RESUMO
IGF1R plays an important role in regulating cellular metabolism and cell growth, and has been identified as an anti-cancer and diabetes drug target. Although research have been reported many crystal and cryo-EM structures of IGF1R, the mechanism of ligand binding remains controversial, mainly because the structure differences among its cryo-EM, crystal and homologous protein insulin receptor structures. Here, we further determined one new high-resolution symmetric cryo-EM structure of ligand-bound IGF1R and be the first to prove that the receptor could bind to two IGFI molecules by single particle cryo-electron microscopy. And the structure is very different from its homologous protein insulin receptor: the two ligands just exist at the binding site 2 with saturating ligand conditions. Then, our findings resolved the major dispute about the comformational changes of IGF1R, and proposed a new theory how IGF1R binds to its ligands. Meanwhile, these findings imply more attention may be needed to study the relationship between the special conformation and their corresponding physiological functions in future.
Assuntos
Fator de Crescimento Insulin-Like I , Receptor IGF Tipo 1 , Humanos , Microscopia Crioeletrônica , Hormônios , Fator de Crescimento Insulin-Like I/química , Ligantes , Domínios Proteicos , Receptor IGF Tipo 1/química , Receptor de Insulina/químicaRESUMO
The human VPS10 domain-containing receptor SorCS3 belongs to the Vps10p-domain receptor family and is an important receptor for regulating normal cellular functions via protein sorting. Here, we determined the cryo-EM structure of the full-length SorCS3 receptor and further found that there were at least three distinct conformations (monomer, M-shaped dimer and N-shaped dimer) of SorCS3 in the apo state. The differences between the two dimer conformations were caused by PKD1-2 assembly. In contrast to its homologous proteins, the conserved residues GLN198, ARG678, TYR430, GLU1020 and ASP1024 may be key points for its dimerization and for protein/polypeptide binding. These results showed the structural details of apo-SorCS3, which provides a foundation for elucidating the mechanism of protein sorting.
Assuntos
Proteínas de Transporte , Proteínas do Tecido Nervoso , Proteínas de Transporte/metabolismo , Microscopia Crioeletrônica , Humanos , Proteínas do Tecido Nervoso/metabolismo , Ligação Proteica , Transporte Proteico , Receptores de Superfície Celular/metabolismoRESUMO
PURPOSE: Mesenchymal stromal cells (MSC) are an attractive tool for treatment of diabetic cardiomyopathy. Syndecan-2/CD362 has been identified as a functional marker for MSC isolation. Imaging mass spectrometry (IMS) allows for the characterization of therapeutic responses in the left ventricle. This study aims to investigate whether IMS can assess the therapeutic effect of CD362+ -selected MSC on early onset experimental diabetic cardiomyopathy. EXPERIMENTAL DESIGN: 1 × 106 wild type (WT), CD362- , or CD362+ MSC are intravenously injected into db/db mice. Four weeks later, mice are hemodynamically characterized and subsequently sacrificed for IMS combined with bottom-up mass spectrometry, and isoform and phosphorylation analyses of cardiac titin. RESULTS: Overall alterations of the cardiac proteome signatures, especially titin, are observed in db/db compared to control mice. Interestingly, only CD362+ MSC can overcome the reduced titin intensity distribution and shifts the isoform ratio toward the more compliant N2BA form. In contrast, WT and CD362- MSCs improve all-titin phosphorylation and protein kinase G activity, which is reflected in an improvement in diastolic performance. CONCLUSIONS AND CLINICAL RELEVANCE: IMS enables the characterization of differences in titin intensity distribution following MSC application. However, further analysis of titin phosphorylation is needed to allow for the assessment of the therapeutic efficacy of MSC.
Assuntos
Cardiomiopatias Diabéticas/patologia , Células-Tronco Mesenquimais , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Animais , Humanos , CamundongosRESUMO
OBJECTIVE: To select potential molecules that can target viral spike proteins, which may potentially interrupt the interaction between the human angiotension-converting enzyme 2 (ACE2) receptor and viral spike protein by virtual screening. METHODS: The three-dimensional (3D)-coordinate file of the receptor-binding domain (RBD)-ACE2 complex for searching a suitable docking pocket was firstly downloaded and prepared. Secondly, approximately 15,000 molecular candidates were prepared, including US Food and Drug Administration (FDA)-approved drugs from DrugBank and natural compounds from Traditional Chinese Medicine Systems Pharmacology (TCMSP), for the docking process. Then, virtual screening was performed and the binding energy in Autodock Vina was calculated. Finally, the top 20 molecules with high binding energy and their Chinese medicine (CM) herb sources were listed in this paper. RESULTS: It was found that digitoxin, a cardiac glycoside in DrugBank and bisindigotin in TCMSP had the highest docking scores. Interestingly, two of the CM herbs containing the natural compounds that had relatively high binding scores, Forsythiae fructus and Isatidis radix, are components of Lianhua Qingwen (), a CM formula reportedly exerting activity against severe acute respiratory syndrome (SARS)-Cov-2. Moreover, raltegravir, an HIV integrase inhibitor, was found to have a relatively high binding score. CONCLUSIONS: A class of compounds, which are from FDA-approved drugs and CM natural compounds, that had high binding energy with RBD of the viral spike protein. Our work provides potential candidates for other researchers to identify inhibitors to prevent SARS-CoV-2 infection, and highlights the importance of CM and integrative application of CM and Western medicine on treating COVID-19.
Assuntos
Infecções por Coronavirus/tratamento farmacológico , Reposicionamento de Medicamentos/métodos , Medicamentos de Ervas Chinesas/farmacologia , Glicoproteínas/efeitos dos fármacos , Imageamento Tridimensional , Simulação de Acoplamento Molecular/métodos , Pneumonia Viral/tratamento farmacológico , COVID-19 , China , Simulação por Computador , Infecções por Coronavirus/diagnóstico , Glicoproteínas/metabolismo , Humanos , Programas de Rastreamento/métodos , Pandemias , Peptidil Dipeptidase A/efeitos dos fármacos , Pneumonia Viral/diagnóstico , Ligação Proteica , Estados Unidos , United States Food and Drug AdministrationRESUMO
BACKGROUND MicroRNAs (miRNAs), which modulate the expression of their target genes, are commonly involved in stimulating and adjusting of many processes that result in cardiovascular diseases, contain cardiac ischemia/reperfusion (I/R) damage. However, the expression and role of miR-149 in pyroptosis mediated myocardial I/R damage remains unclear. MATERIAL AND METHODS Real-time polymerase chain reaction was performed to measure the miR-149 and FoxO3 expression in I/R stimulated H9C2 cells. The cell proliferation, pyroptosis-related inflammatory genes in I/R-treated H9C2 cells transfected miR-149 mimics or miR-149 inhibitor were both explored. We predicted and confirmed miR-149 targets by using bioinformatics analyses and luciferase reporter assay. In addition, the potential relationship between miR-149 and FoxO3 in pyroptosis from I/R treated H9C2 cells was analyzed. RESULTS Our results showed that miR-149 was upregulated, while FoxO3 was downregulated in I/R stimulated H9C2 cells. Over-expression of miR-149 inhibited cell viability and promote pyroptosis, however, down-expression of miR-149 had an opposite effect in I/R treated H9C2 cells. Furthermore, miR-149 could negatively regulate FoxO3 expression by binding 3'UTR, whereas silencing of FoxO3 attenuated the effect of miR-149-mimics on cell proliferation and pyroptosis in I/R treated H9C2 cells. CONCLUSIONS Our study found that miR-149 played a critical role in pyroptosis during cardiac I/R injury, and thus, might provide a novel therapeutic target.
Assuntos
Proteína Forkhead Box O3/genética , MicroRNAs/genética , Piroptose/genética , Animais , Linhagem Celular , Sobrevivência Celular , Proteína Forkhead Box O3/metabolismo , Isquemia/metabolismo , Camundongos , Isquemia Miocárdica/genética , Traumatismo por Reperfusão Miocárdica/metabolismo , Miócitos Cardíacos/metabolismo , Piroptose/fisiologia , Traumatismo por Reperfusão/genética , Traumatismo por Reperfusão/metabolismo , Transdução de SinaisRESUMO
Inflammation in myocarditis induces cardiac injury and triggers disease progression to heart failure. NLRP3 inflammasome activation is a newly identified amplifying step in the pathogenesis of myocarditis. We previously have demonstrated that mesenchymal stromal cells (MSC) are cardioprotective in Coxsackievirus B3 (CVB3)-induced myocarditis. In this study, MSC markedly inhibited left ventricular (LV) NOD2, NLRP3, ASC, caspase-1, IL-1ß, and IL-18 mRNA expression in CVB3-infected mice. ASC protein expression, essential for NLRP3 inflammasome assembly, increased upon CVB3 infection and was abrogated in MSC-treated mice. Concomitantly, CVB3 infection in vitro induced NOD2 expression, NLRP3 inflammasome activation and IL-1ß secretion in HL-1 cells, which was abolished after MSC supplementation. The inhibitory effect of MSC on NLRP3 inflammasome activity in HL-1 cells was partly mediated via secretion of the anti-oxidative protein stanniocalcin-1. Furthermore, MSC application in CVB3-infected mice reduced the percentage of NOD2-, ASC-, p10- and/or IL-1ß-positive splenic macrophages, natural killer cells, and dendritic cells. The suppressive effect of MSC on inflammasome activation was associated with normalized expression of prominent regulators of myocardial contractility and fibrosis to levels comparable to control mice. In conclusion, MSC treatment in myocarditis could be a promising strategy limiting the adverse consequences of cardiac and systemic NLRP3 inflammasome activation.
Assuntos
Inflamassomos/metabolismo , Células-Tronco Mesenquimais/fisiologia , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Animais , Cardiomiopatias/metabolismo , Caspase 1/metabolismo , Infecções por Coxsackievirus/virologia , Coração/fisiologia , Humanos , Inflamação/patologia , Interleucina-1beta/metabolismo , Macrófagos/metabolismo , Masculino , Células-Tronco Mesenquimais/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Miocardite/virologia , Miocárdio/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/fisiologiaRESUMO
A new microporous lanthanide metal-organic framework, {[Yb(BTB)(H2O) (DEF)2}n (1, DEF=N,N-Diethylformamide), with 1D nano-sized channels has been constructed by bridging helical chain secondary building units with 1,3,5-benzenetrisbenzoic acid (H3BTB) ligand. Structural characterization suggests that this complex crystallizes in the hexagonal space group P6122 and possesses 1D triangular channels with coordinated water molecules pointing to the channel center. In addition, anti-myocarditis properties of compound 1 were evaluated in vivo. The results showed that compound 1 can improve hemodynamic parameters of, and it may be a good therapeutic option for heart failure in the future.
Assuntos
Animais , Masculino , Camundongos , Anti-Inflamatórios/química , Cristalografia por Raios X , Elementos da Série dos Lantanídeos/química , Estruturas Metalorgânicas/química , Miocardite/terapia , Anti-Inflamatórios/uso terapêutico , Estruturas Metalorgânicas/uso terapêutico , Modelos Moleculares , Difração de Pó , Termogravimetria , Difração de Raios XRESUMO
Mesenchymal stromal cell (MSC) application in Coxsackievirus B3 (CVB3)-induced myocarditis reduces myocardial inflammation and fibrosis, exerts prominent extra-cardiac immunomodulation, and improves heart function. Although the abovementioned findings demonstrate the benefit of MSC application, the mechanism of the MSC immunomodulatory effects leading to a final cardioprotective outcome in viral myocarditis remains poorly understood. Monocytes are known to be a trigger of myocardial tissue inflammation. The present study aims at investigating the direct effect of MSC on the mobilization and trafficking of monocytes to the heart in CVB3-induced myocarditis. One day post CVB3 infection, C57BL/6 mice were intravenously injected with 1 x 106 MSC and sacrificed 6 days later for molecular biology and flow cytometry analysis. MSC application reduced the severity of myocarditis, and heart and blood pro-inflammatory Ly6Chigh and Ly6Cmiddle monocytes, while those were retained in the spleen. Anti-inflammatory Ly6Clow monocytes increased in the blood, heart, and spleen of MSC-treated CVB3 mice. CVB3 infection induced splenic myelopoiesis, while MSC application slightly diminished the spleen myelopoietic activity in CVB3 mice. Left ventricular (LV) mRNA expression of the chemokines monocyte chemotactic protein-1 (MCP)-1, MCP-3, CCL5, the adhesion molecules intercellular adhesion molecule-1, vascular cell adhesion molecule-1, the pro-inflammatory cytokines interleukin-6, interleukin-12, tumor necrosis factor-α, the pro-fibrotic transforming growth factorß1, and circulating MCP-1 and MCP-3 levels decreased in CVB3 MSC mice, while LV stromal cell-derived factor-1α RNA expression and systemic levels of fractalkine were increased in CVB3 MSC mice. MSC application in CVB3-induced myocarditis modulates monocytes trafficking to the heart and could be a promising strategy for the resolution of cardiac inflammation and prevention of the disease progression. Stem Cells Translational Medicine 2017;6:1249-1261.
Assuntos
Infecções por Coxsackievirus/complicações , Células-Tronco Mesenquimais/fisiologia , Miocardite/etiologia , Miocardite/terapia , Miocárdio/citologia , Animais , Células Cultivadas , Quimiocina CCL2/metabolismo , Quimiocina CCL7/metabolismo , Humanos , Interleucina-12/metabolismo , Interleucina-6/metabolismo , Masculino , Células-Tronco Mesenquimais/citologia , Camundongos , Camundongos Endogâmicos C57BL , Monócitos/citologia , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Rising worldwide cancer incidence and resistance to current anti-cancer drugs necessitate the need for new pharmaceutical compounds and drug delivery system. Two novel series of biscoumarin (1-4) and dihydropyran (5-16) derivatives were synthesized via a one-pot multicomponent condensation reaction and evaluated for their antitumor activity in vitro. The X-ray crystal structure analysis of four representative compounds 2, 7, 10 and 13 confirmed the structures of these compounds. Compounds 1-4 showed the most potent antitumor activity among the total 16 derivatives. More interestingly, preliminary mechanism studies revealed that the most potent compound 4 induced apoptosis and arrested the cell cycle at the S phase in HUTU80 cells. Additionally, the increased accumulation of HUTU80 cells in the sub G1 peak further pointed to the occurence of the cell apoptosis. The selectivity index analysis demonstrated that all the biscoumarin compounds (SI=3.1-7.5) possess higher selectivity towards intestinal epithelial adenocarcinoma cell line (HuTu80) than positive control drug carboplatin (SI=1.6-1.8). The biscoumarin compounds also showed no obvious acute toxicity on mice.
Assuntos
Antineoplásicos/química , Cumarínicos/química , Piranos/química , Antineoplásicos/síntese química , Antineoplásicos/toxicidade , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Cumarínicos/síntese química , Cumarínicos/toxicidade , Cristalografia por Raios X , Ensaios de Seleção de Medicamentos Antitumorais , Pontos de Checagem da Fase G1 do Ciclo Celular/efeitos dos fármacos , Células HEK293 , Células Endoteliais da Veia Umbilical Humana , Humanos , Rim/efeitos dos fármacos , Rim/patologia , Fígado/efeitos dos fármacos , Fígado/patologia , Conformação Molecular , Piranos/síntese química , Piranos/toxicidade , Relação Estrutura-AtividadeRESUMO
We report three patients with acute dysfunction of mechanical aortic valve prosthesis resulting in chest pain, concomitant cardiogenic shock, and electrocardiographic changes mimicking acute occlusion of the left main coronary artery whereas emergency coronary angiography revealed normal coronary arteries. During operation, abnormal proliferation of subvalvular pannus overgrowth on the inflow aspect of the prosthesis was found to impede normal prosthesis closure. We discuss the possible underlying mechanisms of electrocardiographic presentations.
Assuntos
Valva Aórtica/cirurgia , Oclusão Coronária/diagnóstico , Eletrocardiografia , Doenças das Valvas Cardíacas/diagnóstico , Próteses Valvulares Cardíacas/efeitos adversos , Falha de Prótese/efeitos adversos , Doença Aguda , Idoso , Dor no Peito/etiologia , Diagnóstico Diferencial , Feminino , Doenças das Valvas Cardíacas/cirurgia , Implante de Prótese de Valva Cardíaca , Humanos , Masculino , Pessoa de Meia-IdadeRESUMO
OBJECTIVE: To observe the in vitro growth inhibitory and apoptosis-inducing effects of paclitaxel on the human osteosarcoma cell line U-2 OS. METHODS: U-2 OS cells were treated with various concentrations of paclitaxel. Proliferation was determined by cell count in a neubauer cytometer chamber. Viability was assessed by trypan blue dye exclusion. Paclitaxel-induced morphologic alterations were visualized using light and transmission electron microscopy. The extent of paclitaxel-induced apoptosis was determined by flow cytometry and immunohistochemical detection (TdT-mediated dUTP nick end labeling technique, TUNEL). RESULTS: The cells treated with paclitaxel initially showed G(2)/M arrest, which was followed by apoptosis. The characteristic apoptotic changes, including nuclear disintegration and chromatin agglomeration, were displayed. Large amounts of micronuclei cells appeared, something not observed in those cells treated with cisplatin and adriamycin for contrast. Also, extensive DNA-cleavage was detected using TUNEL. CONCLUSION: This study demonstrates that paclitaxel has an apoptotic-inducing effect on osteosarcoma cells through the initiation of G(2)/M arrest and inhibiting mitosis in both a time-dependent and dose-dependent manner.