RESUMEN
Tissue fibrosis is a major cause of mortality that results from the deposition of matrix proteins by an activated mesenchyme. Macrophages accumulate in fibrosis, but the role of specific subgroups in supporting fibrogenesis has not been investigated in vivo. Here, we used single-cell RNA sequencing (scRNA-seq) to characterize the heterogeneity of macrophages in bleomycin-induced lung fibrosis in mice. A novel computational framework for the annotation of scRNA-seq by reference to bulk transcriptomes (SingleR) enabled the subclustering of macrophages and revealed a disease-associated subgroup with a transitional gene expression profile intermediate between monocyte-derived and alveolar macrophages. These CX3CR1+SiglecF+ transitional macrophages localized to the fibrotic niche and had a profibrotic effect in vivo. Human orthologs of genes expressed by the transitional macrophages were upregulated in samples from patients with idiopathic pulmonary fibrosis. Thus, we have identified a pathological subgroup of transitional macrophages that are required for the fibrotic response to injury.
Asunto(s)
Fibrosis Pulmonar Idiopática/inmunología , Pulmón/patología , Activación de Macrófagos , Macrófagos Alveolares/inmunología , Animales , Antígenos de Diferenciación Mielomonocítica/genética , Antígenos de Diferenciación Mielomonocítica/inmunología , Antígenos de Diferenciación Mielomonocítica/metabolismo , Bleomicina/inmunología , Receptor 1 de Quimiocinas CX3C/genética , Receptor 1 de Quimiocinas CX3C/inmunología , Receptor 1 de Quimiocinas CX3C/metabolismo , Células Cultivadas , Modelos Animales de Enfermedad , Femenino , Perfilación de la Expresión Génica/métodos , Humanos , Fibrosis Pulmonar Idiopática/patología , Pulmón/citología , Pulmón/inmunología , Macrófagos Alveolares/metabolismo , Masculino , Ratones , Análisis de Secuencia de ARN/métodos , Lectinas Similares a la Inmunoglobulina de Unión a Ácido Siálico , Análisis de la Célula Individual/métodos , Regulación hacia ArribaRESUMEN
To determine whether double-strand break (DSB) mobility enhances the physical search for an ectopic template during homology-directed repair (HDR), we tested the effects of factors that control chromatin dynamics, including cohesin loading and kinetochore anchoring. The former but not the latter is altered in response to DSBs. Loss of the nonhistone high-mobility group protein Nhp6 reduces histone occupancy and increases chromatin movement, decompaction, and ectopic HDR. The loss of nucleosome remodeler INO80-C did the opposite. To see whether enhanced HDR depends on DSB mobility or the global chromatin response, we tested the ubiquitin ligase mutant uls1Δ, which selectively impairs local but not global movement in response to a DSB. Strand invasion occurs in uls1Δ cells with wild-type kinetics, arguing that global histone depletion rather than DSB movement is rate limiting for HDR. Impaired break movement in uls1Δ correlates with elevated MRX and cohesin loading, despite normal resection and checkpoint activation.
Asunto(s)
Roturas del ADN de Doble Cadena , Nucleosomas/metabolismo , Saccharomyces cerevisiae/metabolismo , Bleomicina/farmacología , Ciclo Celular , Proteínas de Ciclo Celular/metabolismo , Centrómero/metabolismo , Cromatina/metabolismo , Ensamble y Desensamble de Cromatina , Proteínas Cromosómicas no Histona/metabolismo , ADN de Hongos/metabolismo , Histonas/metabolismo , Modelos Biológicos , Fosforilación , Saccharomyces cerevisiae/citología , Proteínas de Saccharomyces cerevisiae/metabolismo , Cuerpos Polares del Huso/metabolismo , CohesinasRESUMEN
Double-strand breaks (DSBs) are the most deleterious DNA lesions, which, if left unrepaired, may lead to genome instability or cell death. Here, we report that, in response to DSBs, the RNA methyltransferase METTL3 is activated by ATM-mediated phosphorylation at S43. Phosphorylated METTL3 is then localized to DNA damage sites, where it methylates the N6 position of adenosine (m6A) in DNA damage-associated RNAs, which recruits the m6A reader protein YTHDC1 for protection. In this way, the METTL3-m6A-YTHDC1 axis modulates accumulation of DNA-RNA hybrids at DSBs sites, which then recruit RAD51 and BRCA1 for homologous recombination (HR)-mediated repair. METTL3-deficient cells display defective HR, accumulation of unrepaired DSBs, and genome instability. Accordingly, depletion of METTL3 significantly enhances the sensitivity of cancer cells and murine xenografts to DNA damage-based therapy. These findings uncover the function of METTL3 and YTHDC1 in HR-mediated DSB repair, which may have implications for cancer therapy.
Asunto(s)
Adenosina/análogos & derivados , Neoplasias de Cabeza y Cuello/genética , Metiltransferasas/genética , Proteínas del Tejido Nervioso/genética , Factores de Empalme de ARN/genética , Reparación del ADN por Recombinación/efectos de los fármacos , Carcinoma de Células Escamosas de Cabeza y Cuello/genética , Adenosina/metabolismo , Animales , Antibióticos Antineoplásicos/farmacología , Proteínas de la Ataxia Telangiectasia Mutada/genética , Proteínas de la Ataxia Telangiectasia Mutada/metabolismo , Proteína BRCA1/genética , Proteína BRCA1/metabolismo , Bleomicina/farmacología , Línea Celular Tumoral , ADN/genética , ADN/metabolismo , Células Epiteliales/efectos de los fármacos , Células Epiteliales/metabolismo , Células Epiteliales/patología , Femenino , Células HEK293 , Neoplasias de Cabeza y Cuello/tratamiento farmacológico , Neoplasias de Cabeza y Cuello/mortalidad , Neoplasias de Cabeza y Cuello/patología , Humanos , Metiltransferasas/metabolismo , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Proteínas del Tejido Nervioso/metabolismo , Hibridación de Ácido Nucleico , Osteoblastos/efectos de los fármacos , Osteoblastos/metabolismo , Osteoblastos/patología , Fosforilación , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Factores de Empalme de ARN/metabolismo , Recombinasa Rad51/genética , Recombinasa Rad51/metabolismo , Ribonucleasa H/genética , Ribonucleasa H/metabolismo , Carcinoma de Células Escamosas de Cabeza y Cuello/tratamiento farmacológico , Carcinoma de Células Escamosas de Cabeza y Cuello/mortalidad , Carcinoma de Células Escamosas de Cabeza y Cuello/patología , Análisis de Supervivencia , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Pulmonary fibrosis is a fatal condition characterized by fibroblast and myofibroblast proliferation and collagen deposition. TGF-ß plays a pivotal role in the development of pulmonary fibrosis. Therefore, modulation of TGF-ß signaling is a promising therapeutic strategy for treating pulmonary fibrosis. To date, however, interventions targeting TGF-ß have not shown consistent efficacy. CD109 is a GPI-anchored glycoprotein that binds to TGF-ß receptor I and negatively regulates TGF-ß signaling. However, no studies have examined the role and therapeutic potential of CD109 in pulmonary fibrosis. The purpose of this study was to determine the role and therapeutic value of CD109 in bleomycin-induced pulmonary fibrosis. CD109-transgenic mice overexpressing CD109 exhibited significantly attenuated pulmonary fibrosis, preserved lung function, and reduced lung fibroblasts and myofibroblasts compared with wild-type (WT) mice. CD109-/- mice exhibited pulmonary fibrosis comparable to WT mice. CD109 expression was induced in variety types of cells, including lung fibroblasts and macrophages, upon bleomycin exposure. Recombinant CD109 protein inhibited TGF-ß signaling and significantly decreased ACTA2 expression in human fetal lung fibroblast cells in vitro. Administration of recombinant CD109 protein markedly reduced pulmonary fibrosis in bleomycin-treated WT mice in vivo. Our results suggest that CD109 is not essential for the development of pulmonary fibrosis, but excess CD109 protein can inhibit pulmonary fibrosis development, possibly through suppression of TGF-ß signaling. CD109 is a novel therapeutic candidate for treating pulmonary fibrosis.
Asunto(s)
Fibrosis Pulmonar , Humanos , Ratones , Animales , Fibrosis Pulmonar/metabolismo , Bleomicina/efectos adversos , Factor de Crecimiento Transformador beta/metabolismo , Pulmón/patología , Fibroblastos/metabolismo , Ratones Transgénicos , Factores de Transcripción/metabolismo , Ratones Endogámicos C57BL , Proteínas de Neoplasias/metabolismo , Antígenos CD/metabolismo , Proteínas Ligadas a GPI/metabolismoRESUMEN
Mycobacterium tuberculosis (Mtb) is an obligate human pathogen and the causative agent of tuberculosis1-3. Although Mtb can synthesize vitamin B12 (cobalamin) de novo, uptake of cobalamin has been linked to pathogenesis of tuberculosis2. Mtb does not encode any characterized cobalamin transporter4-6; however, the gene rv1819c was found to be essential for uptake of cobalamin1. This result is difficult to reconcile with the original annotation of Rv1819c as a protein implicated in the transport of antimicrobial peptides such as bleomycin7. In addition, uptake of cobalamin seems inconsistent with the amino acid sequence, which suggests that Rv1819c has a bacterial ATP-binding cassette (ABC)-exporter fold1. Here, we present structures of Rv1819c, which reveal that the protein indeed contains the ABC-exporter fold, as well as a large water-filled cavity of about 7,700 Å3, which enables the protein to transport the unrelated hydrophilic compounds bleomycin and cobalamin. On the basis of these structures, we propose that Rv1819c is a multi-solute transporter for hydrophilic molecules, analogous to the multidrug exporters of the ABC transporter family, which pump out structurally diverse hydrophobic compounds from cells8-11.
Asunto(s)
Transportadoras de Casetes de Unión a ATP/metabolismo , Proteínas Bacterianas/metabolismo , Bleomicina/metabolismo , Mycobacterium tuberculosis/metabolismo , Transportadoras de Casetes de Unión a ATP/química , Transportadoras de Casetes de Unión a ATP/genética , Proteínas Bacterianas/química , Proteínas Bacterianas/genética , Transporte Biológico , Interacciones Hidrofóbicas e Hidrofílicas , Modelos Moleculares , Mycobacterium tuberculosis/química , Mycobacterium tuberculosis/genética , Estructura Cuaternaria de Proteína , Estructura Terciaria de ProteínaRESUMEN
Excessive deposition of extracellular matrix, mainly collagen protein, is the hallmark of organ fibrosis. The molecular mechanisms regulating fibrotic protein biosynthesis are unclear. Here, we find that chemoattractant receptor homologous molecule expressed on TH2 cells (CRTH2), a plasma membrane receptor for prostaglandin D2, is trafficked to the endoplasmic reticulum (ER) membrane in fibroblasts in a caveolin-1-dependent manner. ER-anchored CRTH2 binds the collagen mRNA recognition motif of La ribonucleoprotein domain family member 6 (LARP6) and promotes the degradation of collagen mRNA in these cells. In line, CRTH2 deficiency increases collagen biosynthesis in fibroblasts and exacerbates injury-induced organ fibrosis in mice, which can be rescued by LARP6 depletion. Administration of CRTH2 N-terminal peptide reduces collagen production by binding to LARP6. Similar to CRTH2, bumetanide binds the LARP6 mRNA recognition motif, suppresses collagen biosynthesis, and alleviates bleomycin-triggered pulmonary fibrosis in vivo. These findings reveal a novel anti-fibrotic function of CRTH2 in the ER membrane via the interaction with LARP6, which may represent a therapeutic target for fibrotic diseases.
Asunto(s)
Autoantígenos/metabolismo , Colágeno/antagonistas & inhibidores , Cirrosis Hepática/prevención & control , Fibrosis Pulmonar/prevención & control , Receptores Inmunológicos/metabolismo , Receptores de Prostaglandina/metabolismo , Ribonucleoproteínas/metabolismo , Animales , Bleomicina , Tetracloruro de Carbono , Células Cultivadas , Colágeno/biosíntesis , Colágeno/genética , Retículo Endoplásmico/metabolismo , Fibroblastos/metabolismo , Membranas Intracelulares/metabolismo , Isoproterenol , Hígado/metabolismo , Hígado/patología , Cirrosis Hepática/inducido químicamente , Cirrosis Hepática/metabolismo , Cirrosis Hepática/patología , Pulmón/metabolismo , Pulmón/patología , Masculino , Ratones Transgénicos , Miocardio/metabolismo , Miocardio/patología , Unión Proteica , Fibrosis Pulmonar/inducido químicamente , Fibrosis Pulmonar/metabolismo , Fibrosis Pulmonar/patología , Receptores Inmunológicos/genética , Receptores de Prostaglandina/genética , Antígeno SS-BRESUMEN
BACKGROUND: Five-year follow-up in a trial involving patients with previously untreated stage III or IV classic Hodgkin's lymphoma showed long-term progression-free survival benefits with first-line therapy with brentuximab vedotin, a CD30-directed antibody-drug conjugate, plus doxorubicin, vinblastine, and dacarbazine (A+AVD), as compared with doxorubicin, bleomycin, vinblastine, and dacarbazine (ABVD). A planned interim analysis indicated a potential benefit with regard to overall survival; data from a median of 6 years of follow-up are now available. METHODS: We randomly assigned patients in a 1:1 ratio to receive up to six cycles of A+AVD or ABVD. The primary end point, modified progression-free survival, has been reported previously. The key secondary end point was overall survival in the intention-to-treat population. Safety was also assessed. RESULTS: A total of 664 patients were assigned to receive A+AVD and 670 to receive ABVD. At a median follow-up of 73.0 months, 39 patients in the A+AVD group and 64 in the ABVD group had died (hazard ratio, 0.59; 95% confidence interval [CI], 0.40 to 0.88; P = 0.009). The 6-year overall survival estimates were 93.9% (95% CI, 91.6 to 95.5) in the A+AVD group and 89.4% (95% CI, 86.6 to 91.7) in the ABVD group. Progression-free survival was longer with A+AVD than with ABVD (hazard ratio for disease progression or death, 0.68; 95% CI, 0.53 to 0.86). Fewer patients in the A+AVD group than in the ABVD group received subsequent therapy, including transplantation, and fewer second cancers were reported with A+AVD (in 23 vs. 32 patients). Primary prophylaxis with granulocyte colony-stimulating factor was recommended after an increased incidence of febrile neutropenia was observed with A+AVD. More patients had peripheral neuropathy with A+AVD than with ABVD, but most patients in the two groups had resolution or amelioration of the event by the last follow-up. CONCLUSIONS: Patients who received A+AVD for the treatment of stage III or IV Hodgkin's lymphoma had a survival advantage over those who received ABVD. (Funded by Takeda Development Center Americas and Seagen; ECHELON-1 ClinicalTrials.gov number, NCT01712490; EudraCT number, 2011-005450-60.).
Asunto(s)
Antineoplásicos Inmunológicos , Protocolos de Quimioterapia Combinada Antineoplásica , Brentuximab Vedotina , Enfermedad de Hodgkin , Antineoplásicos Inmunológicos/efectos adversos , Antineoplásicos Inmunológicos/uso terapéutico , Protocolos de Quimioterapia Combinada Antineoplásica/efectos adversos , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Bleomicina/administración & dosificación , Bleomicina/efectos adversos , Brentuximab Vedotina/administración & dosificación , Brentuximab Vedotina/efectos adversos , Dacarbazina/administración & dosificación , Dacarbazina/efectos adversos , Supervivencia sin Enfermedad , Doxorrubicina/administración & dosificación , Doxorrubicina/efectos adversos , Estudios de Seguimiento , Enfermedad de Hodgkin/tratamiento farmacológico , Enfermedad de Hodgkin/mortalidad , Enfermedad de Hodgkin/patología , Humanos , Estadificación de Neoplasias , Análisis de Supervivencia , Resultado del Tratamiento , Vinblastina/administración & dosificación , Vinblastina/efectos adversosRESUMEN
BACKGROUND: In adults with advanced-stage Hodgkin's lymphoma, the CD30-directed antibody-drug conjugate brentuximab vedotin combined with multiagent chemotherapy has been shown to have greater efficacy, but also more toxic effects, than chemotherapy alone. The efficacy of this targeted therapy approach in children and adolescents with Hodgkin's lymphoma is unclear. METHODS: We conducted an open-label, multicenter, randomized, phase 3 trial involving patients 2 to 21 years of age with previously untreated Hodgkin's lymphoma of stage IIB with bulk tumor or stage IIIB, IVA, or IVB. Patients were assigned to receive five 21-day cycles of brentuximab vedotin with doxorubicin, vincristine, etoposide, prednisone, and cyclophosphamide (brentuximab vedotin group) or the standard pediatric regimen of doxorubicin, bleomycin, vincristine, etoposide, prednisone, and cyclophosphamide (standard-care group). Slow-responding lesions, defined by a score of 4 or 5 (on a 5-point scale, with scores of 1 to 3 indicating rapid-responding lesions), were identified on centrally reviewed positron-emission tomography-computed tomography after two cycles. Involved-site radiation therapy was administered after the fifth cycle of therapy to slow-responding lesions and to large mediastinal adenopathy that was present at diagnosis. The primary end point was event-free survival, defined as the time until disease progression occurred, relapse occurred, a second malignant neoplasm developed, or the patient died. Safety and overall survival were assessed. RESULTS: Of 600 patients who were enrolled across 153 institutions, 587 were eligible. At a median follow-up of 42.1 months (range, 0.1 to 80.9), the 3-year event-free survival was 92.1% (95% confidence interval [CI], 88.4 to 94.7) in the brentuximab vedotin group, as compared with 82.5% (95% CI, 77.4 to 86.5) in the standard-care group (hazard ratio for event or death, 0.41; 95% CI, 0.25 to 0.67; P<0.001). The percentage of patients who received involved-site radiation therapy did not differ substantially between the brentuximab vedotin group and the standard-care group (53.4% and 56.8%, respectively). Toxic effects were similar in the two groups. Overall survival at 3 years was 99.3% (95% CI, 97.3 to 99.8) in the brentuximab vedotin group and 98.5% (95% CI, 96.0 to 99.4) in the standard-care group. CONCLUSIONS: The addition of brentuximab vedotin to standard chemotherapy resulted in superior efficacy, with a 59% lower risk of an event or death, and no increase in the incidence of toxic effects at 3 years. (Funded by the National Institutes of Health and others; AHOD1331 ClinicalTrials.gov number, NCT02166463.).
Asunto(s)
Antineoplásicos Inmunológicos , Protocolos de Quimioterapia Combinada Antineoplásica , Brentuximab Vedotina , Enfermedad de Hodgkin , Adolescente , Adulto , Niño , Humanos , Protocolos de Quimioterapia Combinada Antineoplásica/efectos adversos , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Brentuximab Vedotina/efectos adversos , Brentuximab Vedotina/uso terapéutico , Ciclofosfamida/administración & dosificación , Ciclofosfamida/efectos adversos , Doxorrubicina/administración & dosificación , Doxorrubicina/efectos adversos , Etopósido/administración & dosificación , Etopósido/efectos adversos , Enfermedad de Hodgkin/tratamiento farmacológico , Recurrencia Local de Neoplasia/tratamiento farmacológico , Prednisona/administración & dosificación , Prednisona/efectos adversos , Resultado del Tratamiento , Vincristina/administración & dosificación , Vincristina/efectos adversos , Antineoplásicos Inmunológicos/efectos adversos , Antineoplásicos Inmunológicos/uso terapéutico , Bleomicina/administración & dosificación , Bleomicina/efectos adversosRESUMEN
We induced systemic sclerosis (SSc)-like disease in both wild-type and Dnase1l3-deficient mice using two distinct approaches involving bleomycin and hypochlorous acid injections. Our observations revealed that the deficiency in DNASE1L3 did not affect tissue fibrosis or inflammation caused by these treatments. Despite the association of single nucleotide polymorphisms in humans with SSc pathogenesis, our study demonstrates that DNASE1L3 is dispensable in two inducible murine models of SSc-like pathogenesis.
Asunto(s)
Bleomicina , Modelos Animales de Enfermedad , Endodesoxirribonucleasas , Ratones Noqueados , Esclerodermia Sistémica , Animales , Esclerodermia Sistémica/genética , Esclerodermia Sistémica/patología , Esclerodermia Sistémica/inmunología , Ratones , Endodesoxirribonucleasas/deficiencia , Endodesoxirribonucleasas/genética , Humanos , Ácido Hipocloroso , Fibrosis , Ratones Endogámicos C57BLRESUMEN
Idiopathic pulmonary fibrosis, a fatal interstitial lung disease, is characterized by fibroblast activation and aberrant extracellular matrix accumulation. Effective therapeutic development is limited because of incomplete understanding of the mechanisms by which fibroblasts become aberrantly activated. Here, we show aldehyde dehydrogenase 2 (ALDH2) in fibroblasts as a potential therapeutic target for pulmonary fibrosis. A decrease in ALDH2 expression was observed in patients with idiopathic pulmonary fibrosis and bleomycin-treated mice. ALDH2 deficiency spontaneously induces collagen accumulation in the lungs of aged mice. Furthermore, young ALDH2 knockout mice exhibited exacerbated bleomycin-induced pulmonary fibrosis and increased mortality compared with that in control mice. Mechanistic studies revealed that transforming growth factor (TGF)-ß1 induction and ALDH2 depletion constituted a positive feedback loop that exacerbates fibroblast activation. TGF-ß1 down-regulated ALDH2 through a TGF-ß receptor 1/Smad3-dependent mechanism. The subsequent deficiency in ALDH2 resulted in fibroblast dysfunction that manifested as impaired mitochondrial autophagy and senescence, leading to fibroblast activation and extracellular matrix production. ALDH2 overexpression markedly suppressed fibroblast activation, and this effect was abrogated by PTEN-induced putative kinase 1 (PINK1) knockdown, indicating that the profibrotic effects of ALDH2 are PINK1- dependent. Furthermore, ALDH2 activated by N-(1,3-benzodioxol-5-ylmethyl)-2,6-dichlorobenzamide (Alda-1) reversed the established pulmonary fibrosis in both young and aged mice. In conclusion, ALDH2 expression inhibited the pathogenesis of pulmonary fibrosis. Strategies to up-regulate or activate ALDH2 expression could be potential therapies for pulmonary fibrosis.
Asunto(s)
Aldehído Deshidrogenasa Mitocondrial , Bleomicina , Senescencia Celular , Fibroblastos , Ratones Noqueados , Mitocondrias , Animales , Fibroblastos/metabolismo , Fibroblastos/patología , Aldehído Deshidrogenasa Mitocondrial/metabolismo , Aldehído Deshidrogenasa Mitocondrial/genética , Aldehído Deshidrogenasa Mitocondrial/deficiencia , Ratones , Mitocondrias/metabolismo , Mitocondrias/patología , Humanos , Bleomicina/toxicidad , Bleomicina/efectos adversos , Factor de Crecimiento Transformador beta1/metabolismo , Ratones Endogámicos C57BL , Masculino , Fibrosis Pulmonar Idiopática/patología , Fibrosis Pulmonar Idiopática/metabolismo , Fibrosis Pulmonar Idiopática/genética , Fibrosis Pulmonar/patología , Fibrosis Pulmonar/metabolismo , Fibrosis Pulmonar/genética , Transducción de Señal , Pulmón/patología , Pulmón/metabolismoRESUMEN
The high mortality rates of acute lung injury and acute respiratory distress syndrome challenge the field to identify biomarkers and factors that can be exploited for therapeutic approaches. IL-22 is a cytokine that has antibacterial and reparative properties in the lung. However, it also can exacerbate inflammation and requires tight control by the extracellular inhibitory protein known as IL-22 binding protein (IL-22BP) (Il22ra2). This study showed the necessity of IL-22BP in controlling and preventing acute lung injury using IL-22BP knockout mice (Il22ra2-/-) in the bleomycin model of acute lung injury/acute respiratory distress syndrome. Il22ra2-/- mice had greater sensitivity (weight loss and death) and pulmonary inflammation in the acute phase (first 7 days) of the injury compared with wild-type C57Bl/6 controls. The inflammation was driven by excess IL-22 production, inducing the influx of pathogenic IL-17A+ γδ T cells to the lung. Interestingly, this inflammation was initiated in part by the noncanonical IL-22 signaling to macrophages, which express the IL-22 receptor (Il22ra1) in vivo after bleomycin challenge. This study further showed that IL-22 receptor alpha-1+ macrophages can be stimulated by IL-22 to produce a number of IL-17-inducing cytokines such as IL-1ß, IL-6, and transforming growth factor-ß1. Together, the results suggest that IL-22BP prevents IL-22 signaling to macrophages and reduces bleomycin-mediated lung injury.
Asunto(s)
Lesión Pulmonar Aguda , Lesión Pulmonar , Síndrome de Dificultad Respiratoria , Animales , Ratones , Lesión Pulmonar Aguda/patología , Bleomicina/efectos adversos , Citocinas/metabolismo , Inflamación/patología , Interleucina-22 , Pulmón/patología , Lesión Pulmonar/patología , Ratones Endogámicos C57BL , Ratones Noqueados , Síndrome de Dificultad Respiratoria/metabolismoRESUMEN
Idiopathic pulmonary fibrosis is a progressive interstitial lung disease for which there is no curative therapy available. Repetitive alveolar epithelial injury repair, myofibroblast accumulation, and excessive collagen deposition are key pathologic features of idiopathic pulmonary fibrosis, eventually leading to cellular hypoxia and respiratory failure. The precise mechanism driving this complex maladaptive process remains inadequately understood. WD repeat and suppressor of cytokine signaling box containing 1 (WSB1) is an E3 ubiquitin ligase, the expression of which is associated strongly with hypoxia, and forms a positive feedback loop with hypoxia-inducible factor 1α (HIF-1α) under anoxic condition. This study explored the expression, cellular distribution, and function of WSB1 in bleomycin (BLM)-induced mouse lung injury and fibrosis. WSB1 expression was highly induced by BLM injury and correlated with the progression of lung fibrosis. Significantly, conditional deletion of Wsb1 in adult mice ameliorated BLM-induced pulmonary fibrosis. Phenotypically, Wsb1-deficient mice showed reduced lipofibroblast to myofibroblast transition, but enhanced alveolar type 2 proliferation and differentiation into alveolar type 1 after BLM injury. Proteomic analysis of mouse lung tissues identified caveolin 2 as a potential downstream target of WSB1, contributing to BLM-induced epithelial injury repair and fibrosis. These findings unravel a vital role for WSB1 induction in lung injury repair, thus highlighting it as a potential therapeutic target for pulmonary fibrosis.
Asunto(s)
Fibrosis Pulmonar Idiopática , Lesión Pulmonar , Animales , Ratones , Ubiquitina-Proteína Ligasas/genética , Ubiquitina-Proteína Ligasas/metabolismo , Miofibroblastos/metabolismo , Lesión Pulmonar/patología , Proteómica , Pulmón/patología , Fibrosis , Hipoxia/patología , Fibrosis Pulmonar Idiopática/patología , Bleomicina/toxicidad , Regeneración , Péptidos y Proteínas de Señalización IntracelularRESUMEN
Idiopathic pulmonary fibrosis (IPF) is a progressive interstitial lung disease characterized by pulmonary fibroblast overactivation, resulting in the accumulation of abnormal extracellular matrix and lung parenchymal damage. Although the pathogenesis of IPF remains unclear, aging was proposed as the most prominent nongenetic risk factor. Propionate metabolism undergoes reprogramming in the aging population, leading to the accumulation of the by-product methylmalonic acid (MMA). This study aimed to explore alterations in propionate metabolism in IPF and the impact of the by-product MMA on pulmonary fibrosis. It revealed alterations in the expression of enzymes involved in propionate metabolism within IPF lung tissues, characterized by an increase in propionyl-CoA carboxylase and methylmalonyl-CoA epimerase expression, and a decrease in methylmalonyl-CoA mutase expression. Knockdown of methylmalonyl-CoA mutase, the key enzyme in propionate metabolism, induced a profibrotic phenotype and activated co-cultured fibroblasts in A549 cells. MMA exacerbated bleomycin-induced mouse lung fibrosis and induced a profibrotic phenotype in both epithelial cells and fibroblasts through activation of the canonical transforming growth factor-ß/Smad pathway. Overall, these findings unveil an alteration of propionate metabolism in IPF, leading to MMA accumulation, thus exacerbating lung fibrosis through promoting profibrotic phenotypic transitions via the canonical transforming growth factor-ß/Smad signaling pathway.
Asunto(s)
Envejecimiento , Fibrosis Pulmonar Idiopática , Ácido Metilmalónico , Animales , Humanos , Ratones , Ácido Metilmalónico/metabolismo , Envejecimiento/metabolismo , Envejecimiento/patología , Fibrosis Pulmonar Idiopática/metabolismo , Fibrosis Pulmonar Idiopática/patología , Fibrosis Pulmonar Idiopática/inducido químicamente , Masculino , Fibroblastos/metabolismo , Fibroblastos/patología , Femenino , Ratones Endogámicos C57BL , Anciano , Fibrosis Pulmonar/metabolismo , Fibrosis Pulmonar/patología , Fibrosis Pulmonar/inducido químicamente , Persona de Mediana Edad , Células A549 , Bleomicina/efectos adversos , Pulmón/patología , Pulmón/metabolismoRESUMEN
The optimal first-line treatment for nodular lymphocyte-predominant Hodgkin lymphoma (NLPHL) diagnosed in early stages is largely undefined. We, therefore, analyzed 100 NLPHL patients treated in the randomized HD16 (early-stage favorable; n = 85) and HD17 (early-stage unfavorable; n = 15) studies. These studies investigated the omission of consolidation radiotherapy (RT) in patients with a negative interim positron emission tomography (iPET) (ie, Deauville score <3) after chemotherapy (HD16: 2× doxorubicin, bleomycin, vinblastine, and dacarbazine [ABVD]; HD17: 2× escalated bleomycin, etoposide, doxorubicin, cyclophosphamide, vincristine, procarbazine, and prednisone [BEACOPP] plus 2× ABVD). Patients with NLPHL treated in the HD16 and HD17 studies had 5-year progression-free survival (PFS) rates of 90.3% and 92.9%, respectively. Thus, the 5-year PFS did not differ significantly from that of patients with classical Hodgkin lymphoma treated within the same studies (HD16: P = .88; HD17: P = .50). Patients with early-stage favorable NLPHL who had a negative iPET after 2× ABVD and did not undergo consolidation RT tended to have a worse 5-year PFS than patients with a negative iPET who received consolidation RT (83% vs 100%; P = .05). There were 10 cases of NLPHL recurrence. However, no NLPHL patient died during follow-up. Hence, the 5-year overall survival rate was 100%. Taken together, contemporary Hodgkin lymphoma-directed treatment approaches result in excellent outcomes for patients with newly diagnosed early-stage NLPHL and, thus, represent valid treatment options. In early-stage favorable NLPHL, consolidation RT appears necessary after 2× ABVD to achieve the optimal disease control irrespective of the iPET result.
Asunto(s)
Enfermedad de Hodgkin , Humanos , Enfermedad de Hodgkin/diagnóstico por imagen , Enfermedad de Hodgkin/tratamiento farmacológico , Bleomicina/efectos adversos , Doxorrubicina , Dacarbazina , Vinblastina , Protocolos de Quimioterapia Combinada Antineoplásica/efectos adversos , Ciclofosfamida , Vincristina/efectos adversos , Tomografía de Emisión de Positrones/métodos , PrednisonaRESUMEN
Idiopathic pulmonary fibrosis (IPF) is a chronic progressive disease with an abnormal accumulation of fibrotic tissue in the lung parenchyma and elevated glycolysis level in associated cells without effective therapy options. Lactate accumulation in pulmonary fibrotic tissue is a significant factor aggravating IPF development, but the main mechanism regulating glycolysis needs further investigation. In this study, lung fibrosis model was induced by bleomycin (BLM) intratracheally in female C57BL/6 mice. The changes of lactate level and fibrotic markers were detected. For in vitro studies, cell lines of alveolar epithelial cell and lung fibroblast cell were stimulated with TGF-ß1 and BLM respectively, to detect changes in their fibrotic properties. The function of lactate accumulation on facilitating fibrosis was verified. We demonstrated that BLM-induced pulmonary fibrosis is accompanied by lactate accumulation owing to glycolysis upregulation. Significantly high PDK1 expression in lung fibrotic tissue promotes glycolysis. Moreover, PDK1 stimulated trans-differentiation of lung fibroblasts and epithelial-mesenchymal transition (EMT) of alveolar epithelial cells. Furthermore, phosphorylated Akt2 activated PDK1 to cause pulmonary fibrosis and inhibitors of Akt2 and PDK1 could suppress fibrotic process. This study is the first to consider PDK1 facilitated lactate accumulation through glycolysis as a vital factor in pulmonary fibrosis and could be initiated by Akt2. We concluded that the pro-fibrotic properties of PDK1 are associated with Akt2 phosphorylation and thus provide new potential therapeutic targets for pulmonary fibrosis.
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Fibrosis Pulmonar Idiopática , Ácido Láctico , Femenino , Ratones , Animales , Ratones Endogámicos C57BL , Transducción de Señal , Fibrosis Pulmonar Idiopática/inducido químicamente , Células Epiteliales Alveolares , Bleomicina/toxicidad , Proteínas Proto-Oncogénicas c-aktRESUMEN
Pulmonary fibrosis is a formidable challenge in chronic and age-related lung diseases. Myofibroblasts secrete large amounts of extracellular matrix and induce pro-repair responses during normal wound healing. Successful tissue repair results in termination of myofibroblast activity via apoptosis; however, some myofibroblasts exhibit a senescent phenotype and escape apoptosis, causing over-repair that is characterized by pathological fibrotic scarring. Therefore, the removal of senescent myofibroblasts using senolytics is an important method for the treatment of pulmonary fibrosis. Procyanidin C1 (PCC1) has recently been discovered as a senolytic compound with very low toxicity and few side effects. This study aimed to determine whether PCC1 could improve lung fibrosis by promoting apoptosis in senescent myofibroblasts and to investigate the mechanisms involved. The results showed that PCC1 attenuates bleomycin (BLM)-induced pulmonary fibrosis in mice. In addition, we found that PCC1 inhibited extracellular matrix deposition and promoted the apoptosis of senescent myofibroblasts by increasing PUMA expression and activating the BAX signaling pathway. Our findings represent a new method of pulmonary fibrosis management and emphasize the potential of PCC1 as a senotherapeutic agent for the treatment of pulmonary fibrosis, providing hope for patients with pulmonary fibrosis worldwide. Our results advance our understanding of age-related diseases and highlight the importance of addressing cellular senescence in treatment.
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Bleomicina , Catequina , Senescencia Celular , Ratones Endogámicos C57BL , Miofibroblastos , Fibrosis Pulmonar , Animales , Bleomicina/toxicidad , Miofibroblastos/metabolismo , Miofibroblastos/efectos de los fármacos , Fibrosis Pulmonar/inducido químicamente , Fibrosis Pulmonar/metabolismo , Fibrosis Pulmonar/tratamiento farmacológico , Fibrosis Pulmonar/patología , Ratones , Senescencia Celular/efectos de los fármacos , Catequina/farmacología , Catequina/análogos & derivados , Proantocianidinas/farmacología , Apoptosis/efectos de los fármacos , Masculino , Biflavonoides/farmacología , Transducción de Señal/efectos de los fármacosRESUMEN
Idiopathic pulmonary fibrosis (IPF) is a progressive scarring disease of the lung that leads rapidly to respiratory failure. Novel approaches to treatment are urgently needed. The bioactive lipid sphingosine-1-phosphate (S1P) is increased in IPF lungs and promotes proinflammatory and profibrotic TGF-ß signaling. Hence, decreasing lung S1P represents a potential therapeutic strategy for IPF. S1P is degraded by the intracellular enzyme S1P lyase (SPL). Here we find that a knock-in mouse with a missense SPL mutation mimicking human disease resulted in reduced SPL activity, increased S1P, increased TGF-ß signaling, increased lung fibrosis, and higher mortality after injury compared to wild type (WT). We then tested adeno-associated virus 9 (AAV9)-mediated overexpression of human SGPL1 (AAV-SPL) in mice as a therapeutic modality. Intravenous treatment with AAV-SPL augmented lung SPL activity, attenuated S1P levels within the lungs, and decreased injury-induced fibrosis compared to controls treated with saline or only AAV. We confirmed that AAV-SPL treatment led to higher expression of SPL in the epithelial and fibroblast compartments during bleomycin-induced lung injury. Additionally, AAV-SPL decreased expression of the profibrotic cytokines TNFα and IL1ß as well as markers of fibroblast activation, such as fibronectin (Fn1), Tgfb1, Acta2, and collagen genes in the lung. Taken together, our results provide proof of concept for the use of AAV-SPL as a therapeutic strategy for the treatment of IPF. © 2024 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.
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Dependovirus , Fibrosis Pulmonar Idiopática , Lisofosfolípidos , Esfingosina/análogos & derivados , Humanos , Ratones , Animales , Dependovirus/genética , Pulmón/metabolismo , Fibrosis Pulmonar Idiopática/genética , Fibrosis Pulmonar Idiopática/terapia , Fibrosis Pulmonar Idiopática/metabolismo , Bleomicina , Modelos Animales , Terapia Genética , Aldehído-Liasas/genética , Aldehído-Liasas/metabolismoRESUMEN
Epidemiological evidence indicates that exposure to particulate matter is linked to the development of idiopathic pulmonary fibrosis (IPF) and increases the incidence of acute exacerbations of IPF. In addition to accelerating the rate of lung function decline, exposure to fine particulate matter (particulate matter smaller than 2.5 µm [PM2.5]) is a risk factor for increased mortality in subjects with IPF. In this article, we show that exposure to PM2.5 mediates monocyte recruitment and fibrotic progression in mice with established fibrosis. In mice with established fibrosis, bronchoalveolar lavage cells showed monocyte/macrophage heterogeneity after exposure to PM2.5. These cells had a significant inflammatory and anti-inflammatory signature. The mixed heterogeneity of cells contributed to the proinflammatory and anti-inflammatory response. Although monocyte-derived macrophages were recruited to the lung in bleomycin-injured mice treated with PM2.5, recruitment of monocytes expressing Ly6Chi to the lung promoted progression of fibrosis, reduced lung aeration on computed tomography, and impacted lung compliance. Ly6Chi monocytes isolated from PM2.5-exposed fibrotic mice showed enhanced expression of proinflammatory markers compared with fibrotic mice exposed to vehicle. Moreover, IPF bronchoalveolar lavage cells treated ex vivo with PM2.5 showed an exaggerated inflammatory response. Targeting Ly6Chi monocyte recruitment inhibited fibrotic progression in mice. Moreover, the adoptive transfer of Ly6Chi monocytes exacerbated established fibrosis. These observations suggest that enhanced recruitment of Ly6Chi monocytes with a proinflammatory phenotype mediates acute exacerbations of pulmonary fibrosis, and targeting these cells may provide a potential novel therapeutic target to protect against acute exacerbations of IPF.
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Fibrosis Pulmonar Idiopática , Pulmón , Humanos , Ratones , Animales , Pulmón/patología , Fibrosis Pulmonar Idiopática/patología , Fibrosis , Bleomicina/uso terapéutico , Material Particulado/efectos adversos , Antiinflamatorios/uso terapéuticoRESUMEN
Idiopathic pulmonary fibrosis (IPF) is a fibrotic age-related chronic lung disease characterized by the accumulation of senescent cells. Whether impaired immune response is responsible for the accumulation of senescent cells in the IPF lung remains unknown. In this study, we characterized the NK phenotype in IPF lungs via flow cytometry using 5-dodecanoylaminofluorescein di-ß-d-galactopyranoside, markers of tissue residence, and chemokine receptors. The effect of the lung microenvironment was evaluated using lung fibroblast (LF) conditioned media (CM), and the bleomycin-induced pulmonary fibrosis mouse model was used to assess the in vivo relationship between NK cells and the accumulation of senescent cells. We found that NK cells from the lower lobe of IPF patients exhibited immune-senescent and impaired CD57-NKG2A+ phenotype. We also observed that culture of NK cells from healthy donors in CM from IPF lower lobe lung fibroblasts induced a senescent-like phenotype and impaired cytotoxic capacity. There is an impaired NK recruitment by LF, and NKs presented decreased migration toward their CM. In addition, NK cell-depleted mice treated with bleomycin showed increased collagen deposition and accumulation of different populations of senescent cells compared with controls. The IPF lung microenvironment induces a dysfunctional NK phenotype limiting the clearance of lung senescent cells and the resolution of lung fibrosis. We propose that impaired NK activity could be one of the mechanisms responsible for perpetuating the accumulation of senescent cells in IPF lungs.
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Antineoplásicos , Fibrosis Pulmonar Idiopática , Ratones , Animales , Pulmón/patología , Fibrosis Pulmonar Idiopática/inducido químicamente , Bleomicina/efectos adversos , Fibrosis , Antineoplásicos/farmacología , FibroblastosRESUMEN
The involvement of γδT cells, Th17 cells, and CD4+CD25+ regulatory T cells (Tregs) is crucial in the progression of pulmonary fibrosis (PF), particularly in maintaining immune tolerance and homeostasis. However, the dynamics of these cells in relation to PF progression, especially under pharmacological interventions, remains poorly understood. This study aims to unravel the interplay between the dynamic changes of these cells and the effect of pharmacological agents in a mouse model of PF induced by intratracheal instillation of bleomycin. We analyzed changes in lung histology, lung index, hydroxyproline levels, and the proportions of γδT cells, Th17 cells, and Tregs on the 3rd, 14th, and 28th days following treatment with Neferine, Isoliensinine, Pirfenidone, and Prednisolone. Our results demonstrate that these drugs can partially or dynamically reverse weight loss, decrease lung index and hydroxyproline levels, and ameliorate lung histopathological damage. Additionally, they significantly modulated the abnormal changes in γδT, Th17, and Treg cell proportions. Notably, on day 3, the proportion of γδT cells increased in the Neferine and Prednisolone groups but decreased in the Isoliensinine and Pirfenidone groups, while the proportion of Th17 cells decreased across all treated groups. On day 14, the Neferine group showed an increase in all three cell types, whereas the Pirfenidone group exhibited a decrease. In the Isoliensinine group, γδT and Th17 cells increased, and in the Prednisolone group, only Tregs increased. By day 28, an increase in Th17 cell proportion was observed in all treatment groups, with a decrease in γδT cells noted in the Neferine group. These shifts in cell proportions are consistent with the pathogenesis changes induced by these anti-PF drugs, suggesting a correlation between cellular dynamics and pharmacological interventions in PF progression. Our findings imply potential strategies for assessing the efficacy and timing of anti-PF treatments based on these cellular changes.