL-6 trans-signaling via the ERK1/2 MAPK pathway. The requirement for ADAM17 in Kras G12D-driven LAC was independent of bone marrow-derived immune cells. Furthermore, in KRAS mutant human LAC, there was a significant positive correlation between augmented phospho-ADAM17 levels, observed primarily in epithelial rather than immune cells, and activation of ERK and p38 MAPK pathways. Collectively, these findings identify ADAM17 as a druggable target for oncogenic KRAS-driven LAC and provide the rationale to employ ADAM17-based therapeutic strategies for targeting KRAS mutant cancers.

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