RESUMO
Background & Aims: Lipid droplet (LD) accumulation in cells and tissues is understood to be an evolutionarily conserved tissue tolerance mechanism to prevent lipotoxicity caused by excess lipids; however, the presence of excess LDs has been associated with numerous diseases. Sepsis triggers the reprogramming of lipid metabolism and LD accumulation in cells and tissues, including the liver. The functions and consequences of sepsis-triggered liver LD accumulation are not well known. Methods: Experimental sepsis was induced by CLP (caecal ligation and puncture) in mice. Markers of hepatic steatosis, liver injury, hepatic oxidative stress, and inflammation were analysed using a combination of functional, imaging, lipidomic, protein expression and immune-enzymatic assays. To prevent LD formation, mice were treated orally with A922500, a pharmacological inhibitor of DGAT1. Results: We identified that liver LD overload correlates with liver injury and sepsis severity. Moreover, the progression of steatosis from 24 h to 48 h post-CLP occurs in parallel with increased cytokine expression, inflammatory cell recruitment and oxidative stress. Lipidomic analysis of purified LDs demonstrated that sepsis leads LDs to harbour increased amounts of unsaturated fatty acids, mostly 18:1 and 18:2. An increased content of lipoperoxides within LDs was also observed. Conversely, the impairment of LD formation by inhibition of the DGAT1 enzyme reduces levels of hepatic inflammation and lipid peroxidation markers and ameliorates sepsis-induced liver injury. Conclusions: Our results indicate that sepsis triggers lipid metabolism alterations that culminate in increased liver LD accumulation. Increased LDs are associated with disease severity and liver injury. Moreover, inhibition of LD accumulation decreased the production of inflammatory mediators and lipid peroxidation while improving tissue function, suggesting that LDs contribute to the pathogenesis of liver injury triggered by sepsis. Impact and Implications: Sepsis is a complex life-threatening syndrome caused by dysregulated inflammatory and metabolic host responses to infection. The observation that lipid droplets may contribute to sepsis-associated organ injury by amplifying lipid peroxidation and inflammation provides a rationale for therapeutically targeting lipid droplets and lipid metabolism in sepsis.
RESUMO
Occurrence of hyperglycemia upon infection is associated with worse clinical outcome in COVID-19 patients. However, it is still unknown whether SARS-CoV-2 directly triggers hyperglycemia. Herein, we interrogated whether and how SARS-CoV-2 causes hyperglycemia by infecting hepatocytes and increasing glucose production. We performed a retrospective cohort study including patients that were admitted at a hospital with suspicion of COVID-19. Clinical and laboratory data were collected from the chart records and daily blood glucose values were analyzed to test the hypothesis on whether COVID-19 was independently associated with hyperglycemia. Blood glucose was collected from a subgroup of nondiabetic patients to assess pancreatic hormones. Postmortem liver biopsies were collected to assess the presence of SARS-CoV-2 and its transporters in hepatocytes. In human hepatocytes, we studied the mechanistic bases of SARS-CoV-2 entrance and its gluconeogenic effect. SARS-CoV-2 infection was independently associated with hyperglycemia, regardless of diabetic history and beta cell function. We detected replicating viruses in human hepatocytes from postmortem liver biopsies and in primary hepatocytes. We found that SARS-CoV-2 variants infected human hepatocytes in vitro with different susceptibility. SARS-CoV-2 infection in hepatocytes yields the release of new infectious viral particles, though not causing cell damage. We showed that infected hepatocytes increase glucose production and this is associated with induction of PEPCK activity. Furthermore, our results demonstrate that SARS-CoV-2 entry in hepatocytes occurs partially through ACE2- and GRP78-dependent mechanisms. SARS-CoV-2 infects and replicates in hepatocytes and exerts a PEPCK-dependent gluconeogenic effect in these cells that potentially is a key cause of hyperglycemia in infected patients.
Assuntos
COVID-19 , Hiperglicemia , Humanos , COVID-19/complicações , SARS-CoV-2 , Gluconeogênese , Glicemia , Estudos Retrospectivos , Hepatócitos , Hiperglicemia/complicações , GlucoseRESUMO
Visceral adiposity is a risk factor for severe COVID-19, and a link between adipose tissue infection and disease progression has been proposed. Here we demonstrate that SARS-CoV-2 infects human adipose tissue and undergoes productive infection in fat cells. However, susceptibility to infection and the cellular response depends on the anatomical origin of the cells and the viral lineage. Visceral fat cells express more ACE2 and are more susceptible to SARS-CoV-2 infection than their subcutaneous counterparts. SARS-CoV-2 infection leads to inhibition of lipolysis in subcutaneous fat cells, while in visceral fat cells, it results in higher expression of pro-inflammatory cytokines. Viral load and cellular response are attenuated when visceral fat cells are infected with the SARS-CoV-2 gamma variant. A similar degree of cell death occurs 4-days after SARS-CoV-2 infection, regardless of the cell origin or viral lineage. Hence, SARS-CoV-2 infects human fat cells, replicating and altering cell function and viability in a depot- and viral lineage-dependent fashion.
Assuntos
COVID-19 , SARS-CoV-2 , Tecido Adiposo , Enzima de Conversão de Angiotensina 2 , Citocinas , HumanosRESUMO
Coronavirus disease 2019 (COVID-19) is currently a worldwide emergency caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2). In observational clinical studies, statins have been identified as beneficial to hospitalized patients with COVID-19. However, experimental evidence of underlying statins protection against SARS-CoV-2 remains elusive. Here we reported for the first-time experimental evidence of the protective effects of simvastatin treatment both in vitro and in vivo. We found that treatment with simvastatin significantly reduced the viral replication and lung damage in vivo, delaying SARS-CoV-2-associated physiopathology and mortality in the K18-hACE2-transgenic mice model. Moreover, simvastatin also downregulated the inflammation triggered by SARS-CoV-2 infection in pulmonary tissue and in human neutrophils, peripheral blood monocytes, and lung epithelial Calu-3 cells in vitro, showing its potential to modulate the inflammatory response both at the site of infection and systemically. Additionally, we also observed that simvastatin affected the course of SARS-CoV-2 infection through displacing ACE2 on cell membrane lipid rafts. In conclusion, our results show that simvastatin exhibits early protective effects on SARS-CoV-2 infection by inhibiting virus cell entry and inflammatory cytokine production, through mechanisms at least in part dependent on lipid rafts disruption.
Assuntos
Tratamento Farmacológico da COVID-19 , Regulação para Baixo/efeitos dos fármacos , Inflamação/tratamento farmacológico , Microdomínios da Membrana/efeitos dos fármacos , SARS-CoV-2/patogenicidade , Sinvastatina/farmacologia , Animais , COVID-19/virologia , Modelos Animais de Doenças , Humanos , Inflamação/virologia , Pulmão/virologia , Camundongos , Camundongos Transgênicos , Replicação Viral/efeitos dos fármacosRESUMO
Infection by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been associated with leukopenia and uncontrolled inflammatory response in critically ill patients. A better comprehension of SARS-CoV-2-induced monocyte death is essential for the identification of therapies capable to control the hyper-inflammation and reduce viral replication in patients with 2019 coronavirus disease (COVID-19). Here, we show that SARS-CoV-2 engages inflammasome and triggers pyroptosis in human monocytes, experimentally infected, and from patients under intensive care. Pyroptosis associated with caspase-1 activation, IL-1ß production, gasdermin D cleavage, and enhanced pro-inflammatory cytokine levels in human primary monocytes. At least in part, our results originally describe mechanisms by which monocytes, a central cellular component recruited from peripheral blood to respiratory tract, succumb to control severe COVID-19.
RESUMO
Viruses are obligate intracellular parasites that make use of the host metabolic machineries to meet their biosynthetic needs. Thus, identifying the host pathways essential for the virus replication may lead to potential targets for therapeutic intervention. The mechanisms and pathways explored by SARS-CoV-2 to support its replication within host cells are not fully known. Lipid droplets (LD) are organelles with major functions in lipid metabolism, energy homeostasis and intracellular transport, and have multiple roles in infections and inflammation. Here we described that monocytes from COVID-19 patients have an increased LD accumulation compared to SARS-CoV-2 negative donors. In vitro, SARS-CoV-2 infection were seen to modulate pathways of lipid synthesis and uptake as monitored by testing for CD36, SREBP-1, PPARγ, and DGAT-1 expression in monocytes and triggered LD formation in different human cell lines. LDs were found in close apposition with SARS-CoV-2 proteins and double-stranded (ds)-RNA in infected Vero cells. Electron microscopy (EM) analysis of SARS-CoV-2 infected Vero cells show viral particles colocalizing with LDs, suggestive that LDs might serve as an assembly platform. Pharmacological modulation of LD formation by inhibition of DGAT-1 with A922500 significantly inhibited SARS-CoV-2 replication as well as reduced production of mediators pro-inflammatory response. Taken together, we demonstrate the essential role of lipid metabolic reprograming and LD formation in SARS-CoV-2 replication and pathogenesis, opening new opportunities for therapeutic strategies to COVID-19.
Assuntos
COVID-19/complicações , Mediadores da Inflamação/metabolismo , Inflamação/etiologia , Gotículas Lipídicas/patologia , SARS-CoV-2/isolamento & purificação , Animais , COVID-19/imunologia , COVID-19/patologia , COVID-19/virologia , Estudos de Casos e Controles , Chlorocebus aethiops , Humanos , Inflamação/metabolismo , Inflamação/patologia , Células Vero , Replicação ViralRESUMO
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is an emergent pathogen responsible for the coronavirus disease 2019 (COVID-19). Since its emergence, the novel coronavirus has rapidly achieved pandemic proportions causing remarkably increased morbidity and mortality around the world. A hypercoagulability state has been reported as a major pathologic event in COVID-19, and thromboembolic complications listed among life-threatening complications of the disease. Platelets are chief effector cells of hemostasis and pathological thrombosis. However, the participation of platelets in the pathogenesis of COVID-19 remains elusive. This report demonstrates that increased platelet activation and platelet-monocyte aggregate formation are observed in severe COVID-19 patients, but not in patients presenting mild COVID-19 syndrome. In addition, exposure to plasma from severe COVID-19 patients increased the activation of control platelets ex vivo. In our cohort of COVID-19 patients admitted to the intensive care unit, platelet-monocyte interaction was strongly associated with tissue factor (TF) expression by the monocytes. Platelet activation and monocyte TF expression were associated with markers of coagulation exacerbation as fibrinogen and D-dimers, and were increased in patients requiring invasive mechanical ventilation or patients who evolved with in-hospital mortality. Finally, platelets from severe COVID-19 patients were able to induce TF expression ex vivo in monocytes from healthy volunteers, a phenomenon that was inhibited by platelet P-selectin neutralization or integrin αIIb/ß3 blocking with the aggregation inhibitor abciximab. Altogether, these data shed light on new pathological mechanisms involving platelet activation and platelet-dependent monocyte TF expression, which were associated with COVID-19 severity and mortality.
Assuntos
Betacoronavirus/imunologia , Transtornos da Coagulação Sanguínea/patologia , Plaquetas/patologia , Infecções por Coronavirus/complicações , Monócitos/patologia , Pneumonia Viral/complicações , Tromboplastina/metabolismo , Adulto , Biomarcadores/metabolismo , Transtornos da Coagulação Sanguínea/imunologia , Transtornos da Coagulação Sanguínea/metabolismo , Transtornos da Coagulação Sanguínea/virologia , Plaquetas/metabolismo , Plaquetas/virologia , COVID-19 , Estudos de Casos e Controles , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/metabolismo , Infecções por Coronavirus/virologia , Feminino , Seguimentos , Humanos , Masculino , Pessoa de Meia-Idade , Monócitos/metabolismo , Monócitos/virologia , Selectina-P/metabolismo , Pandemias , Ativação Plaquetária , Pneumonia Viral/imunologia , Pneumonia Viral/metabolismo , Pneumonia Viral/virologia , Prognóstico , Estudos Prospectivos , SARS-CoV-2 , Taxa de SobrevidaRESUMO
Lipid droplets (also known as lipid bodies) are lipid-rich, cytoplasmic organelles that play important roles in cell signaling, lipid metabolism, membrane trafficking, and the production of inflammatory mediators. Lipid droplet biogenesis is a regulated process, and accumulation of these organelles within leukocytes, epithelial cells, hepatocytes, and other nonadipocyte cells is a frequently observed phenotype in several physiologic or pathogenic situations and is thoroughly described during inflammatory conditions. Moreover, in recent years, several studies have described an increase in intracellular lipid accumulation in different neoplastic processes, although it is not clear whether lipid droplet accumulation is directly involved in the establishment of these different types of malignancies. This review discusses current evidence related to the biogenesis, composition and functions of lipid droplets related to the hallmarks of cancer: inflammation, cell metabolism, increased proliferation, escape from cell death, and hypoxia. Moreover, the potential of lipid droplets as markers of disease and targets for novel anti-inflammatory and antineoplastic therapies will be discussed.
Assuntos
Transformação Celular Neoplásica/metabolismo , Gotículas Lipídicas/metabolismo , Neoplasias/metabolismo , Animais , Morte Celular , Proliferação de Células , Transformação Celular Neoplásica/patologia , Metabolismo Energético , Humanos , Mediadores da Inflamação/metabolismo , Gotículas Lipídicas/patologia , Neoplasias/patologia , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Transdução de Sinais , Hipóxia Tumoral , Microambiente TumoralRESUMO
O câncer de esôfago (CE) é a oitava neoplasia mais incidente no mundo e a sexta mais mortal. Há dois subtipos histológicos: o adenocarcinoma (ADE) e o carcinoma epidermoide de esôfago (CEE). A incidência do ADE é a que mais cresce no mundo. Sabe-se que células tumorais consomem glicose avidamente produzindo ácido lático, mesmo em condições de normóxia. Outras alterações metabólicas já foram descritas como o aumento da lipogênese e corpúsculos lipídicos (CLs). Estes são organelas formadas por uma monocamada de fosfolipídeos com um interior enriquecido com lipídeos neutros e proteínas. Os CLs têm papel na sinalização celular, tráfego de membranas, autofagia e síntese de eicosanóides. Um estudo brasileiro mostrou que tumores de ADE têm uma expressão elevada de proteínas estruturais de CLs. Contudo, há uma defasagem no conhecimento sobre como a desregulação do metabolismo lipídico influencia a biogênese e função de CLs em CE. Portanto, o objetivo dessa tese foi avaliar as alterações do metabolismo lipídico e a modulação de CLs em CE. Para tal, foi usado o modelo de linhagens tumorais TE11, TE1 e OE21, oriundas de CEE, OE33 e OE19, oriundas de ADE, e uma linhagem de epitélio esofágico não tumoral, HET1A. Nessas, foi investigada a presença e quantidade de CLs e proteínas estruturais de CLs como ADRP e TIP47. Enquanto a desregulação do metabolismo energético foi estudada pela expressão de enzimas lipogênicas e pela avaliação da composição lipídica da linhagem OE33 por espectrometria de massas. Além disso, as implicações clínico-patológicas destas alterações foram avaliadas em um banco de dados de transcriptômica.
Como resultado, observou-se que a linhagem OE33 apresentava uma maior quantidade de CLs e por isso foi escolhida no estudo como modelo de ADE. A proteína ADRP, foi expressa em todas as linhagens, enquanto TIP47 não foi expresso em HET1A. Além disso, observamos que enzimas lipogênicas ácido graxo sintase e estearoil-CoA dessaturase não estavam presentes em HET1A quando comparadas a OE33. A avaliação do conteúdo lipídico de OE33 revelou que esta linhagem tem CLs enriquecidos com triacilglicerol e ácidos graxos monoinsaturados e, enquanto o ácido oleico é o principal ácido graxo. Por isso, decidimos investigar o papel dos seguintes alvos: FASN (ácido graxo sintase), DGAT1 (diacilglicerol transferase 1) e SCD1 (estearoil-coA dessaturase). A enzima DGAT1 é importante para a biogênese de corpúsculo lipídico na linhagem OE33, no entanto, não é importante para a manutenção tumoral. Enquanto a inibição de FASN diminuiu a viabilidade celular de OE33 e a síntese de corpúsculos lipídicos. A inibição de SCD1 afeta a proliferação, e apesar de não alterar a quantidade de CLs, modula a sua composição. Além disso, a expressão de SCD1 tem uma correlação com pior prognóstico em ADE. Com isso, este estudo sugere a importância das enzimas do metabolismo lipídico: FASN e SCD1 para a manutenção tumoral de ADE. A compreensão do metabolismo lipídico pode ter um impacto na compreensão dos mecanismos de manutenção do ADE e ajudar na identificação de possíveis alvos terapêuticos ou marcadores moleculares.(AU)