RESUMEN
The study investigates the therapeutic effects and mechanisms of ginsenoside Rg_1(GRg_1) on sepsis-induced acute lung injury(SALI). A murine model of SALI was created using cecal ligation and puncture(CLP) surgery, and mice were randomly assigned to groups for GRg_1 intervention. Survival and body weight changes were recorded, lung function was assessed with a non-invasive lung function test system, and lung tissue damage was evaluated through HE staining. The content and expression of inflammatory factors were measured by ELISA and qRT-PCR. Apoptosis was examined using flow cytometry and TUNEL staining. The activation and expression of apoptosis-related molecules cysteinyl aspartate specific proteinase 3(caspase-3), B-cell lymphoma-2(Bcl-2), Bcl-2 associated X protein(Bax), and endoplasmic reticulum stress-related molecules protein kinase R-like endoplasmic reticulum kinase(PERK), eukaryotic initiation factor 2α(eIF2α), activating transcription factor 4(ATF4), and C/EBP homologous protein(CHOP) were studied using Western blot and qRT-PCR. In addition, an in vitro model of lipopolysaccharide(LPS)-induced lung alveolar epithelial cell injury was used, with the application of the endoplasmic reticulum stress inducer tunicamycin to validate the action mechanism of GRg_1. RESULTS:: indicated that, when compared to the model group, GRg_1 intervention significantly enhanced the survival time of CLP mice, mitigated body weight loss, and improved impaired lung function indices. The GRg_1-treated mice also displayed reduced lung tissue pathological scores, a reduced lung tissue wet-to-dry weight ratio, and lower protein content in the bronchoalveolar lavage fluid. Serum levels of interleukin-6(IL-6), interleukin-1ß(IL-1ß), and tumor necrosis factor-α(TNF-α), as well as the mRNA expressions of these cytokines in lung tissues, were decreased. There was a notable decrease in the proportion of apopto-tic alveolar epithelial cells, and down-regulated expressions of caspase-3, Bax, PERK, eIF2α, ATF4, and CHOP and up-regulated expression of Bcl-2 were observed. In vitro findings showed that the apoptosis-lowering and apoptosis-related protein down-regulating effects of GRg_1 were significantly inhibited with the co-application of tunicamycin. Altogether, GRg_1 reduces apoptosis of alveolar epithelial cells, inhibits inflammation in the lungs, alleviates lung injury, and enhances lung function, possibly through the PERK/eIF2α/ATF4/CHOP pathway.
Asunto(s)
Factor de Transcripción Activador 4 , Lesión Pulmonar Aguda , Células Epiteliales Alveolares , Apoptosis , Factor 2 Eucariótico de Iniciación , Ginsenósidos , Sepsis , Factor de Transcripción CHOP , eIF-2 Quinasa , Animales , Lesión Pulmonar Aguda/tratamiento farmacológico , Lesión Pulmonar Aguda/metabolismo , Lesión Pulmonar Aguda/genética , Ginsenósidos/farmacología , Factor de Transcripción Activador 4/metabolismo , Factor de Transcripción Activador 4/genética , Ratones , Apoptosis/efectos de los fármacos , Factor de Transcripción CHOP/metabolismo , Factor de Transcripción CHOP/genética , Sepsis/tratamiento farmacológico , Sepsis/complicaciones , Sepsis/metabolismo , Sepsis/genética , eIF-2 Quinasa/metabolismo , eIF-2 Quinasa/genética , Factor 2 Eucariótico de Iniciación/metabolismo , Factor 2 Eucariótico de Iniciación/genética , Masculino , Células Epiteliales Alveolares/efectos de los fármacos , Células Epiteliales Alveolares/metabolismo , Humanos , Estrés del Retículo Endoplásmico/efectos de los fármacos , Ratones Endogámicos C57BLRESUMEN
Multiple myeloma is a hematological malignancy arising from immunoglobulin-secreting plasma cells. It remains poorly understood how chromatin rewiring of regulatory elements contributes to tumorigenesis and therapy resistance in myeloma. Here we generate a high-resolution contact map of myeloma-associated super-enhancers by integrating H3K27ac ChIP-seq and HiChIP from myeloma cell lines, patient-derived myeloma cells and normal plasma cells. Our comprehensive transcriptomic and phenomic analyses prioritize candidate genes with biological and clinical implications in myeloma. We show that myeloma cells frequently acquire SE that transcriptionally activate an oncogene PPP1R15B, which encodes a regulatory subunit of the holophosphatase complex that dephosphorylates translation initiation factor eIF2α. Epigenetic silencing or knockdown of PPP1R15B activates pro-apoptotic eIF2α-ATF4-CHOP pathway, while inhibiting protein synthesis and immunoglobulin production. Pharmacological inhibition of PPP1R15B using Raphin1 potentiates the anti-myeloma effect of bortezomib. Our study reveals that myeloma cells are vulnerable to perturbation of PPP1R15B-dependent protein homeostasis, highlighting a promising therapeutic strategy.
Asunto(s)
Regulación Neoplásica de la Expresión Génica , Mieloma Múltiple , Proteína Fosfatasa 1 , Proteostasis , Súper Potenciadores , Factor de Transcripción CHOP , Animales , Humanos , Factor de Transcripción Activador 4/metabolismo , Factor de Transcripción Activador 4/genética , Bortezomib/farmacología , Línea Celular Tumoral , Factor 2 Eucariótico de Iniciación/metabolismo , Factor 2 Eucariótico de Iniciación/genética , Mieloma Múltiple/genética , Mieloma Múltiple/metabolismo , Mieloma Múltiple/patología , Proteína Fosfatasa 1/metabolismo , Proteína Fosfatasa 1/genética , Súper Potenciadores/genética , Factor de Transcripción CHOP/metabolismo , Factor de Transcripción CHOP/genéticaRESUMEN
Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal and aggressive tumor that affects the digestive tract, leading to high mortality and poor survival rates. The purpose of the present study was to evaluate the expression levels of DNA damage-inducible transcript 3 (DDIT3) in pancreatic cancer and to investigate its effects in in vitro and in vivo experiments. Bioinformatics analysis indicated that DDIT3 expression was higher in pancreatic cancer tumor tissues and associated with a poor prognosis. Positive or strong positive DDIT3 expression was observed in PDAC, and no or weak expression was observed in normal pancreatic tissues. It was also highly expressed in PDAC cells, while being expressed at lower levels in normal pancreatic ductal epithelial cells. Transfection of short hairpin RNA targeting the DDIT3 gene reduced the proliferation, migration and invasion of PANC-1 cells. In vivo, in an in situ implantation tumor model with Pan02 cells, the size and weight of the tumors were reduced in the DDIT3 knockdown Pan02 cell-implanted group. These data suggested that DDIT3 represents a novel predictive biomarker for the potential treatment of patients presenting with PDAC.
Asunto(s)
Movimiento Celular , Proliferación Celular , Regulación Neoplásica de la Expresión Génica , Neoplasias Pancreáticas , Factor de Transcripción CHOP , Animales , Femenino , Humanos , Masculino , Ratones , Persona de Mediana Edad , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/patología , Carcinoma Ductal Pancreático/metabolismo , Línea Celular Tumoral , Movimiento Celular/genética , Proliferación Celular/genética , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patología , Neoplasias Pancreáticas/metabolismo , Pronóstico , Factor de Transcripción CHOP/metabolismo , Factor de Transcripción CHOP/genéticaRESUMEN
Parkinson's disease (PD) is a neurodegenerative disorder which affects dopaminergic neurons of the midbrain. Accumulation of α-synuclein or exposure to neurotoxins like 6-hydroxydopamine (6-OHDA) induces endoplasmic reticulum (ER) stress along with the unfolded protein response (UPR), which executes apoptosis via activation of PERK/CHOP or IRE1/JNK signaling. The present study aimed to determine which of these pathways is a major contributor to neurodegeneration in an 6-OHDA-induced in vitro model of PD. For this purpose, we have applied pharmacological PERK and JNK inhibitors (AMG44 and JNK V) in differentiated SH-SY5Y cells exposed to 6-OHDA. Inhibition of PERK and JNK significantly decreased genotoxicity and improved mitochondrial respiration, but only JNK inhibition significantly increased cell viability. Gene expression analysis revealed that the effect of JNK inhibition was dependent on a decrease in MAPK10 and XBP1 mRNA levels, whereas inhibition of either PERK or JNK significantly reduced the expression of DDIT3 mRNA. Western blot has shown that JNK inhibition strongly induced the XBP1s protein, and inhibition of each pathway attenuated the phosphorylation of eIF2α and JNK, as well as the expression of CHOP. Collectively, our data suggests that targeting the IRE1/JNK pathway of the UPR is a more effective option for PD treatment as it simultaneously affects more than one pro-apoptotic pathway.
Asunto(s)
Estrés del Retículo Endoplásmico , Endorribonucleasas , Oxidopamina , Proteínas Serina-Treonina Quinasas , Factor de Transcripción CHOP , Respuesta de Proteína Desplegada , eIF-2 Quinasa , Humanos , Apoptosis/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , eIF-2 Quinasa/metabolismo , Estrés del Retículo Endoplásmico/efectos de los fármacos , Endorribonucleasas/metabolismo , Endorribonucleasas/genética , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Proteína Quinasa 10 Activada por Mitógenos/metabolismo , Proteína Quinasa 10 Activada por Mitógenos/genética , Oxidopamina/farmacología , Enfermedad de Parkinson/metabolismo , Enfermedad de Parkinson/patología , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/genética , Transducción de Señal/efectos de los fármacos , Factor de Transcripción CHOP/metabolismo , Factor de Transcripción CHOP/genética , Respuesta de Proteína Desplegada/efectos de los fármacos , Proteína 1 de Unión a la X-Box/metabolismo , Proteína 1 de Unión a la X-Box/genéticaRESUMEN
Montelukast and zafirlukast, cysteinyl leukotriene receptor antagonists (LTRAs), trigger apoptosis and inhibit cell proliferation of triplenegative breast cancer MDAMB231 cells. By contrast, only zafirlukast induces G0/G1 cell cycle arrest. The present study compared the effects of these drugs on proteins regulating cell proliferation, apoptosis, autophagy, and endoplasmic reticulum (ER) and oxidative stress using reverse transcriptionquantitative PCR, western blotting and flow cytometry. The expression of proliferating markers, Ki67 and proliferating cell nuclear antigen, was decreased by both drugs. Zafirlukast, but not montelukast, decreased the expression of cyclin D1 and CDK4, disrupting progression from G1 to S phase. Zafirlukast also increased the expression of p27, a cell cycle inhibitor. Both drugs decreased the expression of antiapoptotic protein Bcl2 and ERK1/2 phosphorylation, and increased levels of the autophagy marker LC3II and DNA damage markers, including cleaved PARP1, phosphorylated (p)ATM and phistone H2AX. The number of caspase 3/7positive cells was greater in montelukasttreated cells compared with zafirlukasttreated cells. Montelukast induced higher levels of the ER stress marker CHOP compared with zafirlukast. Montelukast activated PERK, activating transcription factor 6 (ATF6) and inositolrequiring enzyme type 1 (IRE1) pathways, while zafirlukast only stimulated ATF6 and IRE1 pathways. GSK2606414, a PERK inhibitor, decreased apoptosis mediated by montelukast, but did not affect zafirlukastinduced cell death. The knockdown of CHOP by small interfering RNA reduced apoptosis triggered by montelukast and zafirlukast. In conclusion, the effects on cell cycle regulator proteins may contribute to cell cycle arrest caused by zafirlukast. The greater apoptotic effects of montelukast may be caused by the higher levels of activated caspase enzymes and the activation of three pathways of ER stress: PERK, ATF6, and IRE1.
Asunto(s)
Acetatos , Apoptosis , Autofagia , Ciclopropanos , Daño del ADN , Estrés del Retículo Endoplásmico , Indoles , Quinolinas , Sulfuros , Sulfonamidas , Humanos , Sulfuros/farmacología , Ciclopropanos/farmacología , Quinolinas/farmacología , Apoptosis/efectos de los fármacos , Acetatos/farmacología , Estrés del Retículo Endoplásmico/efectos de los fármacos , Línea Celular Tumoral , Autofagia/efectos de los fármacos , Sulfonamidas/farmacología , Indoles/farmacología , Femenino , Daño del ADN/efectos de los fármacos , Fenilcarbamatos/farmacología , Compuestos de Tosilo/farmacología , Proliferación Celular/efectos de los fármacos , eIF-2 Quinasa/metabolismo , eIF-2 Quinasa/genética , Endorribonucleasas/metabolismo , Endorribonucleasas/genética , Puntos de Control del Ciclo Celular/efectos de los fármacos , Factor de Transcripción CHOP/metabolismo , Factor de Transcripción CHOP/genética , Ciclo Celular/efectos de los fármacos , Antagonistas de Leucotrieno/farmacología , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/genéticaRESUMEN
Resveratrol (RSV) is a polyphenol antioxidant that has been shown to have neuroprotective effects. We sought molecular mechanisms that emphasize the anti-inflammatory activity of RSV in traumatic brain injury (TBI) in mice associated with endoplasmic reticulum stress (ERS). After establishing three experimental groups (sham, TBI, and TBI+RSV), we explored the results of RSV after TBI on ERS and caspase-12 apoptotic pathways. The expression levels of C/EBP homologous protein (CHOP), glucose regulated protein 78kD (GRP78), caspase-3, and caspase-12 in cortical brain tissues were assessed by western blotting. The qPCR analysis was also performed on mRNA expression of tumor necrosis factor (TNF)-α and interleukin (IL)-1ß in cortical brain tissue. In addition, the expression of GRP78 in microglia (ionized calcium binding adaptor molecule 1; Iba-1) and neurons (neuronal nuclei; NeuN) was identified by immunofluorescence staining. The neurological function of mice was assessed by modified neurological severity scores (mNSS). After drug treatment, the expression of CHOP, GRP78, caspase-3 and caspase-12 decreased, and qPCR results showed that TNF-α and IL-1ß were down-regulated. Immunofluorescence staining showed down-regulation of Iba-1+/GRP78+ and NeuN+/GRP78+ cells after RSV treatment. The mNSS analysis confirmed improvement after RSV treatment. RSV improved apoptosis by downregulating the ERS signaling pathway and improved neurological prognosis in mice with TBI.
Asunto(s)
Lesiones Traumáticas del Encéfalo , Chaperón BiP del Retículo Endoplásmico , Estrés del Retículo Endoplásmico , Resveratrol , Animales , Lesiones Traumáticas del Encéfalo/tratamiento farmacológico , Lesiones Traumáticas del Encéfalo/patología , Lesiones Traumáticas del Encéfalo/metabolismo , Resveratrol/farmacología , Resveratrol/uso terapéutico , Estrés del Retículo Endoplásmico/efectos de los fármacos , Ratones , Masculino , Apoptosis/efectos de los fármacos , Pronóstico , Fármacos Neuroprotectores/farmacología , Fármacos Neuroprotectores/uso terapéutico , Neuronas/efectos de los fármacos , Neuronas/patología , Neuronas/metabolismo , Interleucina-1beta/metabolismo , Interleucina-1beta/genética , Caspasa 12/metabolismo , Caspasa 12/genética , Proteínas de Choque Térmico/metabolismo , Proteínas de Choque Térmico/genética , Factor de Necrosis Tumoral alfa/metabolismo , Ratones Endogámicos C57BL , Muerte Celular/efectos de los fármacos , Microglía/efectos de los fármacos , Microglía/metabolismo , Microglía/patología , Factor de Transcripción CHOP/metabolismo , Factor de Transcripción CHOP/genéticaRESUMEN
Dihydropyrazines (DHPs) are formed by non-enzymatic glycation reactions in vivo and in food. We recently reported that 3-hydro-2,2,5,6-tetramethylpyrazine (DHP-3), which is a methyl-substituted DHP, caused severe oxidative stress and cytotoxicity. However, the molecular mechanisms underlying the cytotoxic pathways of the DHP response remain elusive. Because oxidative stress induces endoplasmic reticulum (ER) stress and autophagy, we investigated the ability of DHP-3 to modulate the ER stress and autophagy pathways. DHP-3 activated the ER stress pathway by increasing inositol-requiring enzyme 1 (IRE1) and PKR-like ER kinase (PERK) phosphorylation and transcription factor 6 (ATF6) expression. Moreover, DHP-3 increased the expression of activating transcription factor 4 (ATF4) and C/EBP homologous protein (CHOP), which are downstream targets of PERK. In addition, DHP-3 inhibited the autophagy pathway by increasing the accumulation of microtubule-associated protein 1 light chain 3 alpha-phosphatidylethanolamine conjugate (LC3-II) and p62/sequestosome 1 (p62), while decreasing autophagic flux. Taken together, these results indicate that DHP-3 activates the ER stress pathway and inhibits the autophagy pathway, suggesting that the resulting removal of damaged organelles is inadequate.
Asunto(s)
Factor de Transcripción Activador 4 , Factor de Transcripción Activador 6 , Autofagia , Estrés del Retículo Endoplásmico , Proteínas Serina-Treonina Quinasas , Pirazinas , eIF-2 Quinasa , Humanos , Autofagia/efectos de los fármacos , Estrés del Retículo Endoplásmico/efectos de los fármacos , Pirazinas/farmacología , Células Hep G2 , Factor de Transcripción Activador 4/metabolismo , Factor de Transcripción Activador 4/genética , eIF-2 Quinasa/metabolismo , Factor de Transcripción Activador 6/metabolismo , Factor de Transcripción Activador 6/genética , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/genética , Factor de Transcripción CHOP/metabolismo , Factor de Transcripción CHOP/genética , Endorribonucleasas/metabolismo , Endorribonucleasas/genética , Fosforilación , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patología , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patología , Estrés Oxidativo/efectos de los fármacos , Proteína Sequestosoma-1/metabolismo , Proteína Sequestosoma-1/genética , Transducción de Señal/efectos de los fármacos , Proteínas Asociadas a Microtúbulos/metabolismoRESUMEN
The therapeutic potential of blue light photobiomodulation in cancer treatment, particularly in inhibiting cell proliferation and promoting cell death, has attracted significant interest. Oral squamous cell carcinoma (OSCC) is a prevalent form of oral cancer, necessitating innovative treatment approaches to improve patient outcomes. In this study, we investigated the effects of 420 nm blue LED light on OSCC and explored the underlying mechanisms. Our results demonstrated that 420 nm blue light effectively reduced OSCC cell viability and migration, and induced G2/M arrest. Moreover, we observed that 420 nm blue light triggered endoplasmic reticulum (ER) stress and mitochondrial dysfunction in OSCC cells, leading to activation of the CHOP signal pathway and alterations in the levels of Bcl-2 and Bax proteins, ultimately promoting cell apoptosis. Additionally, blue light suppressed mitochondrial gene expression, likely due to its damage to mitochondrial DNA. This study highlights the distinct impact of 420 nm blue light on OSCC cells, providing valuable insights into its potential application as a clinical treatment for oral cancer.
Asunto(s)
Apoptosis , Carcinoma de Células Escamosas , Supervivencia Celular , Estrés del Retículo Endoplásmico , Luz , Mitocondrias , Neoplasias de la Boca , Humanos , Estrés del Retículo Endoplásmico/efectos de la radiación , Mitocondrias/efectos de la radiación , Mitocondrias/metabolismo , Neoplasias de la Boca/radioterapia , Neoplasias de la Boca/patología , Neoplasias de la Boca/metabolismo , Línea Celular Tumoral , Carcinoma de Células Escamosas/radioterapia , Carcinoma de Células Escamosas/patología , Carcinoma de Células Escamosas/metabolismo , Apoptosis/efectos de la radiación , Supervivencia Celular/efectos de la radiación , Proliferación Celular/efectos de la radiación , Movimiento Celular/efectos de la radiación , Transducción de Señal/efectos de la radiación , Proteína X Asociada a bcl-2/metabolismo , Proteína X Asociada a bcl-2/genética , Factor de Transcripción CHOP/metabolismo , Factor de Transcripción CHOP/genética , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/genética , Luz AzulRESUMEN
Methamphetamine (METH) is a widely abused amphetamine-type psychoactive drug that causes serious health problems. Previous studies have demonstrated that METH can induce neuron autophagy and apoptosis in vivo and in vitro. However, the molecular mechanisms underlying METH-induced neuron autophagy and apoptosis remain poorly understood. Stromal interacting molecule 1 (STIM1) was hypothesized to be involved in METH-induced neuron autophagy and apoptosis. Therefore, the expression of STIM1 protein was measured and the effect of blocking STIM1 expression with siRNA was investigated in cultured neuronal cells, and the hippocampus and striatum of mice exposed to METH. Furthermore, intracellular calcium concentration and endoplasmic reticulum (ER) stress-related proteins were determined in vitro and in vivo in cells treated with METH. The results suggested that STIM1 mediates METH-induced neuron autophagy by activating the p-Akt/p-mTOR pathway. METH exposure also resulted in increased expression of Orai1, which was reversed after STIM1 silencing. Moreover, the disruption of intracellular calcium homeostasis induced ER stress and up-regulated the expression of pro-apoptotic protein CCAAT/enhancer-binding protein homologous protein (CHOP), resulting in classic mitochondria apoptosis. METH exposure can cause neuronal autophagy and apoptosis by increasing the expression of STIM1 protein; thus, STIM1 may be a potential gene target for therapeutics in METH-caused neurotoxicity.
Asunto(s)
Apoptosis , Autofagia , Estrés del Retículo Endoplásmico , Metanfetamina , Neuronas , Molécula de Interacción Estromal 1 , Metanfetamina/toxicidad , Animales , Molécula de Interacción Estromal 1/metabolismo , Molécula de Interacción Estromal 1/genética , Autofagia/efectos de los fármacos , Apoptosis/efectos de los fármacos , Neuronas/efectos de los fármacos , Neuronas/patología , Neuronas/metabolismo , Estrés del Retículo Endoplásmico/efectos de los fármacos , Estrés del Retículo Endoplásmico/fisiología , Ratones , Ratones Endogámicos C57BL , Estimulantes del Sistema Nervioso Central/toxicidad , Calcio/metabolismo , Células Cultivadas , Masculino , Hipocampo/efectos de los fármacos , Hipocampo/metabolismo , Hipocampo/patología , Proteína ORAI1/metabolismo , Proteína ORAI1/genética , Serina-Treonina Quinasas TOR/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Factor de Transcripción CHOP/metabolismo , Factor de Transcripción CHOP/genética , Cuerpo Estriado/efectos de los fármacos , Cuerpo Estriado/metabolismo , Cuerpo Estriado/patologíaRESUMEN
Anticancer strategies using natural products or derivatives are promising alternatives for cancer treatment. Here, we showed that licochalcone D (LCD), a natural flavonoid extracted from Glycyrrhiza uralensis Fisch, suppressed the growth of breast cancer cells, and was less toxic to MCF-10A normal breast cells. LCD-induced DNA damage, cell cycle arrest, and apoptosis in breast cancer cells. Furthermore, LCD potentiated tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced cytotoxicity. Mechanistically, LCD was revealed to reduce survival protein expression and to upregulate death receptor 5 (DR5) expressions. Silencing DR5 blocked the ability of LCD to sensitize cells to TRAIL-mediated apoptosis. LCD increased CCAAT/enhancer-binding protein homologous protein (CHOP) expression in breast cancer cells. Knockdown of CHOP attenuated DR5 upregulation and apoptosis triggered by cotreatment with LCD and TRAIL. Furthermore, LCD suppressed the phosphorylation of extracellular signal-regulated kinase and promoted the phosphorylation of c-Jun amino-terminal kinase (JNK) and p38 mitogen-activated protein kinase (MAPK). Pretreatment with JNK inhibitor SP600125 or p38 MAPK inhibitor SB203580 abolished the upregulation of DR5 and CHOP, and also attenuated LCD plus TRAIL-induced cleavage of poly(ADP-ribose) polymerase. Overall, our results show that LCD exerts cytotoxic effects on breast cancer cells and arguments TRAIL-mediated apoptosis by inhibiting survival protein expression and upregulating DR5 in a JNK/p38 MAPK-CHOP-dependent manner.
Asunto(s)
Apoptosis , Neoplasias de la Mama , Chalconas , Receptores del Ligando Inductor de Apoptosis Relacionado con TNF , Ligando Inductor de Apoptosis Relacionado con TNF , Factor de Transcripción CHOP , Regulación hacia Arriba , Humanos , Receptores del Ligando Inductor de Apoptosis Relacionado con TNF/metabolismo , Receptores del Ligando Inductor de Apoptosis Relacionado con TNF/genética , Chalconas/farmacología , Ligando Inductor de Apoptosis Relacionado con TNF/farmacología , Ligando Inductor de Apoptosis Relacionado con TNF/metabolismo , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Neoplasias de la Mama/tratamiento farmacológico , Apoptosis/efectos de los fármacos , Femenino , Regulación hacia Arriba/efectos de los fármacos , Factor de Transcripción CHOP/metabolismo , Factor de Transcripción CHOP/genética , Línea Celular Tumoral , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Células MCF-7 , Sistema de Señalización de MAP Quinasas/efectos de los fármacosRESUMEN
Hepatocellular carcinoma (HCC) presents a persistent challenge to conventional therapeutic approaches. SLC12A5 is implicated in an oncogenic capacity and facilitates the progression of cancer. The objective of this investigation is to scrutinize the inhibitory effects of borax on endoplasmic reticulum (ER)-stress and apoptosis mediated by SLC12A5 in HepG2 cells. Initially, we evaluated the cytotoxic impact of borax on both HL-7702 and HepG2 cell lines. Subsequently, the effects of borax on cellular morphology and the cell cycle of these lines were examined. Following this, we explored the impact of borax treatment on the mRNA and protein expression levels of SLC12A5, C/EBP homologous protein (CHOP), glucose-regulated protein-78 (GRP78), activating transcription factor-6 (ATF6), caspase-3 (CASP3), and cytochrome c (CYC) in these cellular populations. The determined IC50 value of borax for HL-7702 cells was 40.8 mM, whereas for HepG2 cells, this value was 22.6 mM. The concentrations of IC50 (22.6 mM) and IC75 (45.7 mM) of borax in HepG2 cells did not manifest morphological aberrations in HL-7702 cells. Conversely, these concentrations in HepG2 cells induced observable morphological and nuclear abnormalities, resulting in cell cycle arrest in the G1/G0 phase. Additionally, the levels of SLC12A5, ATF6, CHOP, GRP78, CASP3, and CYC were elevated in HepG2 cells in comparison to HL-7702 cells. Moreover, SLC12A5 levels decreased following borax treatment in HepG2 cells, whereas ATF6, CHOP, GRP78, CASP3, and CYC levels exhibited a significant increase. In conclusion, our data highlight the potential therapeutic effects of borax through the regulation of ER stress in HCC by targeting SLC12A5.
Asunto(s)
Apoptosis , Carcinoma Hepatocelular , Supervivencia Celular , Chaperón BiP del Retículo Endoplásmico , Estrés del Retículo Endoplásmico , Neoplasias Hepáticas , Humanos , Estrés del Retículo Endoplásmico/efectos de los fármacos , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patología , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/tratamiento farmacológico , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patología , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/tratamiento farmacológico , Supervivencia Celular/efectos de los fármacos , Células Hep G2 , Apoptosis/efectos de los fármacos , Factor de Transcripción CHOP/metabolismo , Factor de Transcripción CHOP/genética , Proteínas de Choque Térmico/metabolismo , Proteínas de Choque Térmico/genética , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Factor de Transcripción Activador 6/metabolismo , Factor de Transcripción Activador 6/genética , Proliferación Celular/efectos de los fármacos , Ciclo Celular/efectos de los fármacosRESUMEN
Exosomes generated from mesenchymal stem cells (MSCs) are thought to be a unique therapeutic strategy for several autoimmune deficiency illnesses. The purpose of this study was to elucidate the protective effects of human umbilical cord mesenchymal stem cell-derived exosomes (hUCMSC-Exo) on CD4+ T cells dysfunction during graft-versus-host disease (GVHD) and to identify the underlying processes involved. Here, we showed that hUCMSC-Exo treatment can effectively attenuate GVHD injury by alleviating redox metabolism disorders and inflammatory cytokine bursts in CD4+ T cells. Furthermore, hUCMSC-Exo ameliorate ER stress and ATF6/CHOP signaling-mediated apoptosis in CD4+ T cells and promote the development of CD4+IL-10+ T cells during GVHD. Moreover, downregulating miR-16-5p in hUCMSC-Exo impaired their ability to prevent CD4+ T cells apoptosis and weakened their ability to promote the differentiation of CD4+IL-10+ T cells. Collectively, the obtained data suggested that hUCMSC-Exo suppress ATF6/CHOP signaling-mediated ER stress and apoptosis in CD4+ T cells, enhance the differentiation of CD4+IL-10+ T cells, and reverse the imbalance of immune homeostasis in the GVHD process by transferring miR-16-5p. Our study provided further evidence that GVHD patients can benefit from hUCMSC-Exo-mediated therapy.
Asunto(s)
Factor de Transcripción Activador 6 , Linfocitos T CD4-Positivos , Estrés del Retículo Endoplásmico , Exosomas , Enfermedad Injerto contra Huésped , Células Madre Mesenquimatosas , MicroARNs , Transducción de Señal , Factor de Transcripción CHOP , MicroARNs/metabolismo , MicroARNs/genética , Exosomas/metabolismo , Estrés del Retículo Endoplásmico/inmunología , Enfermedad Injerto contra Huésped/inmunología , Enfermedad Injerto contra Huésped/prevención & control , Humanos , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/metabolismo , Animales , Factor de Transcripción Activador 6/metabolismo , Factor de Transcripción Activador 6/genética , Células Madre Mesenquimatosas/metabolismo , Células Madre Mesenquimatosas/inmunología , Factor de Transcripción CHOP/metabolismo , Factor de Transcripción CHOP/genética , Apoptosis , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Cordón Umbilical/citología , Células CultivadasRESUMEN
HLA-B27 is a major risk factor for spondyloarthritis (SpA), yet the underlying mechanisms remain unclear. HLA-B27 misfolding-induced IL-23, which is mediated by endoplasmic reticulum (ER) stress has been hypothesized to drive SpA pathogenesis. Expression of HLA-B27 and human ß2m (hß2m) in rats (HLA-B27-Tg) recapitulates key SpA features including gut inflammation. Here we determined whether deleting the transcription factor CHOP (Ddit3-/-), which mediates ER-stress induced IL-23, affects gut inflammation in HLA-B27-Tg animals. ER stress-mediated Il23a overexpression was abolished in CHOP-deficient macrophages. Although CHOP-deficiency also reduced Il23a expression in immune cells isolated from the colon of B27+ rats, Il17a levels were not affected, and gut inflammation was not reduced. Rather, transcriptome analysis revealed increased expression of pro-inflammatory genes, including Il1a, Ifng and Tnf in HLA-B27-Tg colon tissue in the absence of CHOP, which was accompanied by higher histological Z-scores. RNAScope localized Il17a mRNA to the lamina propria of the HLA-B27-Tg rats and revealed similar co-localization with Cd3e (CD3) in the presence and absence of CHOP. This demonstrates that CHOP-deficiency does not improve, but rather exacerbates gut inflammation in HLA-B27-Tg rats, indicating that HLA-B27 is not promoting gut disease through ER stress-induced IL-23. Hence, CHOP may protect rats from more severe HLA-B27-induced gut inflammation.
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Colitis , Estrés del Retículo Endoplásmico , Antígeno HLA-B27 , Espondiloartritis , Factor de Transcripción CHOP , Animales , Antígeno HLA-B27/genética , Antígeno HLA-B27/metabolismo , Factor de Transcripción CHOP/metabolismo , Factor de Transcripción CHOP/genética , Colitis/metabolismo , Colitis/genética , Colitis/inducido químicamente , Colitis/patología , Ratas , Espondiloartritis/metabolismo , Espondiloartritis/patología , Espondiloartritis/genética , Modelos Animales de Enfermedad , Interleucina-23/metabolismo , Interleucina-23/genética , Humanos , Subunidad p19 de la Interleucina-23/genética , Subunidad p19 de la Interleucina-23/metabolismo , Ratas Transgénicas , Interleucina-17/metabolismo , Interleucina-17/genética , Colon/patología , Colon/metabolismo , Macrófagos/metabolismo , Macrófagos/inmunologíaRESUMEN
We investigated the relationship between neutrophil apoptosis and endoplasmic reticulum stress (ERS) in sepsis and its mechanism. A prospective cohort study was conducted by recruiting a total of 58 patients with sepsis. Peripheral blood samples were collected on 1, 3, 5 and 7 days after admission to the ICU. The expressions of endoplasmic reticulum specific glucose regulatory protein 78 (GRP78), C/EBP homologous protein (CHOP), apoptosis signal-regulating kinase 1 (ASK1), Bcl-2-like 11 (BIM), death receptor 5 (DR5), c-Jun N-terminal kinases (JNK) and p38 were detected by Western blot and PCR. The subcellular location of CHOP and GRP78 was observed by immunofluorescence analysis. Spearman correlation was used to analyze the correlation between the expression of chop protein and the apoptosis rate of peripheral blood neutrophils. Healthy volunteers in the same period were selected as the healthy control group. The expression of GRP78 protein was significantly elevated on the first day of ICU admission and showed a decreasing trend on the third, fifth and seventh day, but was significantly higher than the corresponding healthy control group. The expression of CHOP protein reached the highest level on the third day. The expression of chop protein in each group was significantly higher than that in the corresponding healthy control group. Immunofluorescence staining clearly showed that the CHOP protein accumulated in the nucleus, with an elevation in the intensity of GRP78. The neutrophil apoptosis rate of sepsis patients on the 1st, 3rd, 5th and 7th day of ICU stay was significantly higher than that of the healthy control group, with the highest apoptosis rate on the 3rd day, and then decreased gradually. CHOP protein expression level was significantly positively correlated with neutrophil apoptosis rate in sepsis patients. Endoplasmic reticulum stress occurs in neutrophils during the development of sepsis. GRP78 protein and CHOP protein may be involved in the pathological process of neutrophil apoptosis in sepsis.
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Apoptosis , Chaperón BiP del Retículo Endoplásmico , Retículo Endoplásmico , Neutrófilos , Sepsis , Factor de Transcripción CHOP , Adulto , Anciano , Femenino , Humanos , Masculino , Persona de Mediana Edad , Retículo Endoplásmico/metabolismo , Retículo Endoplásmico/patología , Retículo Endoplásmico/genética , Estrés del Retículo Endoplásmico , Regulación de la Expresión Génica , Neutrófilos/metabolismo , Neutrófilos/patología , Estudios Prospectivos , Sepsis/patología , Sepsis/metabolismo , Sepsis/genética , Factor de Transcripción CHOP/metabolismo , Factor de Transcripción CHOP/genéticaRESUMEN
Thermal processing of food is likely to form acrylamide (AA) and elaidic acid (EA), which are both mainly metabolized by the liver. The two substances are associated with the pathogenesis of liver disease. In the current study, we investigated the toxic effects of the combined action of AA and EA on HSC-T6 cells, and the mechanism of apoptosis exacerbated by the co-exposure. The results showed a synergistic effect of AA and EA, which exacerbated the damage and oxidative stress (OS) in HSC-T6. Meanwhile, the expression of endoplasmic reticulum stress (ERS) proteins, such as GRP78 and CHOP, was increased, the ERS pathway was activated, and Ca2+ in cells was increased, which exacerbated mitochondrial damage, and opened IP3R-Grp75-VDAC1 channel. Both ERS and mitochondrial damage caused the process of cell apoptosis. Inhibition of ERS by 4-phenylbutyric acid (4-PBA) significantly reversed the synergistic effects on mitochondrial damage via ERS, suggesting that AA and EA exacerbated mitochondrial damage through ERS-mediated Ca2+ overload. AA and EA synergistically damaged the function of mitochondria through exacerbating ERS and led to cell apoptosis.
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Acrilamida , Apoptosis , Estrés del Retículo Endoplásmico , Ácidos Oléicos , Apoptosis/efectos de los fármacos , Estrés del Retículo Endoplásmico/efectos de los fármacos , Acrilamida/toxicidad , Animales , Línea Celular , Ácidos Oléicos/farmacología , Estrés Oxidativo/efectos de los fármacos , Calcio/metabolismo , Ratas , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Factor de Transcripción CHOP/metabolismo , Factor de Transcripción CHOP/genética , Sinergismo Farmacológico , Chaperón BiP del Retículo Endoplásmico , FenilbutiratosRESUMEN
This study investigated the protective effect of ginsenoside Rg_1(GRg_1) on oxygen and glucose deprivation/reoxygenation(OGD/R)-injured rat adrenal pheochromocytoma(PC12) cells and whether the underlying mechanism was related to the regulation of inositol-requiring enzyme 1(IRE1)-c-Jun N-terminal kinase(JNK)-C/EBP homologous protein(CHOP) signaling pathway. An OGD/R model was established in PC12 cells, and PC12 cells were randomly classified into control, model, OGD/R+GRg_1(0.1, 1, 10 µmol·L~(-1)), OGD/R+GRg_1+rapamycin(autophagy agonist), OGD/R+GRg_1+3-methyladenine(3-MA,autophagy inhibitor), OGD/R+GRg_1+tunicamycin(endoplasmic reticulum stress agonist), OGD/R+GRg_1+4-phenylbutyric acid(4-PBA, endoplasmic reticulum stress inhibitor), and OGD/R+GRg_1+3,5-dibromosalicylaldehyde(DBSA, IRE1 inhibitor) groups. Except the control group, the other groups were subjected to OGD/R treatment, i.e., oxygen and glucose deprivation for 6 h followed by reoxygenation for 6 h. Cell viability was detected by the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl tetrazolium bromide(MTT) assay. Apoptosis was detected by Hoechst 33342 staining, and the fluorescence intensity of autophagosomes by the monodansylcadaverine(MDC) assay. Western blot was employed to determine the expression of autophagy-related proteins(Beclin1, LC3-â ¡, and p62) and the pathway-related proteins [IRE1, p-IRE1, JNK, p-JNK, glucose-regulated protein 78(GRP78), and CHOP]. The results showed that GRg_1 dose-dependently increased the viability of PC12 cells and down-regulated the expression of Beclin1, LC3-â ¡, p-IRE1, p-JNK, GRP78, and CHOP, compared with the model group. Furthermore, GRg_1 decreased the apoptosis rate and MDC fluorescence intensity and up-regulated the expression of p62 protein. Compared with the OGD/R+GRg_1(10 µmol·L~(-1)) group, OGD/R+GRg_1+rapamycin and OGD/R+GRg_1+tunicamycin groups showed increased apoptosis rate and MDC fluorescence intensity, up-regulated protein levels of Beclin1, LC3-â ¡, p-IRE1, p-JNK, GRP78, and CHOP, decreased relative cell survival rate, and down-regulated protein level of p62. The 3-MA, 4-PBA, and DBSA groups exerted the opposite effects. Taken together, GRg_1 may ameliorate OGD/R-induced PC12 cell injury by inhibiting autophagy via the IRE1-JNK-CHOP pathway.
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Apoptosis , Ginsenósidos , Glucosa , Proteínas Serina-Treonina Quinasas , Factor de Transcripción CHOP , Animales , Ratas , Células PC12 , Factor de Transcripción CHOP/metabolismo , Factor de Transcripción CHOP/genética , Glucosa/metabolismo , Ginsenósidos/farmacología , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/genética , Apoptosis/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Autofagia/efectos de los fármacos , Endorribonucleasas/metabolismo , Endorribonucleasas/genética , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , Proteínas Quinasas JNK Activadas por Mitógenos/genética , Oxígeno/metabolismo , Estrés del Retículo Endoplásmico/efectos de los fármacos , Complejos MultienzimáticosRESUMEN
OBJECTIVE: DNA damage-inducible transcript 3 (DDIT3), as a downstream transcription factor of endoplasmic reticulum stress, is reported to regulate chondrogenic differentiation under physiological and pathological state. However, the specific involvement of DDIT3 in the degradation of condylar cartilage of temporomandibular joint osteoarthritis (TMJOA) is unclarified. DESIGN: The expression patterns of DDIT3 in condylar cartilage from monosodium iodoacetate-induced TMJOA mice were examined to uncover the potential role of DDIT3 in TMJOA. The Ddit3 knockout (Ddit3-/-) mice and their wildtype littermates (Ddit3+/+) were used to clarify the effect of DDIT3 on cartilage degradation. Primary condylar chondrocytes and ATDC5 cells were applied to explore the mechanisms of DDIT3 on autophagy and extracellular matrix (ECM) degradation in chondrocytes. The autophagy inhibitor chloroquine (CQ) was used to determine the effect of DDIT3-inhibited autophagy in vivo. RESULTS: DDIT3 were highly expressed in condylar cartilage from TMJOA mice. Ddit3 knockout alleviated condylar cartilage degradation and subchondral bone loss, compared with their wildtype littermates. In vitro study demonstrated that DDIT3 exacerbated ECM degradation in chondrocytes induced by TNF-α through inhibiting autophagy. The intraperitoneal injection of CQ further confirmed that Ddit3 knockout alleviated cartilage degradation in TMJOA through activating autophagy in vivo. CONCLUSIONS: Our findings identified the crucial role of DDIT3-inhibited autophagy in condylar cartilage degradation during the development of TMJOA.
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Autofagia , Cartílago Articular , Condrocitos , Ratones Noqueados , Osteoartritis , Factor de Transcripción CHOP , Animales , Factor de Transcripción CHOP/metabolismo , Factor de Transcripción CHOP/genética , Autofagia/fisiología , Cartílago Articular/metabolismo , Ratones , Osteoartritis/metabolismo , Osteoartritis/genética , Condrocitos/metabolismo , Trastornos de la Articulación Temporomandibular/metabolismo , Trastornos de la Articulación Temporomandibular/genética , Cóndilo Mandibular/metabolismo , Cóndilo Mandibular/patología , Proteínas de la Membrana , Factor 2 Relacionado con NF-E2 , Hemo-Oxigenasa 1RESUMEN
The cytotoxic mycotoxin deoxynivalenol (DON) reportedly has adverse effects on oocyte maturation and embryonic development in pigs. Recently, the interplay between cell apoptosis and endoplasmic reticulum (ER) stress has garnered increasing attention in embryogenesis. However, the involvement of the inositol-requiring enzyme 1 (IRE1)/c-jun N-terminal kinase (JNK)/C/EBP homologous protein (CHOP) pathways of unfolded protein response (UPR) signaling in DON-induced apoptosis in porcine embryos remains unknown. In this study, we revealed that exposure to DON (0.25 µM) substantially decreased cell viability until the blastocyst stage in porcine embryos, concomitant with initiation of cell apoptosis through the IRE1/JNK/CHOP pathways in response to ER stress. Quantitative PCR confirmed that UPR signaling-related transcription factors were upregulated in DON-treated porcine blastocysts. Western blot analysis showed that IRE1/JNK/CHOP signaling was activated in DON-exposed porcine embryos, indicating that ER stress-associated apoptosis was instigated. The ER stress inhibitor tauroursodeoxycholic acid protected against DON-induced ER stress in porcine embryos, indicating that the toxic effects of DON on early developmental competence of porcine embryos can be prevented. In conclusion, DON exposure impairs the developmental ability of porcine embryos by inducing ER stress-mediated apoptosis via IRE1/JNK/CHOP signaling.
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Apoptosis , Estrés del Retículo Endoplásmico , Factor de Transcripción CHOP , Tricotecenos , Animales , Estrés del Retículo Endoplásmico/efectos de los fármacos , Apoptosis/efectos de los fármacos , Factor de Transcripción CHOP/metabolismo , Factor de Transcripción CHOP/genética , Porcinos , Tricotecenos/toxicidad , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , Proteínas Quinasas JNK Activadas por Mitógenos/genética , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/genética , Transducción de Señal/efectos de los fármacos , Embrión de Mamíferos/efectos de los fármacos , Respuesta de Proteína Desplegada/efectos de los fármacos , Blastocisto/efectos de los fármacos , Blastocisto/metabolismo , FemeninoRESUMEN
OBJECTIVE: Osteoarthritis (OA) is a degenerative disease characterized by the gradual degeneration of chondrocytes, involving endoplasmic reticulum (ER) stress. Esculin is a natural compound with antioxidant, anti-inflammatory and anti-tumor properties. However, its impact on ER stress in OA therapy has not been thoroughly investigated. We aim to determine the efficiency of Esculin in OA treatment and its underlying mechanism. METHODS: We utilized the tert-butyl hydroperoxide (TBHP) to establish OA model in chondrocytes. The expression of SIRT1, PERK/eIF2α pathway-related proteins, apoptosis-associated proteins and ER stress-related proteins were detected by Western blot and Real-time PCR. The apoptosis was evaluated by flow cytometry and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) staining. X-ray imaging, Hematoxylin & Eosin staining, Safranin O staining and immunohistochemistry were used to assess the pharmacological effects of Esculin in the anterior cruciate ligament transection (ACLT) rat OA model. RESULTS: Esculin downregulated the expression of PERK/eIF2α pathway-related proteins, apoptosis-associated proteins and ER stress-related proteins, while upregulated the expression of SIRT1 and Bcl2 in the TBHP-induced OA model in vitro. It was coincident with the results of TUNEL staining and flow cytometry. We further confirmed the protective effect of Esculin in the rat ACLT-related model. CONCLUSION: Our results suggest the potential therapeutic value of Esculin on osteoarthritis. It probably inhibits the PERK-eIF2α-ATF4-CHOP pathway by upregulating SIRT1, thereby mitigating endoplasmic reticulum stress and protecting chondrocytes from apoptosis.
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Apoptosis , Condrocitos , Modelos Animales de Enfermedad , Factor 2 Eucariótico de Iniciación , Osteoartritis , Estrés Oxidativo , Ratas Sprague-Dawley , Transducción de Señal , Sirtuina 1 , Factor de Transcripción CHOP , eIF-2 Quinasa , Animales , Condrocitos/efectos de los fármacos , Condrocitos/metabolismo , Condrocitos/patología , Sirtuina 1/metabolismo , Sirtuina 1/genética , eIF-2 Quinasa/metabolismo , eIF-2 Quinasa/genética , Osteoartritis/tratamiento farmacológico , Osteoartritis/metabolismo , Osteoartritis/patología , Factor 2 Eucariótico de Iniciación/metabolismo , Factor de Transcripción CHOP/metabolismo , Factor de Transcripción CHOP/genética , Ratas , Estrés Oxidativo/efectos de los fármacos , Masculino , Transducción de Señal/efectos de los fármacos , Apoptosis/efectos de los fármacos , Estrés del Retículo Endoplásmico/efectos de los fármacos , Células CultivadasRESUMEN
BACKGROUND: Myxoid liposarcoma (MLS) displays a distinctive tumor microenvironment and is characterized by the FUS::DDIT3 fusion oncogene, however, the precise functional contributions of these two elements remain enigmatic in tumor development. METHODS: To study the cell-free microenvironment in MLS, we developed an experimental model system based on decellularized patient-derived xenograft tumors. We characterized the cell-free scaffold using mass spectrometry. Subsequently, scaffolds were repopulated using sarcoma cells with or without FUS::DDIT3 expression that were analyzed with histology and RNA sequencing. RESULTS: Characterization of cell-free MLS scaffolds revealed intact structure and a large variation of protein types remaining after decellularization. We demonstrated an optimal culture time of 3 weeks and showed that FUS::DDIT3 expression decreased cell proliferation and scaffold invasiveness. The cell-free MLS microenvironment and FUS::DDIT3 expression both induced biological processes related to cell-to-cell and cell-to-extracellular matrix interactions, as well as chromatin remodeling, immune response, and metabolism. Data indicated that FUS::DDIT3 expression more than the microenvironment determined the pre-adipocytic phenotype that is typical for MLS. CONCLUSIONS: Our experimental approach opens new means to study the tumor microenvironment in detail and our findings suggest that FUS::DDIT3-expressing tumor cells can create their own extracellular niche.