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BACKGROUND: Idiopathic pulmonary fibrosis (IPF) is an age-related disease severely affecting life quality with its prevalence rising as the population ages, yet there is still no effective treatment available. Cell therapy has emerged as a promising option for IPF, however, the absence of mature and stable animal models for IPF immunodeficiency hampers preclinical evaluations of human cell therapies, primarily due to rapid immune clearance of administered cells. This study aims to establish a reliable pulmonary fibrosis (PF) model in immunodeficient mice that supports autologous cell therapy and to investigate underlying mechanism. METHODS: We utilized thirty 5-week-old male NOD/SCID mice, categorizing them into three age groups: 12weeks, 32 weeks and 43 weeks, with 6 mice euthanized randomly from each cohort for lung tissue analysis. We assessed fibrosis using HE staining, Masson's trichrome staining, α-SMA immunohistochemistry and hydroxyproline content measurement. Further, ß-galactosidase staining and gene expression analysis of MMP9, TGF-ß1, TNF-α, IL-1ß, IL-6, IL-8, SOD1, SOD2, NRF2, SIRT1, and SIRT3 were performed. ELISA was employed to quantify protein levels of TNF-α, TGF-ß1, and IL-8. RESULTS: When comparing lung tissues from 32-week-old and 43-week-old mice to those from 12-week-old mice, we noted a marked increase in inflammatory infiltration, fibrosis severity, and hydroxyproline content, alongside elevated expression levels of α-SMA and MMP9. Notably, the degree of fibrosis intensified with age. Additionally, ß-galactosidase staining became more pronounced in older mice. Quantitative PCR analyses revealed age-related, increases in the expression of senescence markers (GLB1, P16, P21), and proinflammatory genes (TGF-ß1, TNF-α, IL-1ß, IL-6, and IL-8). Conversely, the expression of anti-oxidative stress-related genes (SOD1, SOD2, NRF2, SIRT1, and SIRT3) declined, showing statistically significant differences (*P < 0.05, **P < 0.01, ***P < 0.001). ELISA results corroborated these findings, indicating a progressive rise in the protein levels of TGF-ß1, TNF-α, and IL-8 as the mice aged. CONCLUSIONS: The findings suggest that NOD/SCID mice aged 32 weeks and 43 weeks effectively model pulmonary fibrosis in an elderly context, with the disease pathogenesis likely driven by age-associated inflammation and oxidative stress.
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Envejecimiento , Modelos Animales de Enfermedad , Ratones Endogámicos NOD , Ratones SCID , Sirtuina 1 , Animales , Ratones , Masculino , Sirtuina 1/metabolismo , Sirtuina 1/genética , Pulmón/patología , Pulmón/metabolismo , Factor de Crecimiento Transformador beta1/metabolismo , Factor de Crecimiento Transformador beta1/genética , Metaloproteinasa 9 de la Matriz/metabolismo , Metaloproteinasa 9 de la Matriz/genética , Factor de Necrosis Tumoral alfa/metabolismo , Factor de Necrosis Tumoral alfa/genética , Factor 2 Relacionado con NF-E2/metabolismo , Factor 2 Relacionado con NF-E2/genética , Fibrosis Pulmonar Idiopática/genética , Fibrosis Pulmonar Idiopática/patología , Fibrosis Pulmonar Idiopática/metabolismo , Interleucina-8/metabolismo , Interleucina-8/genética , Superóxido Dismutasa-1/genética , Superóxido Dismutasa-1/metabolismo , Superóxido Dismutasa/metabolismo , Superóxido Dismutasa/genética , Sirtuina 3/genética , Sirtuina 3/metabolismo , Hidroxiprolina/metabolismo , Interleucina-6/metabolismo , Interleucina-6/genética , Actinas/metabolismo , Actinas/genética , Interleucina-1beta/metabolismo , Interleucina-1beta/genética , Fibrosis Pulmonar/genética , Fibrosis Pulmonar/patología , Fibrosis Pulmonar/metabolismoRESUMEN
Hepatocellular carcinoma (HCC) remains a lethal malignancy with limited treatment options. Recent discoveries have highlighted the pivotal role of miRNAs in HCC progression. We previously reported that the expression of miR-200b-3p was decreased in HCC cells and exosomal miR-200b-3p from hepatocytes inhibited angiogenesis by suppressing the expression of the endothelial transcription factor ERG (erythroblast transformation-specific (ETS)-related gene), leading to the hypothesis that the delivery of this miRNA may inhibit angiogenesis and suppress HCC growth in vivo. Here, we tested this hypothesis by using human HCC inoculation models. First, we transfected the human HepG2 HCC cells and established a stable cell line that overexpressed a high level of miR-200b-3p. When miR-200b-3p-overexpressing cells were injected into severe combined immunedeficiency (SCID)-beige mice, tumor growth was significantly reduced compared to tumors of control cells, with a reduction in the expression of ERG and vascular endothelial growth factor (VEGF) and subsequent angiogenesis. Intra-tumoral injection of exosomes containing high levels of miR-200b-3p also reduced the growth of parental HepG2 tumors with reduced ERG and VEGF expression and angiogenesis. These results validate the inhibitory role of miR-200b-3p in tumor angiogenesis, thereby suppressing HCC tumor growth, and provide a novel insight into its potential therapeutic application.
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Carcinoma Hepatocelular , Exosomas , Neoplasias Hepáticas , MicroARNs , Neovascularización Patológica , Regulador Transcripcional ERG , Factor A de Crecimiento Endotelial Vascular , MicroARNs/genética , MicroARNs/metabolismo , Humanos , Animales , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patología , Carcinoma Hepatocelular/metabolismo , Exosomas/metabolismo , Exosomas/genética , Neovascularización Patológica/genética , Neovascularización Patológica/metabolismo , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patología , Neoplasias Hepáticas/metabolismo , Células Hep G2 , Factor A de Crecimiento Endotelial Vascular/genética , Factor A de Crecimiento Endotelial Vascular/metabolismo , Ratones , Regulador Transcripcional ERG/genética , Regulador Transcripcional ERG/metabolismo , Ratones SCID , Regulación Neoplásica de la Expresión Génica , Proliferación Celular , AngiogénesisRESUMEN
Pancreatic cancer is associated with a high mortality rate, and there are still very few effective treatment options. Patient-derived xenografts have proven to be invaluable preclinical disease models to study cancer biology and facilitate testing of novel therapeutics. However, the severely immune-deficient mice used to generate standard models lack any functional immune system, thereby limiting their utility as a tool to investigate the tumor-immune cell interface. This chapter will outline a method for establishment of "humanized" patient-derived xenografts, which are reconstituted with human immune cells to imitate the immune-rich microenvironment of pancreatic cancer.
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Modelos Animales de Enfermedad , Neoplasias Pancreáticas , Microambiente Tumoral , Neoplasias Pancreáticas/patología , Neoplasias Pancreáticas/inmunología , Neoplasias Pancreáticas/terapia , Animales , Humanos , Ratones , Microambiente Tumoral/inmunología , Ensayos Antitumor por Modelo de Xenoinjerto/métodos , Xenoinjertos , Ratones SCIDRESUMEN
PURPOSE: Test the feasibility of an image-based method to identify taxane resistance in mouse bearing triple-negative breast cancer (TNBC) tumor xenografts. METHODS: Xenograft tumor-bearing mice from paclitaxel-sensitive and paclitaxel-resistant TNBC cells (MDA-MD-346) were generated by orthotopic injection into female NOD-SCID mice. When tumors reached 100-150 mm3, mice were scanned using [18F]choline PET/CT. Tumors were collected and sliced for autoradiography and immunofluorescence analysis. Quantitative data was analyzed accordingly. RESULTS: From fifteen mice scanned, five had taxane-sensitive cell line tumors of which two underwent taxol-based treatment. From the remaining 10 mice with taxane-resistant cell line tumors, four underwent taxol-based treatment. Only 13 mice had the tumor sample analyzed histologically. When normalized to the blood pool, both cell lines showed differences in metabolic uptake before and after treatment. CONCLUSIONS: Treated and untreated taxane-sensitive and taxane-resistant cell lines have different metabolic properties that could be leveraged before the start of chemotherapy.
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Tomografía Computarizada por Tomografía de Emisión de Positrones , Neoplasias de la Mama Triple Negativas , Humanos , Femenino , Animales , Ratones , Tomografía Computarizada por Tomografía de Emisión de Positrones/métodos , Neoplasias de la Mama Triple Negativas/diagnóstico por imagen , Neoplasias de la Mama Triple Negativas/tratamiento farmacológico , Línea Celular Tumoral , Ratones SCID , Ratones Endogámicos NOD , Tomografía de Emisión de Positrones/métodos , Paclitaxel/farmacología , Paclitaxel/uso terapéutico , Modelos Animales , Resistencia a Medicamentos , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Raman spectroscopy has emerged as a powerful tool in medical, biochemical, and biological research with high specificity, sensitivity, and spatial and temporal resolution. Recent advanced Raman systems, such as portable Raman systems and fiber-optic probes, provide the potential for accurate in vivo discrimination between healthy and cancerous tissues. In our study, a portable Raman probe spectrometer was tested in immunosuppressed mice for the in vivo localization of colorectal cancer malignancies from normal tissue margins. The acquired Raman spectra were preprocessed, and principal component analysis (PCA) was performed to facilitate discrimination between malignant and normal tissues and to highlight their biochemical differences using loading plots. A transfer learning model based on a one-dimensional convolutional neural network (1D-CNN) was employed for the Raman spectra data to assess the classification accuracy of Raman spectra in live animals. The 1D-CNN model yielded an 89.9% accuracy and 91.4% precision in tissue classification. Our results contribute to the field of Raman spectroscopy in cancer diagnosis, highlighting its promising role within clinical applications.
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Neoplasias Colorrectales , Aprendizaje Profundo , Animales , Ratones , Espectrometría Raman/métodos , Redes Neurales de la Computación , Neoplasias Colorrectales/diagnósticoRESUMEN
Liver fibrosis, a critical pathological feature of chronic liver diseases, arises from a multitude of pathogenic factors. Consequently, establishing an appropriate animal model to simulate liver fibrosis holds immense significance for comprehending its underlying pathogenesis. Despite the numerous methodologies available for generating liver fibrosis models, they often deviate substantially from the spontaneous age-related liver fibrosis process. In this study, compared with young (12 weeks) and middle-aged NOD/SCID mice (32 weeks), there were a large number of fibrous septum and collagen in the liver tissue of old NOD/SCID mice (43 weeks, 43 W). Immunohistochemical analysis unequivocally indicated heightened α-SMA content within the liver tissue of the 43 W mice, thereby underscoring aging's role in triggering the epithelial-to-mesenchymal transition. In addition, SA-ß-gal staining as well as P21 expression were increased, and SIRT1 and SIRT3 expression were decreased in 43 W mice. A comprehensive evaluation encompassing transmission electron microscopy and fluorescence quantitative analysis elucidated compromised mitochondrial function and reduced antioxidant capacity in hepatocytes of the 43 W mice. Furthermore, the aging process activated the pro-fibrotic TGF-ß-SMAD pathway, concurrently inducing hepatocellular inflammation. The results of the present study not only validate the successful construction of a spontaneous liver fibrosis mouse model through natural aging induction but also provide initial insights into the mechanisms underpinning age-induced liver fibrosis.
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Patient-derived xenografts (PDXs) are valuable models to study cancer biology, behavior, and response to therapies in vivo. Pancreatic cancer is an aggressive and treatment-resistant disease, and typical biopsies are often of low cellular yield and therefore present challenges for the creation of PDXs. This chapter will describe a method to establish PDX models from tissue biopsies obtained via endoscopic ultrasound-guided fine-needle aspiration, a relatively noninvasive technique which compared to surgery is available to pancreatic cancer patients at all stages of disease. Furthermore, we also describe methods to incorporate "humanization" of PDXs via reconstitution with human immune cells, thus mimicking the immune cell-rich microenvironment of pancreatic tumors.
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Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Animales , Humanos , Neoplasias Pancreáticas/diagnóstico por imagen , Neoplasias Pancreáticas/patología , Carcinoma Ductal Pancreático/diagnóstico por imagen , Carcinoma Ductal Pancreático/patología , Biopsia por Aspiración con Aguja Fina Guiada por Ultrasonido Endoscópico/métodos , Modelos Animales de Enfermedad , Inflamación , Microambiente Tumoral , Neoplasias PancreáticasRESUMEN
The betanodavirus B2 protein targets mitochondria and triggers mitochondrion-mediated cell death signaling in lung cancer cells; however, its molecular mechanism remains unknown. In this study, we observed that B2 triggers hydrogen peroxide/Nrf2-involved stress signals in the dynamic regulation of non-small lung cancer cell (NSCLC)-programmed cell death. Here, the B2 protein works as a necrotic inducer that triggers lung cancer death via p53 upregulation and RIP3 expression, suggesting a new perspective on lung cancer therapy. We employed the B2 protein to target A549 lung cancer cells and solid tumors in NOD/SCID mice. Tumors were collected and processed for the hematoxylin and eosin staining of tissue and cell sections, and their sera were used for blood biochemistry analysis. We observed that B2 killed an A549 cell-induced solid tumor in NOD/SCID mice; however, the mutant ΔB2 did not. In NOD/SCID mice, B2 (but not ΔB2) induced both p53/Bax-mediated apoptosis and RIPK3-mediated necroptosis. Finally, immunochemistry analysis showed hydrogen peroxide /p38/Nrf2 stress strongly inhibited the production of tumor markers CD133, Thy1, and napsin, which correlate with migration and invasion in cancer cells. This B2-triggered, ROS/Nrf2-mediated stress signal triggered multiple signals via pathways that killed A549 lung cancer tumor cells in vivo. Our results provide novel insight into lung cancer management and drug therapy.
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BACKGROUND: The key signals that suffice to induce atopic dermatitis (AD) in human skin remain incompletely understood. Also, current mouse models reflect human AD only unsatisfactorily. Therefore, we have asked whether a humanized AD mouse model can be developed that reflects human AD more faithfully and permit to identify key signals that suffice to induce AD lesions in previously healthy human skin in vivo. METHODS: Healthy human skin from non-atopic donors was transplanted onto SCID/beige mice. After xenotransplant reinnervation by mouse sensory nerve fibers had occurred, mixed autologous human Th2 CD4+ and Tc2 CD8+ T cells that had been pretreated in vitro with IL-2, IL-4, and LPS were injected intradermally into the xenotransplants without skin barrier disruption. RESULTS: Injected non-atopic xenotransplants rapidly developed a morphological, functional, and immunological phenocopy of human AD lesions regarding skin barrier defects, immunopathology including intraepidermal eosinophils, mast cell activation, increase of thymic stromal lymphopoietin, eotaxin-1 and type 2 cytokine circuits, and even showed characteristic neuroimmunological abnormalities such as ß2-adrenergic receptor downregulation. The experimentally induced AD lesions in human skin responded to standard AD therapy (topical dexamethasone or tacrolimus; systemic anti-IL-4Rα antibody [dupilumab]), and relapsed when neurogenic skin inflammation was induced by exposing mice to perceived stress. CONCLUSIONS: This new animal model uniquely mimics the complexity of human AD and its clinical response to standard therapy and psychoemotional stressors in vivo, and shows that Th2-polarized lymphocytes associated with excessive IL-4Rα-mediated signaling suffice to induce human AD skin lesions, while atopy and epidermal barrier disruption are dispensable.
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Dermatitis Atópica , Humanos , Ratones , Animales , Ratones SCID , Piel/patología , Citocinas/metabolismo , Linfocitos T CD8-positivosRESUMEN
Nanodiamonds (NDs) have high potential as a drug carrier and in combination with nitrogen vacancies (NV centers) for highly sensitive MR-imaging after hyperpolarization. However, little remains known about their physiological properties in vivo. PET imaging allows further evaluation due to its quantitative properties and high sensitivity. Thus, we aimed to create a preclinical platform for PET and MR evaluation of surface-modified NDs by radiolabeling with both short- and long-lived radiotracers. Serum albumin coated NDs, functionalized with PEG groups and the chelator deferoxamine, were labeled either with zirconium-89 or gallium-68. Their biodistribution was assessed in two different mouse strains. PET scans were performed at various time points up to 7 d after i.v. injection. Anatomical correlation was provided by additional MRI in a subset of animals. PET results were validated by ex vivo quantification of the excised organs using a gamma counter. Radiolabeled NDs accumulated rapidly in the liver and spleen with a slight increase over time, while rapid washout from the blood pool was observed. Significant differences between the investigated radionuclides were only observed for the spleen (1 h). In summary, we successfully created a preclinical PET and MR imaging platform for the evaluation of the biodistribution of NDs over different time scales.
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Duchenne Muscular Dystrophy (DMD) is a lethal disease caused by mutations in dystrophin encoding gene, causing progressive degeneration of cardiac, respiratory, and skeletal muscles leading to premature death due to cardiac and respiratory failure. Currently, there is no cure for DMD. Therefore, novel therapeutic approaches are needed for DMD patients.We have previously reported functional improvements which correlated with increased dystrophin expression following administration of dystrophin expressing chimeric (DEC) cells of myoblast origin to the mdx mouse models of DMD.In the current study, we confirmed dose-dependent protective effect of human DEC therapy created from myoblasts of normal and DMD-affected donors, on restoration of dystrophin expression and amelioration of cardiac, respiratory, and skeletal muscle function at 180 days after systemic-intraosseous DEC administration to mdx/scid mouse model of DMD. Functional improvements included maintenance of ejection fraction and fractional shortening levels on echocardiography, reduced enhanced pause and expiration time on plethysmography and improved grip strength and maximum stretch induced contraction of skeletal muscles. Improved function was associated with amelioration of mdx muscle pathology revealed by reduced muscle fibrosis, reduced inflammation and improved muscle morphology confirmed by reduced number of centrally nucleated fibers and normalization of muscle fiber diameters. Our findings confirm the long-term systemic effect of DEC therapy in the most severely affected by DMD organs including heart, diaphragm, and long skeletal muscles.These encouraging preclinical data introduces human DEC as a novel therapeutic modality of Advanced Therapy Medicinal Product (ATMP) with the potential to improve or halt the progression of DMD and enhance quality of life of DMD patients. Human DEC as a novel therapeutic modality with the potential to improve or halt progression of the DMD disease and enhance quality of life of DMD patients. Graphical abstract represents manufacturing process of the human DEC therapy for the future clinical applications. 1. We report the long-term efficacy of human DEC therapy resulting in increased dystrophin expression and reduced mdx muscle pathology after systemic-intraosseous administration of human Dystrophin Expressing Chimeric (DEC) Cells to the mdx/scid mouse model of DMD. 2. Systemic administration of human DEC therapy resulted in amelioration of cardiac, respiratory and skeletal muscle function as confirmed by echocardiography, plethysmography and standard muscle strength tests respectively. 3. We introduce human DEC as a novel Advanced Therapy Medicinal Product (ATMP) for future clinical application in DMD patients.
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Distrofia Muscular de Duchenne , Humanos , Ratones , Animales , Distrofia Muscular de Duchenne/genética , Distrofia Muscular de Duchenne/terapia , Distrofia Muscular de Duchenne/metabolismo , Ratones Endogámicos mdx , Distrofina/genética , Distrofina/metabolismo , Ratones SCID , Calidad de Vida , Músculo Esquelético/metabolismo , Modelos Animales de Enfermedad , Tratamiento Basado en Trasplante de Células y TejidosRESUMEN
Combining NSAIDs with conventional therapeutics was recently explored as a new strategy in cancer therapy. Our earlier studies showed that novel oleanolic acid oximes (OAO) conjugated with aspirin or indomethacin may enhance their anti-cancer potential through modulation of the Nrf2 and NF-κB signaling pathways. This study focused on the synthesis and biological evaluation of four diclofenac (DCL)-OAO derivative conjugates in the context of these pathways' modification and hepatic cells survival. Treatment with the conjugates 4d, 3-diclofenacoxyiminoolean-12-en-28-oic acid morpholide, and 4c, 3-diclofenacoxyiminoolean-12-en-28-oic acid benzyl ester significantly reduced cell viability in comparison to the DCL alone. In THLE-2, immortalized normal hepatocytes treated with these conjugates resulted in the activation of Nrf2 and increased expression in SOD-1 and NQO1, while the opposite effect was observed in the HepG2 hepatoma cells. In both cell lines, reduced activation of the NF-κB and COX-2 expression was observed. In HepG2 cells, conjugates increased ROS production resulting from a reduced antioxidant defense, induced apoptosis, and inhibited cell proliferation. In addition, the OAO morpholide derivative and its DCL hybrid reduced the tumor volume in mice bearing xenografts. In conclusion, our study demonstrated that conjugating diclofenac with the OAO morpholide and a benzyl ester might enhance its anti-cancer activity in HCC.
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Lentinan (SLNT) has been shown to be directly cytotoxic to cancer cells. However, this direct antitumour effect has not been thoroughly investigated in vivo, and the mechanism remains unclear. We aimed to examine the direct antitumour effect of SLNT on human colon cancer and the mechanism in vivo and in vitro. SLNT significantly inhibited tumour growth and induced autophagy and endoplasmic reticulum stress (ERS) in HT-29 cells and tumour-bearing nonobese diabetic (NOD)/severe combined immunodeficiency (SCID) mice. Experiments with the autophagy inhibitors chloroquine (CQ) and 3-methyladenine (3-MA) showed that autophagy facilitated the antitumour effect of SLNT. Moreover, ERS was identified as the common upstream regulator of SLNT-induced increases in Ca2+concentrations, autophagy and apoptosis by using ERS inhibitors. In summary, our study demonstrated that SLNT exerted direct antitumour effects on human colon cancer via ERS-mediated autophagy and apoptosis, providing a novel understanding of SLNT as an anti-colon cancer therapy.
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Antineoplásicos/uso terapéutico , Apoptosis/efectos de los fármacos , Autofagia/efectos de los fármacos , Estrés del Retículo Endoplásmico/efectos de los fármacos , Lentinano/uso terapéutico , Neoplasias/tratamiento farmacológico , Animales , Línea Celular Tumoral , Humanos , Masculino , Ratones Endogámicos NOD , Ratones SCID , Transducción de Señal/efectos de los fármacos , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
The sesquiterpene lactone parthenolide is a major component of the feverfew medicinal plant, Tanacetum parthenium. Parthenolide has been extensively studied for its anti-inflammatory and anticancer properties in several tumor models. Parthenolide's antitumor activities depend on several mechanisms but it is mainly known as an inhibitor of the nuclear factor-κB (NF-κB) pathway. This pathway is constitutively activated and induces cell survival in primary effusion lymphoma (PEL), a rare aggressive AIDS-related lymphoproliferative disorder that is commonly caused by the human herpesvirus 8 (HHV-8) infection. The aim of this study is to evaluate the targeted effect of Parthenolide both in vitro and in vivo. Herein, parthenolide significantly inhibited cell growth, induced G0 /G1 cell cycle arrest, and induced massive apoptosis in PEL cells and ascites. In addition, parthenolide inhibited the NF-ĸB pathway suppressing IĸB phosphorylation and p65 nuclear translocation. It also reduced the expression of the DNA methylase inhibitor (DNMT1). Parthenolide induced HHV-8 lytic gene expression without inhibiting latent viral gene expression. Importantly, DMAPT, the more soluble parthenolide prodrug, promoted delay in ascites development and prolonged the survival of PEL xenograft mice. This study supports the therapeutic use of parthenolide in PEL and encourages its further clinical development.
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Antineoplásicos Fitogénicos/farmacología , Linfoma de Efusión Primaria/tratamiento farmacológico , Sesquiterpenos/farmacología , Animales , Apoptosis/efectos de los fármacos , Biomarcadores , Puntos de Control del Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Modelos Animales de Enfermedad , Susceptibilidad a Enfermedades , Evaluación Preclínica de Medicamentos , Humanos , Linfoma de Efusión Primaria/etiología , Linfoma de Efusión Primaria/metabolismo , Ratones , Ratones Endogámicos NOD , Ratones SCID , Transducción de Señal/efectos de los fármacos , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
OBJECTIVE: To establish animal models epidermal growth factor receptor inhibitor-related skin rashes using cetuximab, gefitinib or erlotinib. OBJECTIVE: Female SCID mice were randomly divided into blank control group and high-, moderate-, and low-dose cetuximab groups. The mice in control group received intraperitoneal injection of saline, and those in the 3 cetuximab groups were injected with 80, 40, and 20 mg/kg cetuximab (3 times a week for 4 weeks), respectively. The general skin appearance and skin pathologies of the mice were observed. Female BN rats were randomly divided into blank group, ovalbumin group, gefitinib group and erlotinib group, and in the latter 3 groups, the rats were given ovalbumin (1 mg), gefitinib (37.5 mg/kg), and erlotinib (23.5 mg/kg) by lavage once daily for 45 days, respectively. Skin pathologies of the rats were observed, and serum levels of TNF-α, IL-6 and other inflammatory factors were detected using ELISA. OBJECTIVE: Intraperitoneal injection of cetuximab did not induce typical skin rashes, scabs or obvious skin inflammation in the mice. In female BN rats, lavage of gefitinib caused obvious skin rashes, scabs and exudation, and obvious inflammatory cell infiltration, keratinosis, spinous layer release and epidermal thickening were observed in the skin. No obvious skin inflammation were observed in the rats in the control, ovalbumin or erlotinib groups. While IgE (P=0.061) and TNF-α concentrations (P=0.057) did not differ significantly among the groups, serum levels of IL-6 was significantly higher in gefitinib group than in the blank control group (P=0.016) but similar between erlotinib group and the blank group (P=0.910). OBJECTIVE: Intraperitoneal injection of cetuximab can not induce epidermal growth factor receptor inhibitor-related skin rashes in SCID mice. Lavage of gefitinib, but not erlotinib, can be used to establish models of epidermal growth factor receptor inhibitor-related rashes in BN rats.
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Receptores ErbB , Neoplasias Pulmonares , Animales , Clorhidrato de Erlotinib/toxicidad , Femenino , Ratones , Ratones SCID , Modelos Animales , Quinazolinas/toxicidad , RatasRESUMEN
Ecdysteroids are of interest as potential sport performance enhancers, due to their anabolic effects. The current study aimed to analyze levels of the most abundant ecdysteroid, ecdysterone (20-hydroxyecdysone, 20-OHE) in easily available dietary supplements, and, outline an analytical strategy for its detection, and that, of its metabolites, (1) following administration of pure 20-OHE to uPA(+/+)-SCID mice with humanized liver, (2) in a human volunteer after ingestion of two supplements, one with a relatively low, and the other a high, concentration of 20-OHE, and, (3) to estimate the prevalence of use of 20-OHE in elite athletes (n = 1000). Of the 16 supplements tested, only five showed detectable levels of 20-OHE, with concentrations ranging from undetectable up to 2.3 mg per capsule. Urine of uPA(+/+)-SCID urine showed the presence of 20-OHE and its metabolite, 14 deoxy ecdysterone, within 24 hours (hr) of ingestion. In humans, both the parent and the metabolite were detectable within 2 to 5 hr of ingestion, with the metabolite being detectable for longer than the parent. After ingestion of a low dose supplement, the parent and metabolite were detectable for 70 and 48 hr, while following the higher dose it was 96 and 48 hr, respectively. Analysis of urines from athletes (n = 1000) confirmed four positives for 20-OHE, suggesting a prevalence of use of 0.4%. Prevalence of its use by elite athletes was relatively low, however, this needs to be confirmed in other populations, and with other related ecdysteroids.
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Suplementos Dietéticos/análisis , Doping en los Deportes/prevención & control , Ecdisterona/orina , Detección de Abuso de Sustancias/métodos , Adulto , Animales , Atletas , Ecdisterona/análisis , Ecdisterona/metabolismo , Femenino , Humanos , Hígado/metabolismo , Masculino , Ratones , Ratones SCID , Prevalencia , Factores de TiempoRESUMEN
Chronic nonhealing wounds impact nearly 15% of Medicare beneficiaries (8.2 million) in the United States costing $28-$32 billion annually. Despite advancement in wound management, approximately 8% of diabetic Medicare beneficiaries have a foot ulcer and 1.8% will have an amputation. The development of a regenerative approach is warranted to save these before-mentioned amputations. To this extent, herein, we describe the detailed methods in generating a type 1 diabetes mellitus (T1DM) condition in immunocompromised mice, inducing cutaneous wound, and application of dental pulp stem cell-derived secretory products for therapeutic assessment. This model helps in evaluating the efficacy of stem cell-based therapy and helps with the investigation of involved mechanisms in impaired cutaneous wound healing caused by hyperglycemic stress due to type 1 diabetes.
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Pulpa Dental/trasplante , Pie Diabético/terapia , Trasplante de Células Madre/métodos , Cicatrización de Heridas/genética , Animales , Pulpa Dental/citología , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Experimental/patología , Diabetes Mellitus Tipo 1/complicaciones , Diabetes Mellitus Tipo 1/terapia , Diabetes Mellitus Tipo 2/complicaciones , Diabetes Mellitus Tipo 2/terapia , Pie Diabético/complicaciones , Pie Diabético/patología , Humanos , Ratones , Piel/lesiones , Piel/patología , Células Madre/citologíaRESUMEN
Despite significant advances in diabetic wound management, diabetic wounds remain a significant global problem that decreases patient's quality of life, and chronic wounds may lead to amputation and death to the patients. To develop a potential regenerative therapy, a xenogeneic transplantation compatible laboratory model needs to be developed. This procedure demonstrates how to isolate hematopoietic stem cells (CD133+) from human umbilical cord blood, expand CD34+ stem cells using a nanofiber scaffold (polyether sulfone-coated and amino group-treated), induce diabetes in immunocompromised (NOD/SCID) mice, induce a cutaneous wound in mice, and how to treat the wound with the nanofiber-expanded CD34+ stem cells. This protocol also shows how to measure wound healing.
Asunto(s)
Complicaciones de la Diabetes/terapia , Trasplante de Células Madre Hematopoyéticas/métodos , Células Madre Hematopoyéticas/citología , Nanofibras/química , Enfermedades de la Piel/terapia , Animales , Complicaciones de la Diabetes/patología , Modelos Animales de Enfermedad , Sangre Fetal/trasplante , Supervivencia de Injerto/genética , Humanos , Ratones , Calidad de Vida , Enfermedades de la Piel/patología , Cicatrización de Heridas/genética , Cicatrización de Heridas/fisiologíaRESUMEN
PURPOSE: To evaluate the antitumor efficacy of Ag3Au1Trp1:2NPs in a SCID mouse cancer model, with respect to their effect on tumor growth, on tumor's metastatic potential and the underlying molecular mechanism. SUBJECTS AND METHODS: Ag3Au1Trp1:2NPs were radiolabeled with Gallium-68 and the biodistribution was studied in Swiss mice without tumors and in SCID mice bearing tumors. SCID mice received intratumoral Ag3Au1Trp1:2NPs and tumor size was measured using calipers. Lung and liver tissues were extracted and studied microscopically for the detection of any metastatic sites. Changes in the Caspase-3 and TNF-related apoptosis-inducing ligand (TRAIL) were also investigated using real-time PCR and Western blot techniques, respectively. RESULTS: In the 4T1 tumor-bearing SCID mice, Ag3Au1Trp1:2NPs showed quick passive accumulation at tumor sites at 30 mins post-injection. Mice that received the highest dose of NPs (5.6mg/mL) demonstrated a 1.9-fold lower tumor volume compared to that of the control group at 11 days post-injection, while mice that did not receive NPs showed metastatic sites in liver and lung. Extracted tumor tissue of treated mice revealed increased Casp-3 mRNA levels as well as elevated TRAIL protein levels. CONCLUSION: Based on our results, Ag3Au1Trp1:2NPs express anti-tumor and anti-metastatic effects in vivo. Ag3Au1Trp1:2NPs also reach tumor site via the enhancement and retention effect which results in the apoptotic death of cancerous cells selectively via the extrinsic TRAIL-dependent pathway.
Asunto(s)
Antineoplásicos/química , Antineoplásicos/farmacología , Nanopartículas del Metal/uso terapéutico , Neoplasias Experimentales/patología , Animales , Antineoplásicos/farmacocinética , Apoptosis/efectos de los fármacos , Caspasa 3/metabolismo , Línea Celular Tumoral , Movimiento Celular , Femenino , Radioisótopos de Galio/química , Oro/química , Oro/farmacología , Neoplasias Hepáticas/prevención & control , Neoplasias Hepáticas/secundario , Neoplasias Pulmonares/prevención & control , Neoplasias Pulmonares/secundario , Nanopartículas del Metal/química , Ratones SCID , Neoplasias Experimentales/tratamiento farmacológico , Plata/química , Plata/farmacología , Ligando Inductor de Apoptosis Relacionado con TNF/metabolismo , Distribución Tisular , Carga TumoralRESUMEN
BACKGROUND: DNA-dependent protein kinase (DNA-PK), consisting of a Ku heterodimer (Ku70/80) and a large catalytic subunit (DNA-PKcs), plays an important role in the repair of DNA double-strand breaks via non-homologous end-joining (NHEJ) in mammalian cells. Severe combined immunodeficient (scid) mice carry a mutation in the gene encoding DNA-PKcs and are sensitive to ionizing radiation. To examine the roles of DNA-PKcs in the generation of deletion mutations in vivo, we crossed scid mice with gpt delta transgenic mice for detecting mutations. RESULTS: The scid and wild-type (WT) gpt delta transgenic mice were irradiated with a single X-ray dose of 10 Gy, and Spi- mutant frequencies (MFs) were determined in the brain and spleen 2 days after irradiation. Irradiation with X-rays significantly enhanced Spi- MF in both organs in the scid and WT mice. The MFs in the brain of irradiated scid mice were significantly lower than those in WT mice, i.e., 2.9 ± 1.0 × 10- 6 versus 5.0 ± 1.1 × 10- 6 (P < 0.001), respectively. In the spleen, however, both mouse strains exhibited similar MFs, i.e., 4.1 ± 1.8 × 10- 6 versus 4.8 ± 1.4 × 10- 6. Unirradiated scid and WT mice did not exhibit significant differences in MFs in either organ. CONCLUSIONS: DNA-PKcs is unessential for the induction of deletion mutations in the spleen, while it plays a role in this in the brain. Therefore, the contribution of DNA-PKcs to NHEJ may be organ-specific.