RESUMEN
BACKGROUND: Community-acquired pneumonia (CAP) is a significant cause of morbidity and mortality worldwide. Despite effective antimicrobial therapy, CAP can induce pulmonary endothelial hyperpermeability resulting in life-threatening lung failure due to an exaggerated host-pathogen interaction. Treatment of acute lung injury is mainly supportive because key elements of inflammation-induced barrier disruption remain undetermined. Angiopoietin-1 (Ang-1)-mediated Tie2 activation reduces, and the Ang-1 antagonist Ang-2 increases, inflammation and endothelial permeability in sepsis. Vasculotide (VT) is a polyethylene glycol-clustered Tie2-binding peptide that mimics the actions of Ang-1. The aim of our study was to experimentally test whether VT is capable of diminishing pneumonia-induced lung injury. METHODS: VT binding and phosphorylation of Tie2 were analyzed using tryptophan fluorescence spectroscopy and phospho-Tie-2 enzyme-linked immunosorbent assay. Human and murine lung endothelial cells were investigated by immunofluorescence staining and electric cell-substrate impedance sensing. Pulmonary hyperpermeability was quantified in VT-pretreated, isolated, perfused, and ventilated mouse lungs stimulated with the pneumococcal exotoxin pneumolysin (PLY). Furthermore, Streptococcus pneumoniae-infected mice were therapeutically treated with VT. RESULTS: VT showed dose-dependent binding and phosphorylation of Tie2. Pretreatment with VT protected lung endothelial cell monolayers from PLY-induced disruption. In isolated mouse lungs, VT decreased PLY-induced pulmonary permeability. Likewise, therapeutic treatment with VT of S. pneumoniae-infected mice significantly reduced pneumonia-induced hyperpermeability. However, effects by VT on the pulmonary or systemic inflammatory response were not observed. CONCLUSIONS: VT promoted pulmonary endothelial stability and reduced lung permeability in different models of pneumococcal pneumonia. Thus, VT may provide a novel therapeutic perspective for reduction of permeability in pneumococcal pneumonia-induced lung injury.
Asunto(s)
Permeabilidad Capilar/efectos de los fármacos , Pulmón/efectos de los fármacos , Fragmentos de Péptidos/farmacocinética , Animales , Infecciones Comunitarias Adquiridas/tratamiento farmacológico , Modelos Animales de Enfermedad , Células Endoteliales/metabolismo , Femenino , Ratones , Ratones Endogámicos C57BL , Fragmentos de Péptidos/uso terapéutico , Neumonía Neumocócica/tratamiento farmacológico , Espectrometría de Fluorescencia/métodos , Estadísticas no Paramétricas , Streptococcus pneumoniae/metabolismo , Streptococcus pneumoniae/patogenicidadRESUMEN
BACKGROUND: While therapeutic resistance is difficult to model in vitro in its entirety, in vivo passage and re-derivation of treatment resistant prostate cancer cell variants is a strategy to study therapeutic resistance more comprehensively. However, the process of in vivo passage itself may result in gene expression changes that could confound the analysis of such resistant cell variants compared to their parental cell lines. METHODS: We compared the expression profiles of parental PC-3 human prostate cancer cells and PC-3 cells re-derived after in vivo passage in athymic nude mice. Whole transcriptome information was obtained using the SOLiD 4 system (Applied Biosystems). Differentially expressed genes were mapped to genes in the Database for Annotation, Visualization and Integrated Discovery for gene enrichment and functional annotation analysis. The expression of a panel of these genes was validated using quantitative RT-PCR. RESULTS: Altogether, 21,032 distinct transcripts were found in PC-3 and/or NS1.1. Of these, 906 were differentially regulated (≥2-fold) in NS1.1 versus PC-3. 337 transcripts were upregulated, and 569 were downregulated, including genes previously associated with various aspects of prostate carcinogenesis such as TLR4 and IGFBP5, respectively. Gene ontology analysis of the differentially expressed transcripts revealed enrichment for biological processes such as cell adhesion, migration, and angiogenesis. CONCLUSIONS: When using in vivo as opposed to in vitro derived prostate cancer cell variants for comparative genetic studies of complex traits such as therapeutic resistance, one may be better served to use similarly in vivo passaged control cell variants instead of parental cell lines.
Asunto(s)
Línea Celular Tumoral/trasplante , Regulación Neoplásica de la Expresión Génica , Trasplante de Neoplasias/métodos , Neoplasias de la Próstata/genética , Animales , Perfilación de la Expresión Génica , Humanos , Masculino , Ratones , Ratones Desnudos , Neoplasias de la Próstata/patologíaRESUMEN
ATG4B belongs to the autophagin family of cysteine proteases required for autophagy, an emerging target of cancer therapy. Developing pharmacological ATG4B inhibitors is a very active area of research. However, detailed studies on the role of ATG4B during anticancer therapy are lacking. By analyzing PC-3 and C4-2 prostate cancer cells overexpressing dominant negative ATG4B(C74A)in vitro and in vivo, we show that the effects of ATG4B(C74A) are cell type, treatment, and context-dependent. ATG4B(C74A) expression can either amplify the effects of cytotoxic therapies or contribute to treatment resistance. Thus, the successful clinical application of ATG4B inhibitors will depend on finding predictive markers of response.
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Autofagia/efectos de los fármacos , Cisteína Endopeptidasas/metabolismo , Inhibidores de Cisteína Proteinasa/uso terapéutico , Terapia Molecular Dirigida , Neoplasias de la Próstata/tratamiento farmacológico , Animales , Proteínas Relacionadas con la Autofagia , Línea Celular Tumoral , Cisteína Endopeptidasas/genética , Inhibidores de Cisteína Proteinasa/farmacología , Docetaxel , Doxorrubicina/farmacología , Doxorrubicina/uso terapéutico , Resistencia a Antineoplásicos , Humanos , Masculino , Ratones , Ratones Desnudos , Trasplante de Neoplasias , Tolerancia a Radiación , Taxoides/farmacología , Taxoides/uso terapéutico , Topotecan/farmacología , Topotecan/uso terapéuticoRESUMEN
The endothelial glycocalyx (eGC), a carbohydrate-rich layer lining the luminal surface of the endothelium, provides a first vasoprotective barrier against vascular leakage in sepsis. We hypothesized that angiopoietin-2 (Angpt-2), antagonist of the endothelium-stabilizing receptor Tie2, induces a rapid loss of the eGC in human sepsis. Using intravital microscopy, we measured the perfused boundary region (PBR), an inverse parameter of eGC dimensions in sublingual microvessels, in patients with sepsis and age-matched nonseptic subjects. Median PBR values were significantly higher in patients compared with controls and correlated with serum Angpt-2 levels. To transfer and further explore these findings in a cell culture system, we exposed endothelial cells (ECs) to serum (5%) from a subgroup of septic patients and nonseptic controls. Confocal and atomic force microscopy revealed that sepsis serum, but not control serum, induced thinning of the eGC on human ECs in vitro, which correlated with paired PBR values obtained in vivo (r = 0.96, p < 0.01). Inhibition of Angpt-2 or Tie2 activation completely abolished eGC damage. Mechanistically, sepsis-induced eGC breakdown required the loss of its main constituent heparan sulfate; a result of heparan sulfate-specific enzyme heparanase, which was suppressed by Tie2 activation. Finally, Tie2 activation, but not Angpt-2 inhibition, initiated after septic or enzymatic damage provoked rapid refurbishment of the eGC. Our data indicate that eGC breakdown in human sepsis is mediated via Tie2 deactivation by Angpt-2. Activation of Tie2 seems to accelerate recovery of the eGC and might hold promise as a therapeutic target in human sepsis.
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Células Endoteliales/metabolismo , Glicocálix/metabolismo , Receptor TIE-2/metabolismo , Sepsis/metabolismo , Adulto , Anciano , Angiopoyetina 2/sangre , Estudios de Casos y Controles , Línea Celular , Células Endoteliales/patología , Femenino , Glucuronidasa/metabolismo , Glicocálix/patología , Heparitina Sulfato/metabolismo , Humanos , Microscopía Intravital , Masculino , Microscopía de Fuerza Atómica , Persona de Mediana Edad , Fosforilación , Estudios Prospectivos , Sepsis/sangre , Sepsis/patología , Transducción de SeñalRESUMEN
Castration-resistant prostate cancer (CRPC) is an advanced and incurable stage of the second most frequently diagnosed malignancy in men globally. Current treatment options improve survival modestly but eventually fail due to intrinsic or acquired therapeutic resistance. A hypothesis is presented wherein circulating levels of fibroblast growth factor 23 (FGF23), an endocrine member of the fibroblast growth factor family with phosphaturic properties, are proposed as a prognostic and predictive marker to identify CRPC patients with poor prognosis that are amenable to FGF23 antibody therapy (FGF23i) or treatment with fibroblast growth factor receptor inhibitors (FGFRi). With respect to the latter, FGF23 may also serve as a pharmacodynamic marker enabling individualized FGFRi dosing. We recently discovered that the development of severe and sustained hypophosphatemia in CRPC patients undergoing zoledronic acid therapy for bone metastases was associated with markedly worse prognosis compared to patients without or with only mild and transient hypophosphatemia. Severe hypophosphatemia is a typical manifestation of tumor-induced hypophosphatemic osteomalacia (TIO), a paraneoplastic condition mediated by FGF23 overexpression in most instances. While the postulated tumor-promoting role of FGF23 in CRPC or other malignancies has not yet been studied, several lines of evidence suggest that FGF23 may mediate both severe hypophosphatemia (via its endocrine properties) and aggressive CRPC behavior (via autocrine and paracrine activities): (i) FGF23 and the necessary signalling machinery (i.e. members of the fibroblast growth factor receptor [FGFR] family and the essential co-receptor α-KLOTHO [KL]) are highly expressed in a sizeable subgroup of CRPC patients; (ii) FGF/FGFR signalling plays important roles in prostate cancer; (iii) FGF23 can induce its own expression via a positive autocrine feedback loop involving FGFR1; and (iv) this positive feedback loop may be triggered by bone-targeted therapies frequently used for the treatment of CRPC-associated bone metastases. While there is a lack of personalized treatment strategies in the management of CRPC to date, FGF23 targeted therapy has the potential to fill this unmet clinical need in the not-so-distant future. In fact, FGFRi are currently in advanced clinical testing for a number of malignancies such as kidney and lung cancer, but there is a lack of conclusive data on FGFRi therapy in patients selected for FGF/FGFR pathway activation.
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Factores de Crecimiento de Fibroblastos/metabolismo , Hipofosfatemia/complicaciones , Orquiectomía , Neoplasias de la Próstata/metabolismo , Factor-23 de Crecimiento de Fibroblastos , Humanos , Masculino , Modelos Teóricos , Pronóstico , Neoplasias de la Próstata/complicaciones , Neoplasias de la Próstata/fisiopatología , Estudios RetrospectivosRESUMEN
The authors report a new case of pneumoblastoma concerning a 63 years old man who dies with hepatitic insufficiency and massive metastasis. The study in optic and electronic microscopy shows the three cellular contingents which form the pneumoblastoma: clusters of indifferenciated cells, spindle cells and cubic or cylinder epithelial cells, with few secretory granules and microvilli which form tubes or lobules. All the features show clearly a blastematous character very similar to a foetal lung which allows to differenciate the pneumoblastoma from carcinosarcoma.