Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 28
Filtrar
2.
Cell Death Dis ; 15(3): 224, 2024 Mar 18.
Artículo en Inglés | MEDLINE | ID: mdl-38494482

RESUMEN

Microenvironmental signals strongly influence chronic lymphocytic leukemia (CLL) cells through the activation of distinct membrane receptors, such as B-cell receptors, and inflammatory receptors, such as Toll-like receptors (TLRs). Inflammatory pathways downstream of these receptors lead to NF-κB activation, thus protecting leukemic cells from apoptosis. Dimethyl fumarate (DMF) is an anti-inflammatory and immunoregulatory drug used to treat patients with multiple sclerosis and psoriasis in which it blocks aberrant NF-κB pathways and impacts the NRF2 antioxidant circuit. Our in vitro analysis demonstrated that increasing concentrations of DMF reduce ATP levels and lead to the apoptosis of CLL cells, including cell lines, splenocytes from Eµ-TCL1-transgenic mice, and primary leukemic cells isolated from the peripheral blood of patients. DMF showed a synergistic effect in association with BTK inhibitors in CLL cells. DMF reduced glutathione levels and activated the NRF2 pathway; gene expression analysis suggested that DMF downregulated pathways related to NFKB and inflammation. In primary leukemic cells, DMF disrupted the TLR signaling pathways induced by CpG by reducing the mRNA expression of NFKBIZ, IL6, IL10 and TNFα. Our data suggest that DMF targets a vulnerability of CLL cells linked to their inflammatory pathways, without impacting healthy donor peripheral blood mononuclear cells.


Asunto(s)
Leucemia Linfocítica Crónica de Células B , Ratones , Animales , Humanos , Leucemia Linfocítica Crónica de Células B/genética , Dimetilfumarato/farmacología , Dimetilfumarato/uso terapéutico , FN-kappa B/metabolismo , Leucocitos Mononucleares/metabolismo , Factor 2 Relacionado con NF-E2/genética , Factor 2 Relacionado con NF-E2/metabolismo , Apoptosis , Ratones Transgénicos
3.
Eur Urol ; 85(5): 417-421, 2024 May.
Artículo en Inglés | MEDLINE | ID: mdl-38184414

RESUMEN

Neoadjuvant pembrolizumab has been shown to be a valid treatment for patients affected by muscle-invasive bladder cancer (MIBC), as demonstrated in the PURE-01 clinical trial (NCT02736266). Among the tumor-extrinsic factors influencing immunotherapy efficacy, extensive data highlighted that the microbiome is a central player in immune-mediated anticancer activity. This report aimed to investigate the composition and role of stool microbiome in patients enrolled in the PURE-01 clinical trial. An orthotopic animal model of bladder cancer (MB49-Luc) was used to support some of the findings from human data. An analysis of stool microbiome before pembrolizumab was conducted for 42 patients, of whom 23 showed a pathologic response. The information in the preclinical model of orthotopic bladder cancer treated with anti-PD-1 antibody or control isotype was validated. Linear discriminant analysis effect size and linear models were used to identify the bacterial taxa enriched in either responders or nonresponders. The identified taxa were also tested for their association with event-free survival (EFS). Survival at 31 d after tumor instillation was used as the study endpoint in the preclinical model. Responders and nonresponders emerged to differ in terms of enrichment for 16 bacterial taxa. Of these, the genus Sutterella was enriched in responders, while the species Ruminococcus bromii was enriched in nonresponders. The negative impact of R. bromii on anti-PD-1 antibody activity was also observed in the preclinical model. EFS and survival of the preclinical model showed a negative role of R. bromii. We found different stool bacterial taxa associated with the response or lack of response to neoadjuvant pembrolizumab. Moreover, we provided experimental data about the negative role of R. bromii on immunotherapy response. Further studies are needed to externally validate our findings and provide mechanistic insights about the host-pathogen interactions in MIBC. PATIENT SUMMARY: Using prepembrolizumab stool samples collected from patients enrolled in the PURE-01 clinical trials, we identified some bacterial taxa that were enriched in patients who either responded or did not respond to immunotherapy. Using an animal model of bladder cancer, we gathered further evidence of the negative impact of the Ruminococcus bromii on immunotherapy efficacy. Further studies are needed to confirm the current findings and test the utility of these bacteria as predictive markers of immunotherapy response.


Asunto(s)
Anticuerpos Monoclonales Humanizados , Terapia Neoadyuvante , Ruminococcus , Neoplasias de la Vejiga Urinaria , Animales , Humanos , Neoplasias de la Vejiga Urinaria/patología , Músculos/patología
4.
FEBS Open Bio ; 13(12): 2367-2374, 2023 12.
Artículo en Inglés | MEDLINE | ID: mdl-37881888

RESUMEN

Chronic lymphocytic leukemia (CLL) is a prototypic neoplasia in which malignant cells strongly depend on microenvironmental stimulations in the lymphoid tissues where they accumulate; leukemic cells are exposed to interaction with bystander and accessory cells, as well as inflammatory soluble mediators. Cell lines are frequently used to model the pathobiology of this disease; however, they do not always recapitulate leukemic cell growth and response to stimulation, and no data are available on Toll-like receptors (TLR) signaling in CLL cell lines. To address this gap, we analyzed HG3, MEC2, and PCL12 cell lines, before and after CpG stimulation, by RNA-sequencing followed by bioinformatic analyses and validation experiments. We identified NFKBIZ mRNA and the corresponding IkBz protein as robust markers of TLR9 activation in both MEC2 and PCL12, but not in HG3 cells. Next, we compared our current results with previous results obtained with primary CLL patient samples and were able to conclude that MEC2 is most similar to the patients' cells in terms of global responsiveness to TLR stimulation; in particular, MEC2 better resembles the samples of patients, as it is characterized by high expression levels of IkBz, but with a lower number of genes regulated.


Asunto(s)
Leucemia Linfocítica Crónica de Células B , Humanos , Línea Celular , Leucemia Linfocítica Crónica de Células B/genética , Leucemia Linfocítica Crónica de Células B/metabolismo , Leucemia Linfocítica Crónica de Células B/patología , Transducción de Señal , Receptor Toll-Like 9/genética , Receptor Toll-Like 9/metabolismo , Receptores Toll-Like/genética , Receptores Toll-Like/metabolismo
5.
Eur J Immunol ; 53(12): e2350529, 2023 12.
Artículo en Inglés | MEDLINE | ID: mdl-37741290

RESUMEN

TDC are hematopoietic cells that combine dendritic cell (DC) and conventional T-cell markers and functional properties. They were identified in secondary lymphoid organs (SLOs) of naïve mice as cells expressing CD11c, major histocompatibility molecules (MHC)-II, and the T-cell receptor (TCR). Despite thorough characterization, a physiological role for TDC remains to be determined. Unfortunately, using CD11c as a marker for TDC has the caveat of its upregulation on different cells, including T cells, upon activation. Here, we took advantage of Zbtb46-GFP reporter mice to explore the frequency and localization of TDC in different tissues at steady state and upon viral infection. RNA sequencing analysis confirmed that TDC sorted from Zbtb46-GFP mice have a gene signature that is distinct from conventional T cells and DC. In addition, this reporter model allowed for identification of TDC in situ not only in SLOs but also in the liver and lung of naïve mice. Interestingly, we found that TDC numbers in the SLOs increased upon viral infection, suggesting that TDC might play a role during viral infections. In conclusion, we propose a visualization strategy that might shed light on the physiological role of TDC in several pathological contexts, including infection and cancer.


Asunto(s)
Linfocitos T , Virosis , Ratones , Animales , Células Dendríticas/patología , Antígeno CD11c , Ratones Endogámicos C57BL
6.
Eur J Immunol ; 53(10): e2350418, 2023 10.
Artículo en Inglés | MEDLINE | ID: mdl-37561992

RESUMEN

Chronic lymphocytic leukemia (CLL) co-evolves with its own microenvironment where inflammatory stimuli including toll-like receptors (TLR) signaling can protect CLL cells from spontaneous and drug-induced apoptosis by upregulating IκBζ, an atypical co-transcription factor. To dissect IκBζ-centered signaling pathways, we performed a gene expression profile of primary leukemic cells expressing either high or low levels of IκBζ after stimulation, highlighting that IκBζ is not only an inflammatory gene but it may control metabolic rewiring of malignant cells thus pointing to a novel potential opportunity for therapy. We exploited the capacity of the dimethyl itaconate (DI), an anti-inflammatory electrophilic synthetic derivative of the metabolite Itaconate, to target IκBζ. CLL cells, murine leukemic splenocytes, and leukocytes from healthy donors were treated in vitro with DI that abolished metabolic activation and reduced cell viability of leukemic cells only, even in the presence of robust TLR prestimulation. RNA sequencing highlighted that in addition to the expected electrophilic stress signature observed after DI treatment, novel pathways emerged including the downregulation of distinct MHC class II complex genes. In conclusion, DI not only abrogated the proinflammatory effects of TLR stimulation but also targeted a specific metabolic vulnerability in CLL cells.


Asunto(s)
Leucemia Linfocítica Crónica de Células B , Animales , Ratones , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Leucemia Linfocítica Crónica de Células B/genética , Transducción de Señal/genética , Receptores Toll-Like/metabolismo , Redes y Vías Metabólicas , Microambiente Tumoral
7.
EBioMedicine ; 91: 104567, 2023 May.
Artículo en Inglés | MEDLINE | ID: mdl-37062177

RESUMEN

BACKGROUND: In preclinical models of Type 1 Diabetes (T1D) the integrity of the gut barrier (GB) is instrumental to avoid dysregulated crosstalk between the commensal microbiota and immune cells and to prevent autoimmunity. The GB is composed of the intestinal epithelial barrier (IEB) and of the mucus layer containing mucins and antimicrobial peptides (AMPs) that are crucial to maintain immune tolerance. In preclinical models of T1D the alterations of the GB primarily affect the mucus layer. In human T1D increased gut permeability and IEB damage have been demonstrated but the integrity of the mucus layer was never assessed. METHODS: We evaluated GB integrity by measuring serological markers of IEB damage (serological levels of zonulin) and bacterial translocation such as lipopolysaccharide binding protein (LBP) and myeloid differentiation protein 2 (MD2), and mRNA expression of tight junction proteins, mucins and AMPs in intestinal tissue of T1D patients and healthy controls (HC). Simultaneously, we performed immunological profiling on intestinal tissue and 16S rRNA analysis on the mucus-associated gut microbiota (MAGM). FINDINGS: Our data show a GB damage with mucus layer alterations and reduced mRNA expression of several mucins (MUC2, MUC12, MUC13, MUC15, MUC20, MUC21) and AMPs (HD4 and HD5) in T1D patients. Mucus layer alterations correlated with reduced relative abundance of short chain fatty acids (SCFA)-producing bacteria such as Bifidobacterium dentium, Clostridium butyricum and Roseburia intestinalis that regulate mucin expression and intestinal immune homeostasis. In T1D patients we also found intestinal immune dysregulation with higher percentages of effector T cells such as T helper (Th) 1, Th17 and TNF-α+ T cells. INTERPRETATION: Our data show that mucus layer alterations are present in T1D subjects and associated with dysbiosis and immune dysregulation. FUNDING: Research Grants from the Juvenile Diabetes Foundation (Grant 1-INO-2018-640-A-N to MF and 2-SRA-2019-680-S-B to JD) and from the Italian Ministry of Health (Grant RF19-12370721 to MF).


Asunto(s)
Diabetes Mellitus Tipo 1 , Humanos , Mucosa Intestinal/metabolismo , Disbiosis/metabolismo , ARN Ribosómico 16S/metabolismo , Mucinas/metabolismo , Moco/metabolismo , ARN Mensajero/metabolismo
8.
Cell Death Dis ; 14(2): 129, 2023 02 15.
Artículo en Inglés | MEDLINE | ID: mdl-36792589

RESUMEN

Lipid and cholesterol metabolism play a crucial role in tumor cell behavior and in shaping the tumor microenvironment. In particular, enzymatic and non-enzymatic cholesterol metabolism, and derived metabolites control dendritic cell (DC) functions, ultimately impacting tumor antigen presentation within and outside the tumor mass, dampening tumor immunity and immunotherapeutic attempts. The mechanisms accounting for such events remain largely to be defined. Here we perturbed (oxy)sterol metabolism genetically and pharmacologically and analyzed the tumor lipidome landscape in relation to the tumor-infiltrating immune cells. We report that perturbing the lipidome of tumor microenvironment by the expression of sulfotransferase 2B1b crucial in cholesterol and oxysterol sulfate synthesis, favored intratumoral representation of monocyte-derived antigen-presenting cells, including monocyte-DCs. We also found that treating mice with a newly developed antagonist of the oxysterol receptors Liver X Receptors (LXRs), promoted intratumoral monocyte-DC differentiation, delayed tumor growth and synergized with anti-PD-1 immunotherapy and adoptive T cell therapy. Of note, looking at LXR/cholesterol gene signature in melanoma patients treated with anti-PD-1-based immunotherapy predicted diverse clinical outcomes. Indeed, patients whose tumors were poorly infiltrated by monocytes/macrophages expressing LXR target genes showed improved survival over the course of therapy. Thus, our data support a role for (oxy)sterol metabolism in shaping monocyte-to-DC differentiation, and in tumor antigen presentation critical for responsiveness to immunotherapy. The identification of a new LXR antagonist opens new treatment avenues for cancer patients.


Asunto(s)
Melanoma , Monocitos , Ratones , Animales , Monocitos/metabolismo , Diferenciación Celular , Colesterol/metabolismo , Presentación de Antígeno , Células Dendríticas/metabolismo , Microambiente Tumoral
9.
Front Oncol ; 12: 894413, 2022.
Artículo en Inglés | MEDLINE | ID: mdl-35814450

RESUMEN

Renal cell carcinoma is highly inflamed, and tumor cells are embedded into a microenvironment enriched with IL1. While inflammatory pathways are well characterized in the immune system, less is known about these same pathways in epithelial cells; it is unclear if and how innate immune signals directly impact on cancer cells, and if we could we manipulate these for therapeutic purposes. To address these questions, we first focused on the inflammatory receptors belonging to the IL1- and Toll-like receptor family including negative regulators in a small cohort of 12 clear cell RCC (ccRCC) patients' samples as compared to their coupled adjacent normal tissues. Our data demonstrated that renal epithelial cancer cells showed a specific and distinctive pattern of inflammatory receptor expression marked by a consistent downregulation of the inhibitory receptor SIGIRR mRNA. This repression was confirmed at the protein level in both cancer cell lines and primary tissues. When we analyzed in silico data of different kidney cancer histotypes, we identified the clear cell subtype as the one where SIGIRR was mostly downregulated; nonetheless, papillary and chromophobe tumor types also showed low levels as compared to their normal counterpart. RNA-sequencing analysis demonstrated that IL1 stimulation of the ccRCC cell line A498 triggered an intrinsic signature of inflammatory pathway activation characterized by the induction of distinct "pro-tumor" genes including several chemokines, the autocrine growth factor IL6, the atypical co-transcription factor NFKBIZ, and the checkpoint inhibitor PD-L1. When we looked for the macroareas most represented among the differentially expressed genes, additional clusters emerged including pathways involved in cell differentiation, angiogenesis, and wound healing. To note, SIGIRR overexpression in A498 cells dampened IL1 signaling as assessed by a reduced induction of NFKBIZ. Our results suggest that SIGIRR downregulation unleashes IL1 signaling intrinsic to tumor cells and that manipulating this pathway may be beneficial in ccRCC.

11.
Development ; 147(22)2020 11 30.
Artículo en Inglés | MEDLINE | ID: mdl-33046507

RESUMEN

The choroid plexus (ChP) is a secretory tissue that produces cerebrospinal fluid (CSF) secreted into the ventricular system. It is a monolayer of secretory, multiciliated epithelial cells derived from neuroepithelial progenitors and overlying a stroma of mesenchymal cells of mesodermal origin. Zfp423, which encodes a Kruppel-type zinc-finger transcription factor essential for cerebellar development and mutated in rare cases of cerebellar vermis hypoplasia/Joubert syndrome and other ciliopathies, is expressed in the hindbrain roof plate, from which the IV ventricle ChP arises, and, later, in mesenchymal cells, which give rise to the stroma and leptomeninges. Mouse Zfp423 mutants display a marked reduction of the hindbrain ChP (hChP), which: (1) fails to express established markers of its secretory function and genes implicated in its development and maintenance (Lmx1a and Otx2); (2) shows a perturbed expression of signaling pathways previously unexplored in hChP patterning (Wnt3); and (3) displays a lack of multiciliated epithelial cells and a profound dysregulation of master genes of multiciliogenesis (Gmnc). Our results propose that Zfp423 is a master gene and one of the earliest known determinants of hChP development.


Asunto(s)
Plexo Coroideo/embriología , Proteínas de Unión al ADN/metabolismo , Rombencéfalo/embriología , Factores de Transcripción/metabolismo , Animales , Plexo Coroideo/citología , Proteínas de Unión al ADN/genética , Proteínas con Homeodominio LIM/genética , Proteínas con Homeodominio LIM/metabolismo , Ratones , Ratones Mutantes , Factores de Transcripción Otx/genética , Factores de Transcripción Otx/metabolismo , Rombencéfalo/citología , Factores de Transcripción/genética , Proteína Wnt3/genética , Proteína Wnt3/metabolismo
12.
Front Med (Lausanne) ; 6: 263, 2019.
Artículo en Inglés | MEDLINE | ID: mdl-31803746

RESUMEN

The emergence of data coming from different venues, as several "omic" approaches, is providing already compelling evidence that the smart use of this information could provide invaluable information to prevent, diagnose and treat human diseases. However, the most daunting challenges remain ahead, as the explosive accumulation of data from additional perspectives, including social graphs, biosensors, and imaging, promise to deliver crucial information that could be exploited for the improvement of the entire human race, both in developed, and developing countries, optimizing health expenses and reaching also the less fortunate sections of the societies. And yet, formidable challenges remain, that pertain for the most part to the collection of the data, their organization, and most relevantly their integration. Here we provide few, pointed examples to the present relevance of these big data approaches in human health as well potential road maps toward the implementation of broader data collections and analyses.

13.
Nat Med ; 25(4): 603-611, 2019 04.
Artículo en Inglés | MEDLINE | ID: mdl-30911134

RESUMEN

Transplantation of hematopoietic cells from a healthy individual (allogeneic hematopoietic cell transplantation (allo-HCT)) demonstrates that adoptive immunotherapy can cure blood cancers: still, post-transplantation relapses remain frequent. To explain their drivers, we analyzed the genomic and gene expression profiles of acute myeloid leukemia (AML) blasts purified from patients at serial time-points during their disease history. We identified a transcriptional signature specific for post-transplantation relapses and highly enriched in immune-related processes, including T cell costimulation and antigen presentation. In two independent patient cohorts we confirmed the deregulation of multiple costimulatory ligands on AML blasts at post-transplantation relapse (PD-L1, B7-H3, CD80, PVRL2), mirrored by concomitant changes in circulating donor T cells. Likewise, we documented the frequent loss of surface expression of HLA-DR, -DQ and -DP on leukemia cells, due to downregulation of the HLA class II regulator CIITA. We show that loss of HLA class II expression and upregulation of inhibitory checkpoint molecules represent alternative modalities to abolish AML recognition from donor-derived T cells, and can be counteracted by interferon-γ or checkpoint blockade, respectively. Our results demonstrate that the deregulation of pathways involved in T cell-mediated allorecognition is a distinctive feature and driver of AML relapses after allo-HCT, which can be rapidly translated into personalized therapies.


Asunto(s)
Perfilación de la Expresión Génica , Trasplante de Células Madre Hematopoyéticas , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/inmunología , Regulación Leucémica de la Expresión Génica , Antígenos de Histocompatibilidad Clase II/genética , Antígenos de Histocompatibilidad Clase II/metabolismo , Humanos , Leucemia Mieloide Aguda/terapia , Activación de Linfocitos/inmunología , ARN Mensajero/genética , ARN Mensajero/metabolismo , Recurrencia , Reproducibilidad de los Resultados , Trasplante Homólogo
14.
J Immunother Cancer ; 7(1): 45, 2019 02 13.
Artículo en Inglés | MEDLINE | ID: mdl-30760333

RESUMEN

BACKGROUND: The thymic stromal lymphopoietin (TSLP), a key cytokine for development of Th2 immunity, is produced by cancer associated fibroblasts (CAFs) in pancreatic cancer where predominant tumor infiltrating Th2 over Th1 cells correlates with reduced patients' survival. Which cells and molecules are mostly relevant in driving TSLP secretion by CAFs in pancreatic cancer is not defined. METHODS: We performed in vitro, in vivo and ex-vivo analyses. For in vitro studies we used pancreatic cancer cell lines, primary CAFs cultures, and THP1 cells. TSLP secretion by CAFs was used as a read-out system to identify in vitro relevant tumor-derived inflammatory cytokines and molecules. For in vivo studies human pancreatic cancer cells and CAFs were orthotopically injected in immunodeficient mice. For ex-vivo studies immunohistochemistry was performed to detect ASC (apoptosis-associated speck-like protein containing a caspase recruitment domain) expression in surgical samples. Bioinformatics was applied to interrogate published data sets. RESULTS: We show in vitro that IL-1α and IL-1ß released by pancreatic cancer cells and tumor cell-conditioned macrophages are crucial for TSLP secretion by CAFs. Treatment of immunodeficient mice orthotopically injected with human IL-1 positive pancreatic cancer cells plus CAFs using the IL-1R antagonist anakinra significantly reduced TSLP expression in the tumor. Importantly, we found that pancreatic cancer cells release alarmins, among which ASC, able to induce IL-1ß secretion in macrophages. The relevance of ASC was confirmed ex-vivo by its expression in both tumor cells and tumor associated macrophages in pancreatic cancer surgical samples and survival data analyses showing statistically significant inverse correlation between ASC expression and survival in pancreatic cancer patients. CONCLUSIONS: Our findings indicate that tumor released IL-1α and IL-1ß and ASC are key regulators of TSLP secretion by CAFs and their targeting should ultimately dampen Th2 inflammation and improve overall survival in pancreatic cancer.


Asunto(s)
Proteínas Adaptadoras de Señalización CARD/metabolismo , Fibroblastos Asociados al Cáncer/metabolismo , Citocinas/metabolismo , Interleucina-1alfa/metabolismo , Interleucina-1beta/metabolismo , Neoplasias Pancreáticas/metabolismo , Animales , Proteínas Adaptadoras de Señalización CARD/genética , Línea Celular Tumoral , Humanos , Inflamasomas/metabolismo , Interleucina-1alfa/genética , Interleucina-1beta/genética , Ratones , Neoplasias Pancreáticas/genética , Receptores de Interleucina-1/metabolismo , Células THP-1 , Linfopoyetina del Estroma Tímico
15.
PLoS One ; 13(7): e0200783, 2018.
Artículo en Inglés | MEDLINE | ID: mdl-30020994

RESUMEN

In a variety of species, reduced food intake, and in particular protein or amino acid (AA) restriction, extends lifespan and healthspan. However, the underlying epigenetic and/or transcriptional mechanisms are largely unknown, and dissection of specific pathways in cultured cells may contribute to filling this gap. We have previously shown that, in mammalian cells, deprivation of essential AAs (methionine/cysteine or tyrosine) leads to the transcriptional reactivation of integrated silenced transgenes, including plasmid and retroviral vectors and latent HIV-1 provirus, by a process involving epigenetic chromatic remodeling and histone acetylation. Here we show that the deprivation of methionine/cysteine also leads to the transcriptional upregulation of endogenous retroviruses, suggesting that essential AA starvation affects the expression not only of exogenous non-native DNA sequences, but also of endogenous anciently-integrated and silenced parasitic elements of the genome. Moreover, we show that the transgene reactivation response is highly conserved in different mammalian cell types, and it is reproducible with deprivation of most essential AAs. The General Control Non-derepressible 2 (GCN2) kinase and the downstream integrated stress response represent the best candidates mediating this process; however, by pharmacological approaches, RNA interference and genomic editing, we demonstrate that they are not implicated. Instead, the response requires MEK/ERK and/or JNK activity and is reproduced by ribosomal inhibitors, suggesting that it is triggered by a novel nutrient-sensing and signaling pathway, initiated by translational block at the ribosome, and independent of mTOR and GCN2. Overall, these findings point to a general transcriptional response to essential AA deprivation, which affects the expression of non-native genomic sequences, with relevant implications for the epigenetic/transcriptional effects of AA restriction in health and disease.


Asunto(s)
Aminoácidos Esenciales/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Aminoácidos Esenciales/deficiencia , Animales , Western Blotting , Sistemas CRISPR-Cas , Línea Celular , Edición Génica , Células HeLa , Células Hep G2 , Humanos , Ratones , Proteínas Serina-Treonina Quinasas/genética , Interferencia de ARN , Reacción en Cadena en Tiempo Real de la Polimerasa , Transducción de Señal/genética , Transducción de Señal/fisiología , Activación Transcripcional/genética , Activación Transcripcional/fisiología
16.
Sci Rep ; 7(1): 7383, 2017 08 07.
Artículo en Inglés | MEDLINE | ID: mdl-28785050

RESUMEN

Autosomal dominant tubulointerstitial kidney disease (ADTKD) is an inherited disorder that causes progressive kidney damage and renal failure. Mutations in the UMOD gene, encoding uromodulin, lead to ADTKD-UMOD related. Uromodulin is a GPI-anchored protein exclusively produced by epithelial cells of the thick ascending limb of Henle's loop. It is released in the tubular lumen after proteolytic cleavage and represents the most abundant protein in human urine in physiological condition. We previously generated and characterized a transgenic mouse model expressing mutant uromodulin (Tg UmodC147W) that recapitulates the main features of ATDKD-UMOD. While several studies clearly demonstrated that mutated uromodulin accumulates in endoplasmic reticulum, the mechanisms that lead to renal damage are not fully understood. In our work, we used kidney transcriptional profiling to identify early events of pathogenesis in the kidneys of Tg UmodC147W mice. Our results demonstrate up-regulation of inflammation and fibrosis and down-regulation of lipid metabolism in young Tg UmodC147W mice, before any functional or histological evidence of kidney damage. We also show that pro-inflammatory signals precede fibrosis onset and are already present in the first week after birth. Early induction of inflammation is likely relevant for ADTKD-UMOD pathogenesis and related pathways can be envisaged as possible novel targets for therapeutic intervention.


Asunto(s)
Perfilación de la Expresión Génica/métodos , Redes Reguladoras de Genes , Mutación , Nefritis Intersticial/genética , Uromodulina/genética , Animales , Modelos Animales de Enfermedad , Retículo Endoplásmico/metabolismo , Estrés del Retículo Endoplásmico , Femenino , Regulación de la Expresión Génica , Humanos , Metabolismo de los Lípidos , Masculino , Ratones , Ratones Transgénicos , Nefritis Intersticial/metabolismo , Uromodulina/metabolismo
17.
EMBO Mol Med ; 9(9): 1198-1211, 2017 09.
Artículo en Inglés | MEDLINE | ID: mdl-28667090

RESUMEN

Clinical application of lentiviral vector (LV)-based hematopoietic stem and progenitor cells (HSPC) gene therapy is rapidly becoming a reality. Nevertheless, LV-mediated signaling and its potential functional consequences on HSPC biology remain poorly understood. We unravel here a remarkably limited impact of LV on the HSPC transcriptional landscape. LV escaped innate immune sensing that instead led to robust IFN responses upon transduction with a gamma-retroviral vector. However, reverse-transcribed LV DNA did trigger p53 signaling, activated also by non-integrating Adeno-associated vector, ultimately leading to lower cell recovery ex vivo and engraftment in vivo These effects were more pronounced in the short-term repopulating cells while long-term HSC frequencies remained unaffected. Blocking LV-induced signaling partially rescued both apoptosis and engraftment, highlighting a novel strategy to further dampen the impact of ex vivo gene transfer on HSPC. Overall, our results shed light on viral vector sensing in HSPC and provide critical insight for the development of more stealth gene therapy strategies.


Asunto(s)
Terapia Genética , Vectores Genéticos/genética , Células Madre Hematopoyéticas/inmunología , Lentivirus/genética , Proteína p53 Supresora de Tumor/inmunología , Animales , Vectores Genéticos/inmunología , Trasplante de Células Madre Hematopoyéticas , Humanos , Inmunidad Innata , Lentivirus/inmunología , Ratones , Proteína p53 Supresora de Tumor/genética
18.
Cancer Res ; 77(3): 658-671, 2017 02 01.
Artículo en Inglés | MEDLINE | ID: mdl-27872095

RESUMEN

Donor-derived allogeneic T cells evoke potent graft versus tumor (GVT) effects likely due to the simultaneous recognition of tumor-specific and host-restricted minor histocompatibility (H) antigens. Here we investigated whether such effects could be reproduced in autologous settings by TCR gene-engineered lymphocytes. We report that T cells redirected either to a broadly expressed Y-encoded minor H antigen or to a tumor-associated antigen, although poorly effective if individually transferred, when simultaneously administered enabled acute autochthonous tumor debulking and resulted in durable clinical remission. Y-redirected T cells proved hyporesponsive in peripheral lymphoid organs, whereas they retained effector function at the tumor site, where in synergy with tumor-redirected lymphocytes, they instructed TNFα expression, endothelial cell activation, and intratumoral T-cell infiltration. While neutralizing TNFα hindered GVT effects by the combined T-cell infusion, a single injection of picogram amounts of NGR-TNF, a tumor vessel-targeted TNFα derivative currently in phase III clinical trials, substituted for Y-redirected cells and enabled tumor debulking by tumor-redirected lymphocytes. Together, our results provide new mechanistic insights into allogeneic GVT, validate the importance of targeting the tumor and its associated stroma, and prove the potency of a novel combined approach suitable for immediate clinical implementation. Cancer Res; 77(3); 658-71. ©2016 AACR.


Asunto(s)
Adenocarcinoma/patología , Linfocitos T CD8-positivos/trasplante , Inmunoterapia Adoptiva/métodos , Antígenos de Histocompatibilidad Menor/inmunología , Neoplasias de la Próstata/patología , Factor de Necrosis Tumoral alfa/biosíntesis , Animales , Linfocitos T CD8-positivos/inmunología , Citometría de Flujo , Técnica del Anticuerpo Fluorescente , Perfilación de la Expresión Génica , Inmunohistoquímica , Masculino , Ratones , Ratones Endogámicos C57BL , Reacción en Cadena en Tiempo Real de la Polimerasa
19.
Proc Natl Acad Sci U S A ; 113(51): E8286-E8295, 2016 12 20.
Artículo en Inglés | MEDLINE | ID: mdl-27930306

RESUMEN

Invariant natural killer T cells (iNKT) cells are T lymphocytes displaying innate effector functions, acquired through a distinct thymic developmental program regulated by microRNAs (miRNAs). Deleting miRNAs by Dicer ablation (Dicer KO) in thymocytes selectively impairs iNKT cell survival and functional differentiation. To unravel this miRNA-dependent program, we systemically identified transcripts that were differentially expressed between WT and Dicer KO iNKT cells at different differentiation stages and predicted to be targeted by the iNKT cell-specific miRNAs. TGF-ß receptor II (TGF-ßRII), critically implicated in iNKT cell differentiation, was found up-regulated in iNKT Dicer KO cells together with enhanced TGF-ß signaling. miRNA members of the miR-17∼92 family clusters were predicted to target Tgfbr2 mRNA upon iNKT cell development. iNKT cells lacking all three miR-17∼92 family clusters (miR-17∼92, miR-106a∼363, miR-106b∼25) phenocopied both increased TGF-ßRII expression and signaling, and defective effector differentiation, displayed by iNKT Dicer KO cells. Consistently, genetic ablation of TGF-ß signaling in the absence of miRNAs rescued iNKT cell differentiation. These results elucidate the global impact of miRNAs on the iNKT cell developmental program and uncover the targeting of a lineage-specific cytokine signaling by miRNAs as a mechanism regulating innate-like T-cell development and effector differentiation.


Asunto(s)
MicroARNs/genética , Células T Asesinas Naturales/citología , Factor de Crecimiento Transformador beta/metabolismo , Animales , Antígenos CD1d/metabolismo , Diferenciación Celular , Citocinas/metabolismo , ARN Helicasas DEAD-box/genética , Perfilación de la Expresión Génica , Prueba de Complementación Genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Familia de Multigenes , Fenotipo , ARN Mensajero/metabolismo , Ribonucleasa III/genética , Transducción de Señal , Timo/metabolismo
20.
Sci Rep ; 6: 24647, 2016 Apr 21.
Artículo en Inglés | MEDLINE | ID: mdl-27097888

RESUMEN

Systems biology provides opportunities to fully understand the genes and pathways in disease pathogenesis. We used literature knowledge and unbiased multiple data meta-analysis paradigms to analyze microarray datasets across different mouse strains and acute allergic asthma models. Our combined gene-driven and pathway-driven strategies generated a stringent signature list totaling 933 genes with 41% (440) asthma-annotated genes and 59% (493) ignorome genes, not previously associated with asthma. Within the list, we identified inflammation, circadian rhythm, lung-specific insult response, stem cell proliferation domains, hubs, peripheral genes, and super-connectors that link the biological domains (Il6, Il1ß, Cd4, Cd44, Stat1, Traf6, Rela, Cadm1, Nr3c1, Prkcd, Vwf, Erbb2). In conclusion, this novel bioinformatics approach will be a powerful strategy for clinical and across species data analysis that allows for the validation of experimental models and might lead to the discovery of novel mechanistic insights in asthma.


Asunto(s)
Asma/genética , Asma/metabolismo , Redes Reguladoras de Genes , Transducción de Señal , Enfermedad Aguda , Análisis por Conglomerados , Biología Computacional/métodos , Perfilación de la Expresión Génica , Regulación de la Expresión Génica , Humanos
SELECCIÓN DE REFERENCIAS
DETALLE DE LA BÚSQUEDA