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Mortality rates due to lung cancer are high worldwide. Although PD-1 and PD-L1 immune checkpoint inhibitors boost the survival of patients with non-small-cell lung cancer (NSCLC), resistance often arises. The Warburg Effect, which causes lactate build-up and potential lysine-lactylation (Kla), links immune dysfunction to tumor metabolism. The role of non-histone Kla in tumor immune microenvironment and immunotherapy remains to be clarified. Here, global lactylome profiling and metabolomic analyses of samples from patients with NSCLC is conducted. By combining multi-omics analysis with in vitro and in vivo validation, that intracellular lactate promotes extracellular lipolysis through lactyl-APOC2 is revealed. Mechanistically, lactate enhances APOC2 lactylation at K70, stabilizing it and resulting in FFA release, regulatory T cell accumulation, immunotherapy resistance, and metastasis. Moreover, the anti-APOC2K70-lac antibody that sensitized anti-PD-1 therapy in vivo is developed. This findings highlight the potential of anti lactyl-APOC2-K70 approach as a new combination therapy for sensitizing immunotherapeutic responses.
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The regulation of triglyceride (TG) tissue distribution, storage, and utilization, a fundamental process of energy homeostasis, critically depends on lipoprotein lipase (LPL). We review the intricate mechanisms by which LPL activity is regulated by angiopoietin-like proteins (ANGPTL3, 4, 8), apolipoproteins (APOA5, APOC3, APOC2), and the cAMP-responsive element-binding protein H (CREBH). ANGPTL8 functions as a molecular switch, through complex formation, activating ANGPTL3 while deactivating ANGPTL4 in their LPL inhibition. The ANGPTL3-4-8 model integrates the roles of the aforementioned proteins in TG partitioning between white adipose tissue (WAT) and oxidative tissues (heart and skeletal muscles) during the feed/fast cycle. This model offers a unified perspective on LPL regulation, providing insights into TG metabolism, metabolic diseases, and therapeutics.
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Lipoproteína Lipasa , Humanos , Lipoproteína Lipasa/metabolismo , Animales , Triglicéridos/metabolismo , Proteínas Similares a la Angiopoyetina/metabolismo , Proteínas Similares a la Angiopoyetina/genética , Proteína 8 Similar a la Angiopoyetina , Proteína 4 Similar a la Angiopoyetina/metabolismo , Proteína 4 Similar a la Angiopoyetina/genética , Proteína 3 Similar a la Angiopoyetina/metabolismoRESUMEN
BACKGROUND AND AIM: APO CII, one of several cofactors which regulate lipoprotein lipase enzyme activity, plays an essential role in lipid metabolism. Deficiency of APO CII is an ultra-rare autosomal recessive cause of familial chylomicronemia syndrome. We present the long-term clinical outcomes of 12 children with APO CII deficiency. METHODS AND RESULTS: The data of children with genetically confirmed APO CII deficiency were evaluated retrospectively. Twelve children (8 females) with a mean follow-up of 10.1 years (±3.9) were included. At diagnosis, the median age was 60 days (13 days-10 years). Initial clinical findings included lipemic serum (41.6%), abdominal pain (41.6%), and vomiting (16.6%). At presentation, the median triglyceride (TG) value was 4341 mg/dL (range 1277-14,110). All patients were treated with a restricted fat diet, medium-chain triglyceride (MCT), and omega-3-fatty acids. In addition, seven patients (58.3%) received fibrate. Fibrate was discontinued in two patients due to rhabdomyolysis and in one patient because of cholelithiasis. Seven (58.3%) patients experienced pancreatitis during the follow-up period. One female experienced recurrent pancreatitis and was treated with fresh frozen plasma (FFP). CONCLUSIONS: Apo CII deficiency is an ultra-rare autosomal recessive condition of hypertriglyceridemia associated with significant morbidity and mortality. Low-fat diet and MCT supplementation are the mainstays of therapy, while the benefit of TG-lowering agents are less well-defined.
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Hiperlipoproteinemia Tipo I , Niño , Preescolar , Femenino , Humanos , Lactante , Recién Nacido , Masculino , Factores de Edad , Apolipoproteína C-II/genética , Apolipoproteína C-II/deficiencia , Apolipoproteína C-II/sangre , Biomarcadores/sangre , Dieta con Restricción de Grasas , Ácidos Fíbricos/uso terapéutico , Predisposición Genética a la Enfermedad , Hiperlipoproteinemia Tipo I/sangre , Hiperlipoproteinemia Tipo I/genética , Hiperlipoproteinemia Tipo I/terapia , Hipolipemiantes/uso terapéutico , Fenotipo , Estudios Retrospectivos , Factores de Riesgo , Factores de Tiempo , Resultado del Tratamiento , Triglicéridos/sangreRESUMEN
Introduction: Defects in lipolysis can lead to hypertriglyceridemia, which can trigger acute pancreatitis and is also associated with cardiovascular disease. Decreasing plasma triglycerides (TGs) by activating lipoprotein lipase (LPL) with ApoC2 mimetic peptides is a new treatment strategy for hypertriglyceridemia. We recently described a dual ApoC2 mimetic/ApoC3 antagonist peptide called D6PV that effectively lowered TG in several mouse models but has limitations in terms of drug development. The aim of this study was to create the next generation of ApoC2 mimetic peptides. Methods: We employed hydrocarbon staples, as well as select amino acid substitutions, to make short single helical mimetic peptides based on the last helix of ApoC2. Peptides were first tested for their ability to activate LPL and then in hypertriglyceridemia mouse models. All-atom simulations of peptides were performed in a lipid-trilayer model of TG-rich lipoproteins to discern their possible mechanism of action. Results: We designed a single stapled peptide called SP1 (21 residues), and a double stapled (stitched) peptide called SP2 (21 residues) and its N-terminal acylated analogue, SP2a. The hydrocarbon staples increased the amphipathicity of the peptides and their ability to bind lipids without interfering with LPL activation. Indeed, from all-atom simulations, the conformations of SP1 and SP2a are restrained by the staples and maintains the proper orientation of the LPL activation motif, while still allowing their deeper insertion into the lipid-trilayer model. Intraperitoneal injection of stapled peptides (1-5â umoles/kg) into ApoC2-hypomorphic mice or human ApoC3-transgenic resulted in an 80%-90% reduction in plasma TG within 3â h, similar to the much longer D6PV peptide (41 residues). Other modifications (replacement L-Glu20, L-Glu21 with their D-isomers, N-methylation of Gly19, Met2NorLeu and Ala1alpha-methylAla substitutions, N-terminal octanoylation) were introduced into the SP2a peptide. These changes made SP2a highly resistant to proteolysis against trypsin, pepsin, and Proteinase K, while maintaining similar efficacy in lowering plasma TG in mice. Conclusion: We describe a new generation of ApoC2 mimetic peptides based on hydron carbon stapling that are at least equally potent to earlier peptides but are much shorter and resistant to proteolysis and could be further developed into a new therapy for hypertriglyceridemia.
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The lipolytic processing of triglyceride-rich lipoproteins (TRLs) by lipoprotein lipase (LPL) is crucial for the delivery of dietary lipids to the heart, skeletal muscle, and adipose tissue. The processing of TRLs by LPL is regulated in a tissue-specific manner by a complex interplay between activators and inhibitors. Angiopoietin-like protein 4 (ANGPTL4) inhibits LPL by reducing its thermal stability and catalyzing the irreversible unfolding of LPL's α/ß-hydrolase domain. We previously mapped the ANGPTL4 binding site on LPL and defined the downstream unfolding events resulting in LPL inactivation. The binding of LPL to glycosylphosphatidylinositol-anchored high-density lipoprotein-binding protein 1 protects against LPL unfolding. The binding site on LPL for an activating cofactor, apolipoprotein C2 (APOC2), and the mechanisms by which APOC2 activates LPL have been unclear and controversial. Using hydrogen-deuterium exchange/mass spectrometry, we now show that APOC2's C-terminal α-helix binds to regions of LPL surrounding the catalytic pocket. Remarkably, APOC2's binding site on LPL overlaps with that for ANGPTL4, but their effects on LPL conformation are distinct. In contrast to ANGPTL4, APOC2 increases the thermal stability of LPL and protects it from unfolding. Also, the regions of LPL that anchor the lid are stabilized by APOC2 but destabilized by ANGPTL4, providing a plausible explanation for why APOC2 is an activator of LPL, while ANGPTL4 is an inhibitor. Our studies provide fresh insights into the molecular mechanisms by which APOC2 binds and stabilizes LPL-and properties that we suspect are relevant to the conformational gating of LPL's active site.
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Lipoproteína Lipasa , Lipoproteína Lipasa/metabolismo , Proteína 4 Similar a la Angiopoyetina/metabolismo , Apolipoproteína C-II , Dominios Proteicos , Dominio Catalítico , TriglicéridosRESUMEN
Genetic variations in APOC2 and APOA5 genes involve activating lipoprotein lipase (LPL), responsible for the hydrolysis of triglycerides (TG) in blood and whose impaired functions affect the TG metabolism and are associated with metabolic diseases. In this study, we investigate the biological significance of genetic variations at the DNA sequence and structural level using various computational tools. Subsequently, 8 (APOC2) and 17 (APOA5) non-synonymous SNPs (nsSNPs) were identified as high-confidence deleterious SNPs based on the effects of the mutations on protein conservation, stability, and solvent accessibility. Furthermore, based on our docking results, the interaction of native and mutant forms of the corresponding proteins with LPL depicts differences in root mean square deviation (RMSD), and binding affinities suggest that these mutations may affect their function. Furthermore, in vivo, and in vitro studies have shown that differential expression of these genes in disease conditions due to the influence of nsSNPs abundance may be associated with promoting the development of cancer and cardiovascular diseases. Preliminary screening using computational methods can be a helpful start in understanding the effects of mutations in APOC2 and APOA5 on lipid metabolism; however, further wet-lab experiments would further strengthen the conclusions drawn from the computational study.
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Enfermedades Cardiovasculares , Neoplasias , Humanos , Apolipoproteína A-V/genética , Apolipoproteína C-II/genética , Enfermedades Cardiovasculares/genética , Polimorfismo de Nucleótido Simple , Proteínas PortadorasRESUMEN
Adipogenesis is a complex and precisely orchestrated process mediated by a series of adipogenic regulatory factors. Recent studies have highlighted the importance of microRNAs (miRNAs) in diverse biological processes, most specifically in regulating cell differentiation and proliferation. However, the mechanisms of miRNAs in adipogenesis are largely unknown. In this study, we found that miR-107 expression was higher in bovine adipose tissue than that in other tissues, and there was a downregulation trend during adipocyte differentiation. To explore the function of miR-107 in adipocyte differentiation, agomiR-107 and antiagomiR-107 were transfected into bovine adipocytes, respectively. Oil Red O staining, CCK-8, EdU assays, RT-qPCR, and Western blotting were performed, and the results showed that overexpressed miR-107 significantly suppressed fat deposition and adipocyte differentiation, while knockdown of miR-107 promoted fat deposition and adipocytes differentiation. In addition, through bioinformatics analysis, luciferase reporter assays, RT-qPCR, and Western blotting, we identified apolipoprotein 2 (APOC2) as a target of miR-107. Transfection of siRNA-APOC2 into adipocytes led to suppression in adipocyte differentiation and proliferation, suggesting a positive role of APOC2 in bovine lipogenesis. In summary, our findings suggested that miR-107 regulates bovine adipocyte differentiation and lipogenesis by directly targeting APOC2, and these results. These theoretical and experimental basis for future clarification of the regulation mechanism of adipocyte differentiation and lipogenesis. Moreover, for the highly conserved among different species, miR-107 may be a potential molecular target to be used for the treatment of lipid-related diseases in the future.
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Adipogénesis , MicroARNs , Adipocitos/metabolismo , Adipogénesis/genética , Animales , Apolipoproteína C-II/metabolismo , Bovinos , Diferenciación Celular/genética , MicroARNs/genética , MicroARNs/metabolismoRESUMEN
Objective: Coronary heart disease (CHD) is a complex disease caused by multifaceted interaction between genetic and environmental factors, which makes identification of the most likely disease candidate proteins and their associated risk markers a big challenge. Atherosclerosis is presented by a broad spectrum of heart diseases, including stable coronary artery disease (SCAD) and acute myocardial infarction (AMI), which is the progressive stage of SCAD. As such, the correct and prompt diagnosis of atherosclerosis turns into imperative for precise and prompt disease diagnosis, treatment and prognosis. Methods: The current work aims to look for specific protein markers for differential diagnosis of coronary atherosclerosis. Thirty male patients between 45 and 55 years diagnosed with atherosclerosis were analyzed by tandem mass tag (TMT) mass spectrometry. The study excluded those who were additionally diagnosed with hypertension and type 1 and 2 diabetes. The Mufuzz analysis was applied to select target proteins for precise and prompt diagnosis of atherosclerosis, most of which were most related to high lipid metabolism. The parallel reaction monitoring (PRM) was used to verify the selected target proteins. Finally, The receiver operating characteristic curve (ROC) was calculated by a random forest experiment. Results: One thousand one hundred and forty seven proteins were identified in the TMT mass spectrometry, 907 of which were quantifiable. In the PRM study, six proteins related to lipid metabolism pathway were selected for verification and they were ALB, SHBG, APOC2, APOC3, APOC4, SAA4. Conclusion: Through the detected specific changes in these six proteins, our results provide accuracy in atherosclerosis patients' diagnosis, especially in cases with varying types of the disease.
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BACKGROUND: Peritoneal metastasis (PM) occurs frequently in patients with gastric cancer (GC) and confers poor survival. Lipid metabolism acts as a non-negligible regulator in epithelial-mesenchymal transition (EMT), which is crucial for the metastasis of GC. As apolipoprotein C2 (APOC2) is a key activator of lipoprotein lipase for triglyceride metabolism, the exact mechanism of APOC2 remains largely unknown in GC. METHODS: Tandem mass tags identified differentially expressed proteins between human PM and GC tissues, and showed that APOC2 overexpressed in PM tissues, which was further confirmed by immunoblotting, immunohistochemistry, and ELISA. Global gene expression changes were identified in APOC2 knockdown cells via RNA-sequencing. The role of APOC2 in lipid metabolism of GC cells was assessed via the Seahorse XF analyzer and lipid staining assays. The biological role of APOC2 in GC cells was determined by 3D Spheroid invasion, apoptosis, colony formation, wound healing, transwell assay, and mouse models. The interaction between APOC2 and CD36 was analyzed by co-immunoprecipitation and biolayer interferometry. The underlying mechanisms were investigated using western blot technique. RESULTS: APOC2 overexpressed in GC PM tissues. Upregulation of APOC2 correlated with a poor prognosis in GC patients. APOC2 promoted GC cell invasion, migration, and proliferation via CD36-mediated PI3K/AKT/mTOR signaling activation. Furthermore, APOC2-CD36 axis upregulated EMT markers of GC cells via increasing the phosphorylation of PI3K, AKT, and mTOR. Knockdown either APOC2 or CD36 inhibited the malignant phenotype of cancer cells, and delayed GC PM progression in murine GC models. CONCLUSION: APOC2 cooperates with CD36 to induce EMT to promote GC PM via PI3K/AKT/mTOR pathway. APOC2-CD36 axis may be a potential target for the treatment of aggressive GC.
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Apolipoproteína C-II/metabolismo , Transición Epitelial-Mesenquimal/genética , Neoplasias Peritoneales/genética , Neoplasias Peritoneales/secundario , Neoplasias Gástricas/genética , Neoplasias Gástricas/patología , Serina-Treonina Quinasas TOR/metabolismo , Animales , Apolipoproteína C-II/genética , Regulación Neoplásica de la Expresión Génica/genética , Humanos , Masculino , Ratones , Ratones Endogámicos BALB C , Transducción de Señal , Serina-Treonina Quinasas TOR/genéticaAsunto(s)
Apolipoproteína C-II/genética , Apolipoproteína C-II/metabolismo , Biomarcadores de Tumor/análisis , Metilación de ADN , Regulación Neoplásica de la Expresión Génica , Neoplasias/patología , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/metabolismo , Estudios de Casos y Controles , Humanos , Neoplasias/genética , Neoplasias/metabolismo , Pronóstico , Tasa de Supervivencia , Células Tumorales CultivadasRESUMEN
Familial chylomicronemia is caused by deficiency of lipoprotein lipase or its co-activators. Here, we report an infant with apolipoprotein C-II (APOC2) deficiency, who developed acute pancreatitis 37 days after birth. He presented as abdominal sepsis with fever, irritability and abdominal distention. Amylase levels were low, but lipase levels and imaging findings were consistent with acute pancreatitis. He had severe hypertriglyceridemia (1091 mg/dl). Keeping him nil orally for two days resulted in rapid decrease in triglyceride levels and resolution of the clinical findings. APOC2 gene sequencing revealed a homozygous splice-site mutation (c.55+1G>C). To the best of our knowledge, this patient is not only the youngest reported patient with APOC2 deficiency, but also the youngest such patient who developed pancreatitis. Although he had a severe presentation, invasive methods to treat hypertriglyceridemia were not necessary. We emphasize that clinical findings and amylase levels are not reliable to diagnose pancreatitis in this age group.
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Hiperlipoproteinemia Tipo I/complicaciones , Hiperlipoproteinemia Tipo I/genética , Pancreatitis/etiología , Predisposición Genética a la Enfermedad , Humanos , Lactante , Masculino , Mutación , Pancreatitis/terapiaRESUMEN
Dysregulation of microRNAs (miRNAs) expression has been demonstrated in gastrointestinal stromal tumor (GIST). In this study, we aimed to determine the differential miRNAs expression in GISTs and explore the functional mechanism of differential miRNAs in GIST cells. We measured differential miRNAs in six pairs of GIST tissues and matched adjacent tissues through a high-throughput sequencing, which was confirmed in 64 pairs of GIST tissues and adjacent tissues by real-time polymerase chain reaction. We found that miR-4510 expression was significantly downregulated in GIST tissues compared to matched control tissues. Luciferase reporter assay identified apolipoprotein C-II (APOC2) as a direct target of miR-4510. Overexpression of miR-4510 inhibited the mRNA and protein expression of APOC2. In addition, overexpression of miR-4510 suppressed GIST cell proliferation, migration, and invasion. Overexpression of miR-4510 also inhibited the phosphorylation of AKT and ERK1/2, reduced the expression of matrix metallopeptidase 2 (MMP2) and MMP9. APOC2 knockdown mimicked the effect of miR-4510 overexpression. Further investigation confirmed that APOC2 was notably upregulated in GIST tissues compared to adjacent control tissues. These results suggested that miR-4510 downregulation could promote GIST progression, including growth, invasion, and metastasis, through increasing APOC2 expression.
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Apolipoproteína C-II/genética , Tumores del Estroma Gastrointestinal/genética , Genes Supresores de Tumor , MicroARNs/genética , Proliferación Celular , Femenino , Tumores del Estroma Gastrointestinal/patología , Regulación Neoplásica de la Expresión Génica/genética , Humanos , Masculino , Persona de Mediana EdadRESUMEN
Background Familial apo C-II deficiency is a rare hereditary disorder frequently caused by lipoprotein lipase (LPL) and APOC2 gene mutations. To date, less than 30 patients with familial apo C-II deficiency with 24 different mutations have been identified in the literature. Here, we describe two familial chylomicronemia syndrome cases in infants with two novel mutations of the APOC2 gene. Case presentation Case 1, a 46-day-old female, was admitted to our hospital for evaluation due to the lipemic appearance of the blood sample. A clinical examination revealed hepatomegaly and lipemia retinalis. Triglyceride level of 6295 mg/dL was decreased with a strict low-fat diet, medium-chain triglycerides (MCT) oil-rich formula and omega-3 fatty acid supplementation. Due to low adherence to the diet, TG elevation was detected and fresh frozen plasma (10 mL/kg/day) was administered for 2 days. A novel homozygous p.Q25X (c.73C>T) mutation in the APOC2 gene was detected. Case 2, a 10-month-old female patient, referred to our center for the differential diagnosis of hyperlipidemia as her blood sample could not be assessed due to its lipemic appearance. Laboratory examinations showed a TG level of 4520 mg/dL which was reduced with a low-fat diet, MCT oil-rich formula and omega-3 fatty acid supplementation. Hepatosteatosis and splenomegaly were determined using abdominal sonography. A novel homozygous IVS2+6T>G (c.55+6T>G) mutation in the APOC2 gene was identified. Conclusions We describe two novel homozygous mutations (p.Q25X [c.73C>T] and IVS2+6T>G [c.55+6T>G]) in the APOC2 gene in infants with hyperchylomicronemia. To the best of our knowledge, Case 1 is the youngest patient with familial apo C-II deficiency in the literature to date.
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Apolipoproteína C-II/genética , Hiperlipoproteinemia Tipo I/genética , Mutación , Femenino , Humanos , Lactante , LinajeRESUMEN
Apolipoprotein C-II (APOC2) is an obligatory activator of lipoprotein lipase. Human patients with APOC2 deficiency display severe hypertriglyceridemia while consuming a normal diet, often manifesting xanthomas, lipemia retinalis and pancreatitis. Hypertriglyceridemia is also an important risk factor for development of cardiovascular disease. Animal models to study hypertriglyceridemia are limited, with no Apoc2-knockout mouse reported. To develop a genetic model of hypertriglyceridemia, we generated an apoc2 mutant zebrafish characterized by the loss of Apoc2 function. apoc2 mutants show decreased plasma lipase activity and display chylomicronemia and severe hypertriglyceridemia, which closely resemble the phenotype observed in human patients with APOC2 deficiency. The hypertriglyceridemia in apoc2 mutants is rescued by injection of plasma from wild-type zebrafish or by injection of a human APOC2 mimetic peptide. Consistent with a previous report of a transient apoc2 knockdown, apoc2 mutant larvae have a minor delay in yolk consumption and angiogenesis. Furthermore, apoc2 mutants fed a normal diet accumulate lipid and lipid-laden macrophages in the vasculature, which resemble early events in the development of human atherosclerotic lesions. In addition, apoc2 mutant embryos show ectopic overgrowth of pancreas. Taken together, our data suggest that the apoc2 mutant zebrafish is a robust and versatile animal model to study hypertriglyceridemia and the mechanisms involved in the pathogenesis of associated human diseases.