RESUMO
The complement system, a key component of innate immunity, provides the first line of defense against bacterial infection; however, the COVID-19 pandemic has revealed that it may also engender severe complications in the context of viral respiratory disease. Here, we review the mechanisms of complement activation and regulation and explore their roles in both protecting against infection and exacerbating disease. We discuss emerging evidence related to complement-targeted therapeutics in COVID-19 and compare the role of the complement in other respiratory viral diseases like influenza and respiratory syncytial virus. We review recent mechanistic studies and animal models that can be used for further investigation. Novel knockout studies are proposed to better understand the nuances of the activation of the complement system in respiratory viral diseases.
Assuntos
COVID-19 , Influenza Humana , Vírus Sincicial Respiratório Humano , Animais , Humanos , Pandemias , Proteínas do Sistema ComplementoRESUMO
Kidney intercalated cells (ICs) maintain acid-base homeostasis and recent studies have demonstrated that they function in the kidney's innate defense. To study kidney innate immune function, ICs have been enriched using vacuolar ATPase (V-ATPase) B1 subunit (Atp6v1b1)-Cre (B1-Cre) mice. Although Atp6v1b1 is considered kidney specific, it is expressed in multiple organ systems, both in mice and humans, raising the possibility of off-target effects when using the Cre-lox system. We have recently shown using single-cell RNA sequencing that the gene that codes for the V-ATPase G3 subunit (mouse gene: Atp6v1g3; human gene: ATP6V1G3; protein abbreviation: G3) mRNA is selectively enriched in human kidney ICs. In this study, we generated Atp6v1g3-Cre (G3-Cre) reporter mice using CRISPR/CAS technology and crossed them with Tdtomatoflox/flox mice. The resultant G3-Cre+Tdt+ progeny was evaluated for kidney specificity in multiple tissues and found to be highly specific to kidney cells with minimal or no expression in other organs evaluated compared with B1-Cre mice. Tdt+ cells were flow sorted and were enriched for IC marker genes on RT-PCR analysis. Next, we crossed these mice to ihCD59 mice to generate an IC depletion mouse model (G3-Cre+ihCD59+/+). ICs were depleted in these mice using intermedilysin, which resulted in lower blood pH, suggestive of a distal renal tubular acidosis phenotype. The G3-Cre mice were healthy, bred normally, and produce regular-sized litter. Thus, this new "IC reporter" mice can be a useful tool to study ICs.NEW & NOTEWORTHY This study details the development, validation, and experimental use of a new mouse model to study the collecting duct and intercalated cells. Kidney intercalated cells are a cell type increasingly recognized to be important in several human diseases including kidney infections, acid-base disorders, and acute kidney injury.
Assuntos
Acidose Tubular Renal , Túbulos Renais Coletores , ATPases Vacuolares Próton-Translocadoras , Camundongos , Humanos , Animais , Rim/metabolismo , Integrases/genética , Integrases/metabolismo , Acidose Tubular Renal/genética , ATPases Vacuolares Próton-Translocadoras/genética , ATPases Vacuolares Próton-Translocadoras/metabolismo , Túbulos Renais Coletores/metabolismoRESUMO
Within arterial plaque, HIV infection creates a state of inflammation and immune activation, triggering NLRP3/caspase-1 inflammasome, tissue damage, and monocyte/macrophage infiltration. Previously, we documented that caspase-1 activation in myeloid cells was linked with HIV-associated atherosclerosis in mice and people with HIV. Here, we mechanistically examined the direct effect of caspase-1 on HIV-associated atherosclerosis. Caspase-1-deficient (Casp-1-/-) mice were crossed with HIV-1 transgenic (Tg26+/-) mice with an atherogenic ApoE-deficient (ApoE-/-) background to create global caspase-1-deficient mice (Tg26+/-/ApoE-/-/Casp-1-/-). Caspase-1-sufficient (Tg26+/-/ApoE-/-/Casp-1+/+) mice served as the controls. Next, we created chimeric hematopoietic cell-deficient mice by reconstituting irradiated ApoE-/- mice with bone marrow cells transplanted from Tg26+/-/ApoE-/-/Casp-1-/- (BMT Casp-1-/-) or Tg26+/-/ApoE-/-/Casp-1+/+ (BMT Casp-1+/+) mice. Global caspase-1 knockout in mice suppressed plaque deposition in the thoracic aorta, serum IL-18 levels, and ex vivo foam cell formation. The deficiency of caspase-1 in hematopoietic cells resulted in reduced atherosclerotic plaque burden in the whole aorta and aortic root, which was associated with reduced macrophage infiltration. Transcriptomic analyses of peripheral mononuclear cells and splenocytes indicated that caspase-1 deficiency inhibited caspase-1 pathway-related genes. These results document the critical atherogenic role of caspase-1 in chronic HIV infection and highlight the implication of this pathway and peripheral immune activation in HIV-associated atherosclerosis.
Assuntos
Aterosclerose , Infecções por HIV , HIV-1 , Placa Aterosclerótica , Animais , Camundongos , Apolipoproteínas E/genética , Aterosclerose/genética , Caspase 1/genética , Infecções por HIV/complicações , Infecções por HIV/genética , Placa Aterosclerótica/genéticaRESUMO
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) induces a wide range of disease severity, ranging from asymptomatic infection to a life-threating illness, particularly in the elderly population and individuals with comorbid conditions. Among individuals with serious coronavirus 2019 (COVID-19) disease, acute respiratory distress syndrome (ARDS) is a common and often fatal presentation. Animal models of SARS-CoV-2 infection that manifest severe disease are needed to investigate the pathogenesis of COVID-19-induced ARDS and evaluate therapeutic strategies. We report two cases of ARDS in two aged African green monkeys (AGMs) infected with SARS-CoV-2 that had pathological lesions and disease similar to severe COVID-19 in humans. We also report a comparatively mild COVID-19 phenotype characterized by minor clinical, radiographic, and histopathologic changes in the two surviving, aged AGMs and four rhesus macaques (RMs) infected with SARS-CoV-2. Notable increases in circulating cytokines were observed in three of four infected, aged AGMs but not in infected RMs. All the AGMs had increased levels of plasma IL-6 compared with baseline, a predictive marker and presumptive therapeutic target in humans infected with SARS-CoV-2. Together, our results indicate that both RMs and AGMs are capable of modeling SARS-CoV-2 infection and suggest that aged AGMs may be useful for modeling severe disease manifestations, including ARDS.
Assuntos
COVID-19/etiologia , Pulmão/virologia , SARS-CoV-2/patogenicidade , Envelhecimento , Animais , Chlorocebus aethiops/virologia , Infecções por Coronavirus/tratamento farmacológico , Citocinas/metabolismo , Humanos , Pulmão/patologia , Macaca mulatta/virologia , Carga Viral/métodosRESUMO
Preclinical mouse models that recapitulate some characteristics of coronavirus disease (COVID-19) will facilitate focused study of pathogenesis and virus-host responses. Human agniotensin-converting enzyme 2 (hACE2) serves as an entry receptor for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) to infect people via binding to envelope spike proteins. Herein we report development and characterization of a rapidly deployable COVID-19 mouse model. C57BL/6J (B6) mice expressing hACE2 in the lung were transduced by oropharyngeal delivery of the recombinant human adenovirus type 5 that expresses hACE2 (Ad5-hACE2). Mice were infected with SARS-CoV-2 at Day 4 after transduction and developed interstitial pneumonia associated with perivascular inflammation, accompanied by significantly higher viral load in lungs at Days 3, 6, and 12 after infection compared with Ad5-empty control group. SARS-CoV-2 was detected in pneumocytes in alveolar septa. Transcriptomic analysis of lungs demonstrated that the infected Ad5-hACE mice had a significant increase in IFN-dependent chemokines Cxcl9 and Cxcl10, and genes associated with effector T-cell populations including Cd3 g, Cd8a, and Gzmb. Pathway analysis showed that several Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways were enriched in the data set, including cytokine-cytokine receptor interaction, the chemokine signaling pathway, the NOD-like receptor signaling pathway, the measles pathway, and the IL-17 signaling pathway. This response is correlative to clinical response in lungs of patients with COVID-19. These results demonstrate that expression of hACE2 via adenovirus delivery system sensitized the mouse to SARS-CoV-2 infection and resulted in the development of a mild COVID-19 phenotype, highlighting the immune and inflammatory host responses to SARS-CoV-2 infection. This rapidly deployable COVID-19 mouse model is useful for preclinical and pathogenesis studies of COVID-19.
Assuntos
Células Epiteliais Alveolares/imunologia , COVID-19/imunologia , Expressão Gênica , SARS-CoV-2/imunologia , Transdução de Sinais/imunologia , Adenoviridae/genética , Adenoviridae/metabolismo , Células Epiteliais Alveolares/metabolismo , Células Epiteliais Alveolares/virologia , Enzima de Conversão de Angiotensina 2/biossíntese , Enzima de Conversão de Angiotensina 2/genética , Enzima de Conversão de Angiotensina 2/imunologia , Animais , COVID-19/genética , COVID-19/metabolismo , COVID-19/patologia , Citocinas/genética , Citocinas/imunologia , Modelos Animais de Doenças , Humanos , Camundongos , Camundongos Transgênicos , SARS-CoV-2/genética , SARS-CoV-2/metabolismo , Transdução de Sinais/genética , Transdução GenéticaRESUMO
Adipocyte death occurs under various physiopathological conditions, including obesity and alcohol drinking, and can trigger organ damage particularly in the liver, but the underlying mechanisms remain obscure. To explore these mechanisms, we developed a mouse model of inducible adipocyte death by overexpressing the human CD59 (hCD59) on adipocytes (adipocyte-specific hCD59 transgenic mice). Injection of these mice with intermedilysin (ILY), which rapidly lyses hCD59 expressing cells exclusively by binding to the hCD59 but not mouse CD59, resulted in the acute selective death of adipocytes, adipose macrophage infiltration, and elevation of serum free fatty acid (FFA) levels. ILY injection also resulted in the secondary damage to multiple organs with the strongest injury observed in the liver, with inflammation and hepatic macrophage activation. Mechanistically, acute adipocyte death elevated epinephrine and norepinephrine levels and activated lipolysis pathways in adipose tissue in a chemokine (C-C motif) receptor 2-positive (CCR2+ ) macrophage-dependent manner, which was followed by FFA release and lipotoxicity in the liver. Additionally, acute adipocyte death caused hepatic CCR2+ macrophage activation and infiltration, further exacerbating liver injury. Conclusion: Adipocyte death predominantly induces liver injury and inflammation, which is probably due to the superior sensitivity of hepatocytes to lipotoxicity and the abundance of macrophages in the liver.
Assuntos
Adipócitos/fisiologia , Tecido Adiposo/enzimologia , Hepatopatias/etiologia , Macrófagos/fisiologia , Receptores CCR2/metabolismo , Animais , Bacteriocinas , Morte Celular , Modelos Animais de Doenças , Epinefrina/sangue , Ácidos Graxos não Esterificados/sangue , Feminino , Inflamação/etiologia , Isoproterenol , Lipólise , Hepatopatias/sangue , Masculino , Camundongos Transgênicos , Norepinefrina/sangue , Receptores CCR2/genéticaRESUMO
Targeted cell ablation is a powerful approach for studying the role of specific cell populations in a variety of organotypic functions, including cell differentiation, and organ generation and regeneration. Emerging tools for permanently or conditionally ablating targeted cell populations and transiently inhibiting neuronal activities exhibit a diversity of application and utility. Each tool has distinct features, and none can be universally applied to study different cell types in various tissue compartments. Although these tools have been developed for over 30 years, they require additional improvement. Currently, there is no consensus on how to select the tools to answer the specific scientific questions of interest. Selecting the appropriate cell ablation technique to study the function of a targeted cell population is less straightforward than selecting the method to study a gene's functions. In this review, we discuss the features of the various tools for targeted cell ablation and provide recommendations for optimal application of specific approaches.
Assuntos
Bacteriocinas/metabolismo , Ácido Clodrônico/química , Toxina Diftérica/genética , Optogenética/métodos , Simplexvirus/fisiologia , Animais , Ácido Clodrônico/toxicidade , Toxina Diftérica/metabolismo , Humanos , Intoxicação por MPTP/metabolismo , Intoxicação por MPTP/patologia , Neurônios/fisiologia , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Simplexvirus/enzimologiaRESUMO
AIMS: Endothelial progenitor cells (EPCs) play a crucial role in postnatal angiogenesis and neovascularization. Inward rectifier potassium channel 2.1 (Kir2.1) have been identified in EPCs. However, the effect of Kir2.1 on EPC function is not known. Here, we try to establish the role of Kir2.1 channels in EPC function and to provide first insights into the mechanisms. METHODS AND RESULTS: We first observed that the expression of Kir2.1 gradually decreased with the differentiation of EPCs into ECs in gene and protein levels. Treatment with the Kir2.1-selective inhibitor ML133 or knockdown of Kir2.1 by shRNA triggered EPC depolarization and promoted EPC biological functions, such as migration, adhesion, angiogenesis and differentiation into ECs in vitro. Transplantation of ML133-treated or Kir2.1 knockdown EPCs facilitated re-endothelialization in the rat injured arterial segment and inhibited neointima formation in vivo. In parallel, ML133 significantly enhanced autophagy and autophagic flux. After suppression of autophagy by 3-methyladenine (3-MA), the effects of ML133 on in vitro function and in vivo endothelialization capacity of EPCs were significantly inhibited. Mechanistically, ML133-induced autophagy was mediated at least partly by increased the activity of reactive oxygen species (ROS) that likely through intracellular calcium. CONCLUSION: Our study indicates that blocking or knockdown Kir2.1 results in a moderate depolarization of EPCs, which directly participated in enhancing EPC functions both in vitro and in vivo. In the mean time, autophagy signaling pathway is, at least in part, involved in this process. It may provide a potential target for the treatment or prevention of vascular injury and disease.
Assuntos
Potenciais de Ação , Autofagia , Diferenciação Celular , Células Progenitoras Endoteliais/metabolismo , Canais de Potássio Corretores do Fluxo de Internalização/metabolismo , Potenciais de Ação/efeitos dos fármacos , Animais , Artérias/efeitos dos fármacos , Artérias/patologia , Autofagia/efeitos dos fármacos , Cálcio/metabolismo , Diferenciação Celular/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Células Progenitoras Endoteliais/efeitos dos fármacos , Imidazóis/farmacologia , Neointima/patologia , Neovascularização Fisiológica/efeitos dos fármacos , Fenantrolinas/farmacologia , Ratos Sprague-Dawley , Espécies Reativas de Oxigênio/metabolismoRESUMO
In the antiretroviral therapy (ART) era, chronic HIV infection is primarily associated with chronic inflammation driving comorbidities such as cardiovascular disease and neurocognitive impairment. Caspase-1 activation in leukocytes has been documented in HIV infection; however, whether caspase-1 activation and the downstream pro-inflammatory cytokines interleukin-1beta (IL-1ß) and interleukin-18 (IL-18) contribute to chronic inflammation in HIV comorbidities remains undetermined. The relationship between the caspase-1 cascade and persistent inflammation in HIV has not been investigated. Here, we used an accelerated simian immunodeficiency virus (SIV)-infected rhesus macaque model with or without ART to investigate the dynamics of caspase-1 and immune cell activation before infection, 21 days post infection (dpi), and necropsy. Caspase-1, IL-18, IL-1ß, and immune markers were measured both in the circulation and lymphoid tissues. We found a significant increase in caspase-1 and IL-18 in SIV infection that positively correlated with inflammatory monocytes and negatively correlated with CD4+ T cell counts. ART attenuated these effects at necropsy in the circulation. Further, lymph nodes from SIV+ or SIV+ART animals had increased activation of caspase-1 and potential upstream priming of the NF-κB pathway, indicating that tissue-specific immune activation persists with ART. Together, these results shed light on the interconnectedness of the caspase-1 pathway and peripheral immune activation and further indicate that ART is not sufficient for suppressing inflammation. The caspase-1 pathway may provide novel therapeutic targets to improve HIV-associated comorbidities and health outcomes in the context of viral suppression.
Assuntos
Caspase 1/imunologia , Síndrome de Imunodeficiência Adquirida dos Símios/imunologia , Vírus da Imunodeficiência Símia/imunologia , Animais , Inflamação/imunologia , Inflamação/virologia , Macaca mulattaRESUMO
RATIONALE: Endothelial injury is an initial mechanism mediating cardiovascular disease. OBJECTIVE: Here, we investigated the effect of hyperhomocysteinemia on programed cell death in endothelial cells (EC). METHODS AND RESULTS: We established a novel flow-cytometric gating method to define pyrotosis (Annexin V(-)/Propidium iodide(+)). In cultured human EC, we found that: (1) homocysteine and lipopolysaccharide individually and synergistically induced inflammatory pyroptotic and noninflammatory apoptotic cell death; (2) homocysteine/lipopolysaccharide induced caspase-1 activation before caspase-8, caspase-9, and caspase-3 activations; (3) caspase-1/caspase-3 inhibitors rescued homocysteine/lipopolysaccharide-induced pyroptosis/apoptosis, but caspase-8/caspase-9 inhibitors had differential rescue effect; (4) homocysteine/lipopolysaccharide-induced nucleotide-binding oligomerization domain, and leucine-rich repeat and pyrin domain containing protein 3 (NLRP3) protein caused NLRP3-containing inflammasome assembly, caspase-1 activation, and interleukin (IL)-1ß cleavage/activation; (5) homocysteine/lipopolysaccharide elevated intracellular reactive oxygen species, (6) intracellular oxidative gradient determined cell death destiny as intermediate intracellular reactive oxygen species levels are associated with pyroptosis, whereas high reactive oxygen species corresponded to apoptosis; (7) homocysteine/lipopolysaccharide induced mitochondrial membrane potential collapse and cytochrome-c release, and increased B-cell lymphoma 2-associated X protein/B-cell lymphoma 2 ratio which were attenuated by antioxidants and caspase-1 inhibitor; and (8) antioxidants extracellular superoxide dismutase and catalase prevented homocysteine/lipopolysaccharide -induced caspase-1 activation, mitochondrial dysfunction, and pyroptosis/apoptosis. In cystathionine ß-synthase-deficient (Cbs(-/-)) mice, severe hyperhomocysteinemia-induced caspase-1 activation in isolated lung EC and caspase-1 expression in aortic endothelium, and elevated aortic caspase-1, caspase-9 protein/activity and B-cell lymphoma 2-associated X protein/B-cell lymphoma 2 ratio in Cbs(-/-) aorta and human umbilical vein endothelial cells. Finally, homocysteine-induced DNA fragmentation was reversed in caspase-1(-/-) EC. Hyperhomocysteinemia-induced aortic endothelial dysfunction was rescued in caspase-1(-/-) and NLRP3(-/-) mice. CONCLUSIONS: Hyperhomocysteinemia preferentially induces EC pyroptosis via caspase-1-dependent inflammasome activation leading to endothelial dysfunction. We termed caspase-1 responsive pyroptosis and apoptosis as pyrop-apoptosis.
Assuntos
Caspase 1/metabolismo , Células Endoteliais da Veia Umbilical Humana/metabolismo , Inflamassomos/metabolismo , Piroptose , Animais , Cistationina beta-Sintase/genética , Cistationina beta-Sintase/metabolismo , Homocisteína/toxicidade , Humanos , Interleucina-1beta/metabolismo , Lipopolissacarídeos/toxicidade , Camundongos , Camundongos Endogâmicos C57BL , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Proteína X Associada a bcl-2/metabolismoRESUMO
CRISPR-associated protein 9 (Cas9)-mediated genome editing provides a promising cure for HIV-1/AIDS; however, gene delivery efficiency in vivo remains an obstacle to overcome. Here, we demonstrate the feasibility and efficiency of excising the HIV-1 provirus in three different animal models using an all-in-one adeno-associated virus (AAV) vector to deliver multiplex single-guide RNAs (sgRNAs) plus Staphylococcus aureus Cas9 (saCas9). The quadruplex sgRNAs/saCas9 vector outperformed the duplex vector in excising the integrated HIV-1 genome in cultured neural stem/progenitor cells from HIV-1 Tg26 transgenic mice. Intravenously injected quadruplex sgRNAs/saCas9 AAV-DJ/8 excised HIV-1 proviral DNA and significantly reduced viral RNA expression in several organs/tissues of Tg26 mice. In EcoHIV acutely infected mice, intravenously injected quadruplex sgRNAs/saCas9 AAV-DJ/8 reduced systemic EcoHIV infection, as determined by live bioluminescence imaging. Additionally, this quadruplex vector induced efficient proviral excision, as determined by PCR genotyping in the liver, lungs, brain, and spleen. Finally, in humanized bone marrow/liver/thymus (BLT) mice with chronic HIV-1 infection, successful proviral excision was detected by PCR genotyping in the spleen, lungs, heart, colon, and brain after a single intravenous injection of quadruplex sgRNAs/saCas9 AAV-DJ/8. In conclusion, in vivo excision of HIV-1 proviral DNA by sgRNAs/saCas9 in solid tissues/organs can be achieved via AAV delivery, a significant step toward human clinical trials.
Assuntos
Endonucleases/genética , Terapia Genética/métodos , Genoma Viral , Infecções por HIV/terapia , HIV-1/genética , Provírus/genética , RNA Guia de Cinetoplastídeos/genética , Animais , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Sequência de Bases , Sistemas CRISPR-Cas , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Dependovirus/genética , Dependovirus/metabolismo , Modelos Animais de Doenças , Endonucleases/metabolismo , Edição de Genes/métodos , Vetores Genéticos/química , Vetores Genéticos/metabolismo , Infecções por HIV/patologia , Infecções por HIV/virologia , Repetição Terminal Longa de HIV , HIV-1/metabolismo , Humanos , Camundongos , Camundongos Transgênicos , Oligonucleotídeos/genética , Oligonucleotídeos/metabolismo , Provírus/metabolismo , RNA Guia de Cinetoplastídeos/metabolismo , Staphylococcus aureus/química , Staphylococcus aureus/enzimologia , Produtos do Gene gag do Vírus da Imunodeficiência Humana/genética , Produtos do Gene gag do Vírus da Imunodeficiência Humana/metabolismo , Produtos do Gene pol do Vírus da Imunodeficiência Humana/genética , Produtos do Gene pol do Vírus da Imunodeficiência Humana/metabolismoRESUMO
Effective methods for cell ablation are important tools for examining the anatomical, functional, and behavioral consequences of selective loss of specific cell types in animal models. We have developed an ablation system based on creating genetically modified animals that express human CD59 (hCD59), a membrane receptor, and administering intermedilysin (ILY), a toxin produced by Streptococcus intermedius, which binds specifically to hCD59 to induce cell lysis. As proof-of-concept in the rat, we generated an anemia model, SD-Tg(CD59-HBA1)Bryd, which expresses hCD59 on erythrocytes. Hemolysis is a common complication of inherited or acquired blood disorders, which can result in cardiovascular compromise and death. A rat model that can replicate hemolysis through specific ablation of erythrocytes would allow further study of disease and novel treatments. In vitro, complete lysis of erythrocytes expressing hCD59 was observed at and above 250 pM ILY, while no lysis was observed in wild-type erythrocytes at any ILY concentration (8-1,000 pM). In vivo, ILY intravenous injection (100 ng/g body wt) dramatically reduced the hematocrit within 10 min, with a mean hematocrit reduction of 43% compared with 1.4% in the saline control group. Rats injected with ILY at 500 ng/g intraperitoneally developed gross signs of anemia. Histopathology confirmed anemia and revealed hepatic necrosis, with microthrombi present. These studies validate the hCD59-ILY cell ablation technology in the rat and provide the scientific community with a new rapid conditional targeted ablation model for hemolytic anemia and hemolysis-associated sequelae.
Assuntos
Anemia Hemolítica/genética , Animais Geneticamente Modificados/genética , Anemia Hemolítica/metabolismo , Animais , Animais Geneticamente Modificados/metabolismo , Antígenos CD59/genética , Antígenos CD59/metabolismo , DNA Complementar/genética , Eritrócitos/metabolismo , Feminino , Hemólise/genética , Humanos , Masculino , RatosRESUMO
Deficient angiogenesis may contribute to worsen the prognosis of myocardial ischemia, peripheral arterial disease, ischemic stroke, etc. Dyslipidemic and inflammatory environments attenuate endothelial cell (EC) proliferation and angiogenesis, worsening the prognosis of ischemia. Under these dyslipidemic and inflammatory environments, EC-caspase-1 becomes activated and induces inflammatory cell death that is defined as pyroptosis. However, the underlying mechanism that correlates caspase-1 activation with angiogenic impairment and the prognosis of ischemia remains poorly defined. By using flow cytometric analysis, enzyme and receptor inhibitors, and hind limb ischemia model in caspase-1 knock-out (KO) mice, we examined our novel hypothesis, i.e. inhibition of caspase-1 in ECs under dyslipidemic and inflammatory environments attenuates EC pyroptosis, improves EC survival mediated by vascular endothelial growth factor receptor 2 (VEGFR-2), angiogenesis, and the prognosis of ischemia. We have made the following findings. Proatherogenic lipids induce higher caspase-1 activation in larger sizes of human aortic endothelial cells (HAECs) than in smaller sizes of HAECs. Proatherogenic lipids increase pyroptosis significantly more in smaller sizes of HAECs than in larger sizes of the cells. VEGFR-2 inhibition increases caspase-1 activation in HAECs induced by lysophosphatidylcholine treatment. Caspase-1 activation inhibits VEGFR-2 expression. Caspase-1 inhibition improves the tube formation of lysophosphatidylcholine-treated HAECs. Finally, caspase-1 depletion improves angiogenesis and blood flow in mouse hind limb ischemic tissues. Our results have demonstrated for the first time that inhibition of proatherogenic caspase-1 activation in ECs improves angiogenesis and the prognosis of ischemia.
Assuntos
Caspase 1/metabolismo , Inibidores de Caspase/farmacologia , Isquemia/tratamento farmacológico , Neovascularização Fisiológica/efeitos dos fármacos , Animais , Caspase 1/deficiência , Caspase 1/genética , Morte Celular/efeitos dos fármacos , Tamanho Celular , Células Cultivadas , Modelos Animais de Doenças , Células Endoteliais/citologia , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/enzimologia , Técnicas de Silenciamento de Genes , Membro Posterior/irrigação sanguínea , Humanos , Isquemia/enzimologia , Isquemia/patologia , Lipídeos/química , Lipídeos/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/antagonistas & inibidores , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/genética , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismoRESUMO
There is increased interest in immune-based monoclonal antibody therapies for different malignancies because of their potential specificity and limited toxicity. The activity of some therapeutic monoclonal antibodies is partially dependent on complement-dependent cytolysis (CDC), in which the immune system surveys for invading pathogens, infected cells, and malignant cells and facilitates their destruction. CD59 is a ubiquitously expressed cell-surface glycosylphosphatidylinositol-anchored protein that protects cells from CDC. However, in certain tumors, CD59 expression is enhanced, posing a significant obstacle for treatment, by hindering effective monoclonal antibody-induced CDC. In this study, we used non-small lung carcinoma cells to characterize the mechanism of a novel CD59 inhibitor: the 114-amino acid recombinant form of the 4th domain of intermedilysin (rILYd4), a pore forming toxin secreted by Streptococcus intermedius. We compared the rates of internalization of CD59 in the presence of rILYd4 or anti-CD59 antibodies and determined that rILYd4 induces more rapid CD59 uptake at early time points. Most significantly, upon binding to rILYd4, CD59 is internalized and undergoes massive degradation in lysosomes within minutes. The remaining rILYd4·CD59 complexes recycle to the PM and are shed from the cell. In comparison, upon internalization of CD59 via anti-CD59 antibody binding, the antibody·CD59 complex is recycled via early and recycling endosomes, mostly avoiding degradation. Our study supports a novel role for rILYd4 in promoting internalization and rapid degradation of the complement inhibitor CD59, and highlights the potential for improving CDC-based immunotherapy.
Assuntos
Antígenos CD59/metabolismo , Antígenos CD59/genética , Antígenos CD59/imunologia , Linhagem Celular Tumoral , Endocitose , Humanos , Hidrólise , Interferência de RNA , RNA Interferente Pequeno/genéticaRESUMO
The various neurological complications associated with HIV-1 infection, specifically HIV-associated neurocognitive disorders (HAND) persist as a major public health burden worldwide. Despite the widespread use of anti-retroviral therapy, the prevalence of HAND is significantly high. HAND results from the direct effects of an HIV-1 infection as well as secondary effects of HIV-1-induced immune reaction and inflammatory response. Complement, a critical mediator of innate and acquired immunity, plays important roles in defeating many viral infections by the formation of a lytic pore or indirectly by opsonization and recruitment of phagocytes. While the role of complement in the pathogenesis of HIV-1 infection and HAND has been previously recognized for over 15 years, it has been largely underestimated thus far. Complement can be activated through HIV-1 envelope proteins, mannose-binding lectins (MBL), and anti-HIV-1 antibodies. Complement not only fights against HIV-1 infection but also enhances HIV-1 infection. In addition, HIV-1 can hijack complement regulators such as CD59 and CD55 and can utilize these regulators and factor H to escape from complement attack. Normally, complement levels in brain are much lower than plasma levels and there is no or little complement deposition in brain cells. Interestingly, local production and deposition of complement are dramatically increased in HIV-1-infected brain, indicating that complement may contribute to the pathogenesis of HAND. Here, we review the current understanding of the role of complement in HIV-1 infection and HAND, as well as potential therapeutic approaches targeting the complement system for the treatment and eradications of HIV-1 infection.
Assuntos
Complexo AIDS Demência/imunologia , Anticorpos Antivirais/sangue , Proteínas do Sistema Complemento/metabolismo , Anticorpos Anti-HIV/sangue , HIV-1/imunologia , Complexo AIDS Demência/sangue , Complexo AIDS Demência/patologia , Complexo AIDS Demência/virologia , Imunidade Adaptativa , Complexo Antígeno-Anticorpo/sangue , Antígenos CD/imunologia , HIV-1/patogenicidade , Humanos , Evasão da Resposta Imune , Imunidade Inata , Lectinas de Ligação a Manose/imunologia , Proteínas do Envelope Viral/imunologiaRESUMO
Macrophages are exceptionally diversified cell types and perform unique features and functions when exposed to different stimuli within the specific microenvironment of various kidney diseases. In instances of kidney tissue necrosis or infection, specific patterns associated with damage or pathogens prompt the development of pro-inflammatory macrophages (M1). These M1 macrophages contribute to exacerbating tissue damage, inflammation, and eventual fibrosis. Conversely, anti-inflammatory macrophages (M2) arise in the same circumstances, contributing to kidney repair and regeneration processes. Impaired tissue repair causes fibrosis, and hence macrophages play a protective and pathogenic role. In response to harmful stimuli within the body, inflammasomes, complex assemblies of multiple proteins, assume a pivotal function in innate immunity. The initiation of inflammasomes triggers the activation of caspase 1, which in turn facilitates the maturation of cytokines, inflammation, and cell death. Macrophages in the kidneys possess the complete elements of the NLRP3 inflammasome, including NLRP3, ASC, and pro-caspase-1. When the NLRP3 inflammasomes are activated, it triggers the activation of caspase-1, resulting in the release of mature proinflammatory cytokines (IL)-1ß and IL-18 and cleavage of Gasdermin D (GSDMD). This activation process therefore then induces pyroptosis, leading to renal inflammation, cell death, and renal dysfunction. The NLRP3-ASC-caspase-1-IL-1ß-IL-18 pathway has been identified as a factor in the development of the pathophysiology of numerous kidney diseases. In this review, we explore current progress in understanding macrophage behavior concerning inflammation, injury, and fibrosis in kidneys. Emphasizing the pivotal role of activated macrophages in both the advancement and recovery phases of renal diseases, the article delves into potential strategies to modify macrophage functionality and it also discusses emerging approaches to selectively target NLRP3 inflammasomes and their signaling components within the kidney, aiming to facilitate the healing process in kidney diseases.
RESUMO
In the field of brain-to-text communication, it is difficult to finish highly dexterous behaviors of writing multi-character by motor-imagery-based brain-computer interface (MI-BCI), setting a barrier to restore communication in people who have lost the ability to move and speak. In this paper, we design and implement a multi-character classification scheme based on 29 characters of motor imagery (MI) electroencephalogram (EEG) signals, which contains 26 English letters and 3 punctuation marks. Firstly, we design a novel experimental paradigm to increase the variety of BCI inputs by asking subjects to imagine the movement of writing 29 characters instead of gross motor skills such as reaching or grasping. Secondly, because of the high dimension of EEG signals, we adopt power spectral density (PSD), principal components analysis (PCA), kernel principal components analysis (KPCA) respectively to decompose EEG signals and extract feature, and then test the results with pearson product-moment correlation coefficient (PCCs). Thirdly, we respectively employ k-nearest neighbor (kNN), support vector machine (SVM), extreme learning machine (ELM) and light gradient boosting machine (LightGBM) to classify 29 characters and compare the results. We have implemented a complete scheme, including paradigm design, signal acquisition, feature extraction and classification, which can effectively classify 29 characters. The experimental results show that the KPCA has the best feature extraction effect and the kNN has the highest classification accuracy, with the final classification accuracy reaching 96.2%, which is better than other studies.
Assuntos
Interfaces Cérebro-Computador , Eletroencefalografia , Humanos , Eletroencefalografia/métodos , Imagens, Psicoterapia , Movimento , Encéfalo , Algoritmos , ImaginaçãoRESUMO
Brain-machine interface (BMI) can convert electroencephalography signals (EEGs) into the control instructions of external devices, and the key of control performance is the accuracy and efficiency of decoder. However, the performance of different decoders obtaining control instructions from complex and variable EEG signals is very different and irregular in the different neural information transfer model. Aiming at this problem, the off-line and on-line performance of eight decoders based on the improved single-joint information transmission (SJIT) model is compared and analyzed in this paper, which can provide a theoretical guidance for decoder design. Firstly, in order to avoid the different types of neural activities in the decoding process on the decoder performance, eight decoders based on the improved SJIT model are designed. And then the off-line decoding performance of these decoders is tested and compared. Secondly, a closed-loop BMI system which combining by the designed decoder and the random forest encoder based on the improved SJIT model is constructed. Finally, based on the constructed closed-loop BMI system, the on-line decoding performance of decoders is compared and analyzed. The results show that the LSTM-based decoder has better on-line decoding performance than others in the improved SJIT model.
RESUMO
Computational modeling has emerged as a time-saving and cost-effective alternative to traditional animal testing for assessing chemicals for their potential hazards. However, few computational modeling studies for immunotoxicity were reported, with few models available for predicting toxicants due to the lack of training data and the complex mechanisms of immunotoxicity. In this study, we employed a data-driven quantitative structure-activity relationship (QSAR) modeling workflow to extensively enlarge the limited training data by revealing multiple targets involved in immunotoxicity. To this end, a probe data set of 6,341 chemicals was obtained from a high-throughput screening (HTS) assay testing for the activation of the aryl hydrocarbon receptor (AhR) signaling pathway, a key event leading to immunotoxicity. Searching this probe data set against PubChem yielded 3,183 assays with testing results for varying proportions of these 6,341 compounds. 100 assays were selected to develop QSAR models based on their correlations to AhR agonism. Twelve individual QSAR models were built for each assay using combinations of four machine-learning algorithms and three molecular fingerprints. 5-fold cross-validation of the resulting models showed good predictivity (average CCR = 0.73). A total of 20 assays were further selected based on QSAR model performance, and their resulting QSAR models showed good predictivity of potential immunotoxicants from external chemicals. This study provides a computational modeling strategy that can utilize large public toxicity data sets for modeling immunotoxicity and other toxicity endpoints, which have limited training data and complicated toxicity mechanisms.