RESUMEN
Drosophila matrix metalloproteinase 2 (MMP2) is specifically expressed in posterior follicle cells of stage-14 egg chambers (mature follicles) and is crucial for the breakdown of the follicular wall during ovulation, a process that is highly conserved from flies to mammals. The factors that regulate spatiotemporal expression of MMP2 in follicle cells remain unknown. Here, we demonstrate crucial roles for the ETS-family transcriptional activator Pointed (Pnt) and its endogenous repressor Yan in the regulation of MMP2 expression. We found that Pnt is expressed in posterior follicle cells and overlaps with MMP2 expression in mature follicles. Genetic analysis demonstrated that pnt is both required and sufficient for MMP2 expression in follicle cells. In addition, Yan was temporally upregulated in stage-13 follicle cells to fine-tune Pnt activity and MMP2 expression. Furthermore, we identified a 1.1â kb core enhancer that is responsible for the spatiotemporal expression of MMP2 and contains multiple pnt/yan binding motifs. Mutation of pnt/yan binding sites significantly impaired the Mmp2 enhancer activity. Our data reveal a mechanism of transcriptional regulation of Mmp2 expression in Drosophila ovulation, which could be conserved in other biological systems.
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Proteínas de Drosophila , Drosophila , Animales , Femenino , Drosophila/metabolismo , Metaloproteinasa 2 de la Matriz/genética , Metaloproteinasa 2 de la Matriz/metabolismo , Proteínas Proto-Oncogénicas c-ets/genética , Proteínas Proto-Oncogénicas c-ets/metabolismo , Proteínas de Unión al ADN/genética , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Transducción de Señal/fisiología , Ovulación/genética , Mamíferos/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Factores de Transcripción/genéticaRESUMEN
Across species, ovulation is a process induced by a myriad of signaling cascades that ultimately leads to the release of encapsulated oocytes from follicles. Follicles first need to mature and gain ovulatory competency before ovulation; however, the signaling pathways regulating follicle maturation are incompletely understood in Drosophila and other species. Our previous work has shown that the bHLH-PAS transcription factor Single-minded (Sim) plays important roles in follicle maturation downstream of the nuclear receptor Ftz-f1 in Drosophila. Here, we demonstrate that Tango (Tgo), another bHLH-PAS protein, acts as a co-factor of Sim to promote follicle cell differentiation from stages 10 to 12. In addition, we discover that re-upregulation of Sim in stage-14 follicle cells is also essential to promote ovulatory competency by upregulating octopamine receptor in mushroom body (OAMB), matrix metalloproteinase 2 (Mmp2) and NADPH oxidase (NOX), either independently of or in conjunction with the zinc-finger protein Hindsight (Hnt). All these factors are crucial for successful ovulation. Together, our work indicates that the transcriptional complex Sim:Tgo plays multiple roles in late-stage follicle cells to promote follicle maturation and ovulation.
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Proteínas de Drosophila , Metaloproteinasa 2 de la Matriz , Animales , Femenino , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Drosophila/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Metaloproteinasa 2 de la Matriz/metabolismo , Oogénesis/genética , Ovulación/genéticaRESUMEN
Dendrites possess distinct structural and functional properties that enable neurons to receive information from the environment as well as other neurons. Despite their key role in neuronal function, current understanding of the ability of neurons to regenerate dendrites is lacking. This study characterizes the structural and functional capacity for dendrite regeneration in vivo in adult animals and examines the effect of neuronal maturation on dendrite regeneration. We focused on the class IV dendritic arborization (c4da) neuron of the Drosophila sensory system, which has a dendritic arbor that undergoes dramatic remodeling during the first 3 d of adult life and then maintains a relatively stable morphology thereafter. Using a laser severing paradigm, we monitored regeneration after acute and spatially restricted injury. We found that the capacity for regeneration was present in adult neurons but diminished as the animal aged. Regenerated dendrites recovered receptive function. Furthermore, we found that the regenerated dendrites show preferential alignment with the extracellular matrix (ECM). Finally, inhibition of ECM degradation by inhibition of matrix metalloproteinase 2 (Mmp2) to preserve the extracellular environment characteristics of young adults led to increased dendrite regeneration. These results demonstrate that dendrites retain regenerative potential throughout adulthood and that regenerative capacity decreases with aging.
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Dendritas/fisiología , Drosophila/fisiología , Metaloproteinasa 2 de la Matriz/metabolismo , Regeneración , Células Receptoras Sensoriales/fisiología , Envejecimiento/fisiología , Animales , Dendritas/enzimología , Drosophila/citología , Drosophila/enzimología , Proteínas de Drosophila/metabolismo , Epidermis/enzimología , Matriz Extracelular/fisiología , Regulación del Desarrollo de la Expresión Génica , Integrinas/genética , Integrinas/metabolismo , Células Receptoras Sensoriales/enzimologíaRESUMEN
The basement membrane zone is the interface between the epidermis and dermis, and it is disrupted in several skin conditions. Here, we report the results of a comprehensive investigation into the structural and molecular factors of the basement membrane zone in vitiligo, a dermatological disorder characterised by depigmented patches on the skin. Using electron microscopy and immunofluorescence staining, we confirmed abnormal basement membrane zone morphology and disrupted basement membrane zone architecture in human vitiliginous skin. Furthermore, we identified elevated expression of matrix metalloproteinase 2 (MMP2) in human dermal fibroblasts as a key factor responsible for basement membrane zone matrix degradation. In our in vitro and ex vivo models, overexpression of MMP2 in fibroblasts led to basement membrane zone disruption and melanocyte disappearance. Importantly, we reveal that the loss of melanocytes in vitiligo is primarily linked to their weakened adhesion to the basement membrane, mediated by binding between integrin ß1 and laminin and discoidin domain receptor 1 and collagen IV. Finally, inhibition of matrix metalloproteinase 2 expression reversed depigmentation in a mouse model of vitiligo. In conclusion, our research shows the importance of basement membrane zone integrity in melanocyte residence and offers new avenues for therapeutic interventions to address this challenging skin condition. © 2024 The Author(s). 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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Membrana Basal , Melanocitos , Vitíligo , Vitíligo/patología , Vitíligo/metabolismo , Melanocitos/patología , Melanocitos/metabolismo , Membrana Basal/patología , Membrana Basal/metabolismo , Humanos , Animales , Ratones , Metaloproteinasa 2 de la Matriz/metabolismo , Fibroblastos/patología , Fibroblastos/metabolismo , Masculino , Femenino , Modelos Animales de Enfermedad , Ratones Endogámicos C57BLRESUMEN
OBJECTIVE: A wide range of cardiac diseases is associated with inflammation. "Inflamed" heart tissue is infiltrated with pro-inflammatory macrophages which extensively secrete matrix metalloproteinase 9 (MMP9), a regulator of extracellular matrix turnover. As MMP9 is released from macrophages in a latent form, it requires activation. The present study addresses the role of cardiomyocytes in the course of this activation process. METHODS AND RESULTS: In mono- and co-cultures of pro-inflammatory rat macrophages (bone marrow-derived and peritoneal) and cardiomyocytes (H9C2 cell line) gelatin zymography demonstrated that activated macrophages robustly secreted latent pro-MMP9, whereas cardiomyocytes could not produce the enzyme. Co-culturing of the two cell species was critical for pro-MMP9 activation and was also accompanied by processing of cardiomyocyte-secreted pro-MMP2. A cascade of pro-MMP9 activation was initiated on macrophage membrane with pro-MMP2 cleavage. Namely, pro-inflammatory macrophages expressed an active membrane type 1 MMP (MT1MMP), which activated pro-MMP2, which in turn converted pro-MMP9. Downregulation of MT1MMP in macrophages by siRNA abolished activation of both pro-MMP2 and pro-MMP9 in co-culture. In addition, both cell species secreted MMP13 as a further pro-MMP9 activator. In co-culture, activation of pro-MMP13 occurred on membranes of macrophages and was enhanced in presence of active MMP2. Using incubations with recombinant MMPs and isolated macrophage membranes, we demonstrated that while both MMP2 and MMP13 individually had the ability to activate pro-MMP9, their combined action provided a synergistic effect. CONCLUSION: Activation of pro-MMP9 in a co-culture of pro-inflammatory macrophages and cardiomyocytes was the result of a complex interaction of several MMPs on the cell membrane and in the extracellular space. Both cell types contributed critically to pro-MMP9 processing.
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Metaloproteinasa 2 de la Matriz , Metaloproteinasa 9 de la Matriz , Animales , Ratas , Células Cultivadas , Técnicas de Cocultivo , Macrófagos/metabolismo , Metaloproteinasa 13 de la Matriz , Metaloproteinasa 2 de la Matriz/genética , Metaloproteinasa 2 de la Matriz/metabolismo , Metaloproteinasa 9 de la Matriz/genética , Metaloproteinasa 9 de la Matriz/metabolismo , Miocitos Cardíacos/metabolismoRESUMEN
Epithelial-mesenchymal transition (EMT) is an important biological process contributing to kidney fibrosis and chronic kidney disease. This process is characterized by decreased epithelial phenotypes/markers and increased mesenchymal phenotypes/markers. Tubular epithelial cells (TECs) are commonly susceptible to EMT by various stimuli, for example, transforming growth factor-ß (TGF-ß), cellular communication network factor 2, angiotensin-II, fibroblast growth factor-2, oncostatin M, matrix metalloproteinase-2, tissue plasminogen activator (t-PA), plasmin, interleukin-1ß, and reactive oxygen species. Similarly, glomerular podocytes can undergo EMT via these stimuli and by high glucose condition in diabetic kidney disease. EMT of TECs and podocytes leads to tubulointerstitial fibrosis and glomerulosclerosis, respectively. Signaling pathways involved in EMT-mediated kidney fibrosis are diverse and complex. TGF-ß1/Smad and Wnt/ß-catenin pathways are the major venues triggering EMT in TECs and podocytes. These two pathways thus serve as the major therapeutic targets against EMT-mediated kidney fibrosis. To date, a number of EMT inhibitors have been identified and characterized. As expected, the majority of these EMT inhibitors affect TGF-ß1/Smad and Wnt/ß-catenin pathways. In addition to kidney fibrosis, these EMT-targeted antifibrotic inhibitors are expected to be effective for treatment against fibrosis in other organs/tissues.
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Factor de Crecimiento Transformador beta1 , beta Catenina , Humanos , Factor de Crecimiento Transformador beta1/metabolismo , Factor de Crecimiento Transformador beta1/farmacología , beta Catenina/metabolismo , Metaloproteinasa 2 de la Matriz/metabolismo , Metaloproteinasa 2 de la Matriz/farmacología , Activador de Tejido Plasminógeno/metabolismo , Activador de Tejido Plasminógeno/farmacología , Células Epiteliales/metabolismo , Vía de Señalización Wnt , Transición Epitelial-Mesenquimal , Riñón , FibrosisRESUMEN
Exosomes derived from bone marrow-derived mesenchymal stem cells (BMSCs) can alleviate the symptoms of pelvic floor dysfunction (PFD) in rats. However, the potential therapeutical effects of exosomes derived from BMSCs treated with tumour necrosis factor (TNF)-α on the symptoms of PFD in rats are unknown. Exosomes extracted from BMSCs treated with or without TNF-α were applied to treat PFD rats. Our findings revealed a significant elevation in interleukin (IL)-6 and TNF-α, and matrix metalloproteinase-2 (MMP2) levels in the vaginal wall tissues of patients with pelvic organ prolapse (POP) compared with the control group. Daily administration of exosomes derived from BMSCs, treated either with or without TNF-α (referred to as Exo and TNF-Exo), resulted in increased void volume and bladder void pressure, along with reduced peak bladder pressure and leak point pressure in PFD rats. Notably, TNF-Exo treatment demonstrated superior efficacy in restoring void volume, bladder void pressure and the mentioned parameters compared with Exo treatment. Importantly, TNF-Exo exhibited greater potency than Exo in restoring the levels of multiple proteins (Elastin, Collagen I, Collagen III, IL-6, TNF-α and MMP2) in the anterior vaginal walls of PFD rats. The application of exosomes derived from TNF-α-treated BMSCs holds promise as a novel therapeutic approach for treating PFD.
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Exosomas , Metaloproteinasa 2 de la Matriz , Células Madre Mesenquimatosas , Prolapso de Órgano Pélvico , Factor de Necrosis Tumoral alfa , Animales , Exosomas/metabolismo , Exosomas/trasplante , Células Madre Mesenquimatosas/metabolismo , Femenino , Factor de Necrosis Tumoral alfa/metabolismo , Ratas , Humanos , Prolapso de Órgano Pélvico/terapia , Prolapso de Órgano Pélvico/metabolismo , Metaloproteinasa 2 de la Matriz/metabolismo , Ratas Sprague-Dawley , Interleucina-6/metabolismo , Diafragma Pélvico , Modelos Animales de Enfermedad , Células de la Médula Ósea/metabolismo , Vagina/patología , Trasplante de Células Madre Mesenquimatosas/métodos , Trastornos del Suelo Pélvico/terapia , Persona de Mediana EdadRESUMEN
Nasopharyngeal carcinoma (NPC) is prevalent in Asia and exhibits highly metastatic characteristics, leading to uncontrolled disease progression. Isoliquiritigenin (ISL) have attracted attention due to their diverse biological and pharmacological properties, including anticancer activities. However, the impact of ISL on the invasive and migratory ability of NPC remains poorly understood. Hence, this study aimed to investigate the in vitro anti-metastatic effects of ISL on NPC cells and elucidate the underlying signalling pathways. Human NPC cell NPC-39 and NPC-BM were utilized as cell models. Migratory and invasive capabilities were evaluated through wound healing and invasion assays, respectively. Gelatin zymography was employed to demonstrate matrix metalloproteinase-2 (MMP-2) activity, while western blotting was conducted to analyse protein expression levels and explore signalling cascades. Overexpression of signal transducer and activator of transcription 3 (STAT3) was carried out by transduction of STAT3-expressing vector. Our findings revealed that ISL effectively suppressed the migration and invasion of NPC cells. Gelatin zymography and Western blotting assays demonstrated that ISL treatment led to a reduction in MMP-2 enzyme activity and protein expression. Investigation of signalling cascades revealed that ISL treatment resulted in the inhibition of STAT3 phosphorylation. Moreover, overexpression of STAT3 restored the migratory ability of NPC cells in the presence of ISL. Collectively, these findings indicate that ISL inhibits the migration and invasion of NPC cells associating with MMP-2 downregulation through suppressing STAT3 activation. This suggests that ISL has an anti-metastatic effect on NPC cells and has potential therapeutic benefit for NPC treatment.
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Movimiento Celular , Chalconas , Metaloproteinasa 2 de la Matriz , Carcinoma Nasofaríngeo , Neoplasias Nasofaríngeas , Invasividad Neoplásica , Factor de Transcripción STAT3 , Transducción de Señal , Humanos , Factor de Transcripción STAT3/metabolismo , Metaloproteinasa 2 de la Matriz/metabolismo , Chalconas/farmacología , Carcinoma Nasofaríngeo/metabolismo , Carcinoma Nasofaríngeo/patología , Carcinoma Nasofaríngeo/tratamiento farmacológico , Carcinoma Nasofaríngeo/genética , Transducción de Señal/efectos de los fármacos , Movimiento Celular/efectos de los fármacos , Línea Celular Tumoral , Neoplasias Nasofaríngeas/patología , Neoplasias Nasofaríngeas/metabolismo , Neoplasias Nasofaríngeas/tratamiento farmacológico , Neoplasias Nasofaríngeas/genética , Regulación Neoplásica de la Expresión Génica/efectos de los fármacosRESUMEN
Chlorotoxin (CTX), a scorpion venom-derived 36-residue miniprotein, binds to and is taken up selectively by glioblastoma cells. Previous studies provided controversial results concerning target protein(s) of CTX. These included CLC3 chloride channel, matrix metalloproteinase 2 (MMP-2), regulators of MMP-2, annexin A2, and neuropilin 1 (NRP1). The present study aimed at clarifying which of the proposed binding partners can really interact with CTX using biochemical methods and recombinant proteins. For this purpose, we established two new binding assays based on anchoring the tested proteins to microbeads and quantifying the binding of CTX by flow cytometry. Screening of His-tagged proteins anchored to cobalt-coated beads indicated strong interaction of CTX with MMP-2 and NRP1, whereas binding to annexin A2 was not confirmed. Similar results were obtained with fluorophore-labeled CTX and CTX-displaying phages. Affinity of CTX to MMP-2 and NRP1 was assessed by the "immunoglobulin-coated bead" test, in which the proteins were anchored to beads by specific antibodies. This assay yielded highly reproducible data using both direct titration and displacement approach. The affinities of labeled and unlabeled CTX appeared to be similar for both MMP-2 and NRP1 with estimated KD values of 0.5 to 0.7 µM. Contrary to previous reports, we found that CTX does not inhibit the activity of MMP-2 and that CTX not only with free carboxyl end but also with carboxamide terminal end binds to NRP1. We conclude that the presented robust assays could also be applied for affinity-improving studies of CTX to its genuine targets using phage display libraries.
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Glioblastoma , Metaloproteinasa 2 de la Matriz , Neuropilina-1 , Venenos de Escorpión , Humanos , Glioblastoma/metabolismo , Metaloproteinasa 2 de la Matriz/metabolismo , Neuropilina-1/metabolismo , Venenos de Escorpión/metabolismo , Línea Celular Tumoral , Unión ProteicaRESUMEN
miR-146a, a microRNA (miRNA) that regulates inflammatory responses, plays an important role in many inflammatory diseases. Although an in vitro study had suggested that miR-146a is involved in abnormal inflammatory response, being a critical factor in the pathogenesis of chronic obstructive pulmonary disease (COPD), in vivo evidence of its pathogenic role in COPD remains limited. Eight-week-old male B6(FVB)-Mir146tm1.1Bal/J [miR-146a knockout (KO)] and C57BL/6J mice were intratracheally administered elastase and evaluated after 28 days or exposed to cigarette smoke (CS) and evaluated after 5 mo. miR-146a expression was significantly increased in C57BL/6J mouse lungs due to elastase administration (P = 0.027) or CS exposure (P = 0.019) compared with that in the control group. Compared with C57BL/6J mice, elastase-administered miR-146a-KO mice had lower average computed tomography (CT) values (P = 0.017) and increased lung volume-to-weight ratio (P = 0.016), mean linear intercept (P < 0.001), and destructive index (P < 0.001). Moreover, total cell (P = 0.006), macrophage (P = 0.001), neutrophil (P = 0.026), chemokine (C-X-C motif) ligand 2/macrophage inflammatory protein-2 [P = 0.045; in bronchoalveolar lavage fluid (BALF)], cyclooxygenase-2, and matrix metalloproteinase-2 levels were all increased (in the lungs). Following long-term CS exposure, miR-146a-KO mice showed a greater degree of emphysema formation in their lungs and inflammatory response in the BALF and lungs than C57BL/6J mice. Collectively, miR-146a protected against emphysema formation and the associated abnormal inflammatory response in two murine models.NEW & NOTEWORTHY This study demonstrates that miR-146a expression is upregulated in mouse lungs because of elastase- and CS-induced emphysema and that the inflammatory response by elastase or CS is enhanced in the lungs of miR-146a-KO mice than in those of control mice, resulting in the promotion of emphysema. This is the first study to evaluate the protective role of miR-146a in emphysema formation and the associated abnormal inflammatory response in different in vivo models.
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Enfisema , MicroARNs , Enfermedad Pulmonar Obstructiva Crónica , Enfisema Pulmonar , Animales , Masculino , Ratones , Enfisema/etiología , Inflamación/patología , Pulmón/metabolismo , Metaloproteinasa 2 de la Matriz/metabolismo , Ratones Endogámicos C57BL , Ratones Noqueados , MicroARNs/genética , MicroARNs/metabolismo , Elastasa Pancreática/metabolismo , Enfermedad Pulmonar Obstructiva Crónica/patología , Enfisema Pulmonar/inducido químicamente , Enfisema Pulmonar/genéticaRESUMEN
PURPOSE: Obesity has known detrimental effects on breast cancer (BC) development and progression. However, it's essential to consider the obesity phenotype based on metabolic health. This study aims to evaluate the impact of circulating extracellular vesicles (EVs) from women with metabolically healthy or unhealthy normal weight, overweight, and obesity on MDA-MB-231 cell migration, invasion, and apoptosis. METHODS: Plasma EVs were isolated from different obesity phenotypes in women. EVs were characterized and EVs uptake by MDA-MB-231 cells was assessed. MDA-MB-231 cell lines were treated with EVs obtained from various studied groups, and migration, invasion, MMP-2 and MMP-9 activity, Bax and Bcl-2 mRNA expression, p-53 and Thr55 p-p53 protein expression, and apoptosis were assessed. RESULTS: EVs from obese individuals, regardless of phenotype, increased invasion and MMP-2 activity compared to healthy normal-weight EVs. Normal-weight EVs led to higher invasion under unhealthy conditions. BC cell migration was enhanced by EVs from healthy obese individuals compared to healthy normal-weight EVs. EVs from unhealthy obese women exhibited significantly lower p53/p-p53 levels and reduced apoptosis compared to healthy obese groups. CONCLUSION: It appears that EVs from both normal-weight women with unhealthy conditions and those with obesity or overweight, irrespective of metabolic status, worsened the cancerous behavior of TNBC cells. Therefore, considering metabolic health, in addition to BMI, is crucial for understanding obesity-related disorders.
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Vesículas Extracelulares , Neoplasias de la Mama Triple Negativas , Humanos , Femenino , Sobrepeso/complicaciones , Sobrepeso/metabolismo , Metaloproteinasa 2 de la Matriz/metabolismo , Proteína p53 Supresora de Tumor , Obesidad/metabolismo , Vesículas Extracelulares/metabolismoRESUMEN
BACKGROUND: Thoracic aortic aneurysm (TAA) is a silent but life-threatening cardiovascular disease. Heme oxygenase 1 (HO-1) plays an important role in the cardiovascular diseases but is poorly understood in TAA. This study aims at investigating the role of HO-1 in TAA. METHODS: Single-cell RNA sequencing, Western blot and histological assay were performed to identify specific cellular expression of HO-1 in both human and ß-aminopropionitrile (BAPN)-induced mice TAA. Zinc protoporphyrin (ZnPP), a pharmacological inhibitor of HO-1, was used to investigate whether inhibition of HO-1 could attenuate BAPN-induced TAA in rodent model. Histological assay, Western blot assay, and mRNA sequencing were further performed to explore the underlying mechanisms. RESULTS: Single-cell transcriptomic analyses of 113,800 thoracic aortic cells identified an increase of HO-1(+) macrophage in aneurysmal thoracic aorta from BAPN-induced TAA mice and TAA patients. Histological assay verified HO-1 overexpression in clinical TAA specimens, which was co-localized with CD68(+) macrophage. HO-1(+) macrophage was closely associated with pro-inflammatory response and immune activation. Inhibition of HO-1 through ZnPP significantly alleviated BAPN-induced TAA in mice and restored extracellular matrix (ECM) in vivo. Further experiments showed that ZnPP treatment suppressed the expression of matrix metalloproteinases (MMPs) in aneurysmal thoracic aortic tissues from BAPN-induced TAA mice, including MMP2 and MMP9. Macrophages from myeloid specific HO-1 knockout mice displayed weakened pro-inflammatory activity and ECM degradation capability. CONCLUSION: HO-1(+) macrophage subgroup is a typical hallmark of TAA. Inhibition of HO-1 through ZnPP alleviates BAPN-induced TAA in mice, which might work through restoration of ECM via suppressing MMP2 and MMP9 expression.
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Aneurisma de la Aorta Torácica , Metaloproteinasa 2 de la Matriz , Animales , Humanos , Ratones , Aminopropionitrilo/efectos adversos , Aminopropionitrilo/metabolismo , Aorta Torácica/metabolismo , Aneurisma de la Aorta Torácica/genética , Modelos Animales de Enfermedad , Matriz Extracelular/metabolismo , Hemo-Oxigenasa 1/metabolismo , Metaloproteinasa 2 de la Matriz/metabolismo , Metaloproteinasa 9 de la Matriz/metabolismo , Ratones NoqueadosRESUMEN
Acute myeloid leukemia (AML) is one of the most prevalent types of leukemia and is challenging to cure for most patients. Basic Leucine Zipper ATF-Like Transcription Factor (BATF) has been reported to participate in the development and progression of numerous tumors. However, its role in AML is largely unknown. In this study, the expression and prognostic value of BATF were examined in AML. Our results demonstrated that BATF expression was upregulated in AML patients, which was significantly correlated with poor clinical characteristics and survival. Afterward, functional experiments were performed after knocking down or overexpressing BATF by transfecting small interfering RNAs and overexpression plasmids into AML cells. Our findings revealed that BATF promoted the migratory and invasive abilities of AML cells in vitro and in vivo. Moreover, the target genes of BATF were searched from databases to explore the binding of BATF to the target gene using ChIP and luciferase assays. Notably, our observations validated that BATF is bound to the promoter region of TGF-ß1, which could transcriptionally enhance the expression of TGF-ß1 and activate the TGF-ß1/Smad/MMPs signaling pathway. In summary, our study established the aberrantly high expression of BATF and its pro-migratory function via the TGF-ß1-Smad2/3-MMP2/9 axis in AML, which provides novel insights into extramedullary infiltration of AML.
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Factores de Transcripción con Cremalleras de Leucina de Carácter Básico , Leucemia Mieloide Aguda , Factor de Crecimiento Transformador beta1 , Animales , Femenino , Humanos , Masculino , Ratones , Persona de Mediana Edad , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/genética , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/metabolismo , Línea Celular Tumoral , Movimiento Celular , Regulación Leucémica de la Expresión Génica , Leucemia Mieloide Aguda/metabolismo , Leucemia Mieloide Aguda/patología , Leucemia Mieloide Aguda/genética , Metaloproteinasa 2 de la Matriz/metabolismo , Metaloproteinasa 2 de la Matriz/genética , Metaloproteinasa 9 de la Matriz/metabolismo , Metaloproteinasa 9 de la Matriz/genética , Invasividad Neoplásica , Pronóstico , Transducción de Señal , Proteínas Smad/metabolismo , Proteínas Smad/genética , Factor de Crecimiento Transformador beta1/metabolismo , Factor de Crecimiento Transformador beta1/genéticaRESUMEN
INTRODUCTION: Previous studies have confirmed that low shear stress (LSS) induces glycocalyx disruption, leading to endothelial dysfunction. However, the role of autophagy in LSS-induced glycocalyx disruption and relevant mechanism are not clear. In this study, we hypothesized that LSS may promote autophagy, disrupting the endothelium glycocalyx. METHODS: Human umbilical vein endothelial cells were subjected to physiological shear stress and LSS treatments, followed by the application of autophagy inducers and inhibitors. Additionally, cells were treated with specific matrix metalloproteinase-2 (MMP-2) and matrix metalloproteinase-9 (MMP-9) inhibitor. The expression of autophagic markers, glycocalyx, MMP-2, and MMP-9 was measured. RESULTS: LSS impacted the expression of endothelium autophagy markers, increasing the expression of LC3II.LC3I-1 and Beclin-1, and decreasing the levels of p62, accompanied by glycocalyx disturbance. Moreover, LSS upregulated the expression of MMP-2 and MMP-9 and downregulated the levels of syndecan-1 and heparan sulfate (HS). Additionally, expression of MMP-2 and MMP-9 was increased by an autophagy promoter but was decreased by autophagy inhibitor treatment under LSS. Autophagy and MMP-2 and MMP-9 further caused glycocalyx disruption. CONCLUSION: LSS promotes autophagy, leading to glycocalyx disruption. Autophagy increases the expression of MMP-2 and MMP-9, which are correlated with the glycocalyx destruction induced by LSS.
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Glicocálix , Metaloproteinasa 2 de la Matriz , Humanos , Glicocálix/metabolismo , Metaloproteinasa 2 de la Matriz/metabolismo , Metaloproteinasa 9 de la Matriz/metabolismo , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Autofagia , Estrés MecánicoRESUMEN
Cadmium (Cd) is a naturally occurring, toxic environmental metal found in foods. Humans do not have an efficient mechanism for Cd elimination; thus, Cd burden in humans increases with age. Cell and mouse studies show that Cd burden from low environmental levels of exposure impacts lung cell metabolism, proliferation signaling and cell growth as part of disease-promoting profibrotic responses in the lungs. Prior integrative analysis of metabolomics and transcriptomics identified the zDHHC11 transcript as a central functional hub in response to Cd dose. zDHHC11 encodes a protein S-palmitoyltransferase, but no evidence is available for effects of Cd on protein S-palmitoylation. In the present research, we studied palmitoylation changes in response to Cd and found increased protein S-palmitoylation in human lung fibroblasts that was inhibited by 2-bromopalmitate (2-BP), an irreversible palmitoyltransferase inhibitor. Mass spectrometry-based proteomics showed palmitoylation of proteins involved in divalent metal transport and in fibrotic signaling. Mechanistic studies showed that 2-BP inhibited palmitoylation of divalent metal ion transporter ZIP14 and also inhibited cellular Cd uptake. Transcription analyses showed that Cd stimulated transforming growth factor (TGF)-ß1 and ß3 expression within 8 h and lung fibrotic markers α-smooth muscle actin, matrix metalloproteinase-2, and collagen 1α1 gene expression and that these effects were blocked by 2-BP. Because 2-BP also blocked palmitoylation of proteins controlled by TGFß1, these results show that palmitoylation impacts Cd-dependent fibrotic signaling both by enhancing cellular Cd accumulation and by supporting post-translational processing of TGFß1-dependent proteins.
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Cadmio , Metaloproteinasa 2 de la Matriz , Humanos , Ratones , Animales , Cadmio/toxicidad , Cadmio/metabolismo , Metaloproteinasa 2 de la Matriz/metabolismo , Lipoilación , Pulmón , Transducción de Señal , Fibrosis , Fibroblastos , Factor de Crecimiento Transformador beta1/metabolismoRESUMEN
Secreted protein acidic and cysteine rich/osteonectin, cwcv and kazal-like domain proteoglycan 2 (SPOCK2) is a protein that regulates cell differentiation and growth. Recent studies have reported that SPOCK2 plays important roles in the progression of various human cancers; however, the role of SPOCK2 in melanoma remains unknown. Therefore, this study investigated the roles of SPOCK2 and the related mechanisms in melanoma progression. To evaluate the clinical significance of SPOCK2 expression in patients with melanoma, we analysed the association between SPOCK2 expression and its prognostic value for patients with melanoma using systematic multiomic analysis. Subsequently, to investigate the roles of Spock2 in melanoma progression in vitro and in vivo, we knocked down Spock2 in the B16F10 melanoma cell line. High SPOCK2 levels were positively associated with good prognosis and long survival rate of patients with melanoma. Spock2 knockdown promoted melanoma cell proliferation by inducing the cell cycle and inhibiting apoptosis. Moreover, Spock2 downregulation significantly increased cell migration and invasion by upregulating MMP2 and MT1-MMP. The increased cell proliferation and migration were inhibited by MAPK inhibitor, and ERK phosphorylation was considerably enhanced in Spock2 knockdown cells. Therefore, Spock2 could function as a tumour suppressor gene to regulate melanoma progression by regulating the MAPK/ERK signalling pathway. Additionally, Spock2 knockdown cell injection induced considerable tumour growth and lung metastasis in C57BL6 mice compared to that in the control group. Our findings suggest that SPOCK2 plays crucial roles in malignant progression of melanoma and functions as a novel therapeutic target of melanoma.
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Apoptosis , Movimiento Celular , Proliferación Celular , Progresión de la Enfermedad , Melanoma , Neoplasias Cutáneas , Animales , Femenino , Humanos , Masculino , Ratones , Biomarcadores de Tumor/metabolismo , Biomarcadores de Tumor/genética , Ciclo Celular , Línea Celular Tumoral , Técnicas de Silenciamiento del Gen , Sistema de Señalización de MAP Quinasas , Metaloproteinasa 14 de la Matriz/metabolismo , Metaloproteinasa 14 de la Matriz/genética , Metaloproteinasa 2 de la Matriz/metabolismo , Metaloproteinasa 2 de la Matriz/genética , Melanoma/genética , Melanoma/patología , Melanoma/metabolismo , Melanoma Experimental/genética , Melanoma Experimental/patología , Melanoma Experimental/metabolismo , Ratones Endogámicos C57BL , Invasividad Neoplásica , Pronóstico , Proteoglicanos/metabolismo , Proteoglicanos/genética , Neoplasias Cutáneas/patología , Neoplasias Cutáneas/genética , Neoplasias Cutáneas/metabolismoRESUMEN
BACKGROUND: Cancer-associated fibroblasts (CAFs) play an important role in the tumor microenvironment. Despite the well-known in vitro antitumoral effect of vitamin D3 (VD3), its impact on breast CAFs is almost unknown. In this study, we analyzed the ex vivo effects of calcitriol on CAFs isolated from breast cancer tissues. METHODS: CAFs were cultured with 1 and 10 nM calcitriol and their phenotype; gene expression, protein expression, and secretion were assessed. Calcitriol-treated CAFs-conditioned media (CM) were used to analyze the effect of CAFs on the migration and protein expression of MCF-7 and MDA-MB-231 cells. RESULTS: Tumor tissues from VD3-deficient patients exhibited lower levels of ß-catenin and TGFß1, along with higher levels of CYP24A1 compared to VD3-normal patients. In VD3-deficient patients, CAF infiltration was inversely associated with CYP24A1 levels and positively correlated with OPN levels. Calcitriol diminished CAFs' viability, but this effect was weaker in premenopausal and VD3-normal patients. Calcitriol reduced mRNA expression of CCL2, MMP9, TNC, and increased PDPN, SPP1, and TIMP1. It also decreased the secretion of CCL2, TNC, and the activity of MMP-2, while increasing cellular levels of TIMP1 in CAFs from all patient groups. In nonmetastatic and postmenopausal patients, PDPN surface expression increased, and CAFs CM from these groups decreased MCF-7 cell migration after ex vivo calcitriol treatment. In premenopausal and VD3-deficient patients, calcitriol reduced IDO1 expression in CAFs. Calcitriol-treated CAFs CM from these patients decreased OPN expression in MCF-7 and/or MDA-MB-231 cells. However, in premenopausal patients, calcitriol-treated CAFs CM also decreased E-cadherin expression in both cell lines. CONCLUSION: The effects of calcitriol on breast CAFs, both at the gene and protein levels, are complex, reflecting the immunosuppressive or procancer properties of CAFs. The anticancer polarization of CAFs following ex vivo calcitriol treatment may result from decreased CCL2, TNC (gene and protein), MMP9, and MMP-2, while the opposite effect may result from increased PDPN, TIMP1 (gene and protein), and SPP1. Despite these multifaceted effects of calcitriol on molecule expression, CAFs' CMs from nonmetastatic and postmenopausal patients treated ex vivo with calcitriol decreased the migration of MCF-7 cells.
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Neoplasias de la Mama , Fibroblastos Asociados al Cáncer , Humanos , Femenino , Fibroblastos Asociados al Cáncer/metabolismo , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Metaloproteinasa 9 de la Matriz/metabolismo , Metaloproteinasa 2 de la Matriz/metabolismo , Vitamina D3 24-Hidroxilasa/genética , Vitamina D3 24-Hidroxilasa/metabolismo , Colecalciferol , Calcitriol/farmacología , Fibroblastos/metabolismo , Movimiento Celular/genética , Línea Celular Tumoral , Microambiente Tumoral/genéticaRESUMEN
Hypertrophic ligamentum flavum (LF) is a main factor responsible for lumbar spinal stenosis (LSS); however, the exact mechanisms of the pathogenesis of these processes remain unknown. This study aimed to elucidate whether circular RNAs and microRNAs regulate the pathogenesis of LF and LSS, especially focusing on circPDK1 (hsa_circ_0057105), a circRNA targeting pyruvate dehydrogenase kinase 1 and differentially expressed in LF tissues between lumbar disk herniation and LSS patients. The circPDK1/miR-4731 and miR-4731/TNXB (Tenascin XB) interactions were predicted and validated by luciferase reporter assay. Colony formation, wound-healing, and MTT assays were used for estimating cell proliferation and migration. Protein expression levels were evaluated using Western blotting. TNXB expression was verified using immunohistochemistry (IHC). Overexpressing circPDK1 promoted the proliferation, migration, and expression of fibrosis-related protein (alpha smooth muscle actin (α-SMA), lysyl oxidase like 2 (LOXL2), Collagen I, matrix metalloproteinase-2 (MMP-2) and TNXB) in LF whereas miR-4731-5p showed opposite effects. The expression of TNXB was promoted by circPDK1; contrary results were observed with miR-4731-5p. Co-overexpression of miR-4731-5p partially reversed the proliferative and fibrosis-prompting effects of circPDK1 or TNXB. The circPDK1-miR-4731-TNXB pathway may be proposed as a regulatory axis in LF hypertrophy, which might shed light on in-depth research of LSS, as well as providing a novel therapeutic target for LF hypertrophy-induced LSS.
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Ligamento Amarillo , MicroARNs , Humanos , ARN Circular/genética , ARN Circular/metabolismo , Metaloproteinasa 2 de la Matriz/metabolismo , Ligamento Amarillo/metabolismo , MicroARNs/genética , MicroARNs/metabolismo , Fibrosis , Hipertrofia/metabolismoRESUMEN
BACKGROUND: Psoriasis is a T cell-mediated chronic inflammatory skin condition characterized by the interaction of T cells with various cell types, forming an inflammatory microenvironment that sustains psoriatic inflammation. Homeostasis of these tissue-resident T cells is supported by fibroblasts, the primary structural cells in the dermis. In psoriasis, there is increased expression of matrix metalloproteinase 2 (MMP2), mediating structural alterations in skin tissues and modulating inflammation. Additionally, the CD100-plexin-B2 (PLXNB2) axis is known to enhance psoriasis inflammation via keratinocytes, and CD103 levels are associated with the severity of psoriasis upon relapse. OBJECTIVES: To elucidate the role of fibroblasts and the MMP2-CD100 axis in modulating psoriasis inflammation. METHODS: CD100 expression and function in psoriasis were assessed using immunofluorescence, enzyme-linked immunosorbent assay, single-cell transcriptome sequencing, cellular interaction analyses and quantitative reverse transcriptase polymerase chain reaction. CD8+ T cells from people with psoriasis were isolated using magnetic beads, to investigate the regulatory effect of MMP2 on CD100 expression on their membranes. Single-cell transcriptome sequencing, spatial transcriptome sequencing, mimetic timing analysis, immunofluorescence and flow cytometry were used to determine the origin of MMP2 and its impact on CD103+ CD8+ T cells. The hypotheses were further validated in vivo using MMP2 and CD100 inhibitors. RESULTS: Soluble CD100 (sCD100) was significantly upregulated in both psoriatic lesions and peripheral blood, amplifying psoriasis inflammation by promoting the production of inflammatory cytokines by keratinocytes, fibroblasts and endothelial cells via the sCD100-PLXNB2 axis. Fibroblasts that highly expressed MMP2 (MMP2hi) exacerbated psoriasis symptoms by facilitating CD100 shedding from CD8+ T-cell membranes. Additionally, it was shown that fibroblasts enhance the upregulation of the CD8+ T-cell residency factor CD103 in co-cultures with CD8+ T cells. Inhibitors targeting MMP2 and CD100 were effective in reducing inflammation in an imiquimod-induced psoriasis model. CONCLUSIONS: Our findings underscore the pivotal role of MMP2hi fibroblasts in the amplification and recurrence of inflammatory responses in psoriasis. These fibroblasts augment psoriasis inflammation through the CD100-PLXNB2 axis by facilitating CD100 shedding on CD8+ T-cell membranes and by upregulating CD103, thereby enhancing CD8+ T-cell residency.
Psoriasis is a chronic skin condition. It involves the interaction of cells of the immune system (called T cells) with other cells in the body, causing inflammation. The main structural cells in the skin are called fibroblasts. Fibroblasts are important in skin healing and disease. In psoriasis, the body produces more of an enzyme called MMP2. MMP2 brings about structural alterations in skin tissues and controls inflammation. Proteins called CD100 and PLXNB2 increase the inflammation in psoriasis through another type of skin cell called keratinocytes. We investigated the role of fibroblasts and MMP2 and CD100 in psoriasis. To do this, the production and function of CD100 were assessed using a variety of laboratory techniques. We found that by controlling the enzymes MMP2 and MMP9, fibroblasts cause the release of CD100 from some T cells, encouraging the production of substances that promote inflammation, worsening psoriasis. This also caused fibroblasts to produce more MMP2, forming a cycle that increases the inflammation seen in psoriasis. Fibroblasts were found to increase the level of another protein called CD103 on some T cells, affecting how often psoriasis flares up. Our study highlights the important role of fibroblasts that express a lot of MMP2 in the inflammation found in people with psoriasis.
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Antígenos CD , Linfocitos T CD8-positivos , Fibroblastos , Metaloproteinasa 2 de la Matriz , Psoriasis , Psoriasis/inmunología , Humanos , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/metabolismo , Fibroblastos/metabolismo , Metaloproteinasa 2 de la Matriz/metabolismo , Antígenos CD/metabolismo , Ratones , Animales , Queratinocitos/metabolismo , Masculino , Femenino , Piel/inmunología , Piel/patología , Cadenas alfa de Integrinas/metabolismo , Células Cultivadas , SemaforinasRESUMEN
RESEARCH QUESTION: Is four and a half LIM domain 2 (FHL2) involved in trophoblast migration, invasion and epithelial-mesenchymal transition (EMT) in recurrent miscarriage? DESIGN: Villus tissue was collected from 24 patients who had experienced recurrent miscarriage and 24 healthy controls. FHL2 mRNA and protein expression in villus specimens were observed by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot. Small interfering RNA and overexpression plasmid were used to change the FHL2 expression. JAR and HTR8/SVneo cell lines were used to conduct scratch-wound assay and transwell assay to detect trophoblast migration and invasion of FHL2. Downstream molecule expression of mRNA and protein and EMT markers were verified by qRT-PCR and Western blot. RESULTS: Significantly lower FHL2 mRNA (Pâ¯=â¯0.019) and protein (Pâ¯=â¯0.0014) expression was found in trophoblasts from the recurrent miscarriage group compared with healthy controls. FHL2 knockdown repressed migration (Pâ¯=â¯0.0046), invasion (P < 0.001) and EMT, as shown by significant differences in mRNA and protein expression of the EMT markers N-cadherin, E-cadherin, Vimentin and Snail (all P < 0.05) of extravillus trophoblasts. FHL2 overexpression enhanced migration (Pâ¯=â¯0.025), invasion (P < 0.001) and EMT of extravillus trophoblasts (all EMT markers P < 0.05). The positive upstream factor FHL2 in the extracellular signal-related kinase pathway induced JunD expression, thereby promoting trophoblast migration and invasion via matrix metalloproteinase 2. CONCLUSIONS: FHL2 is involved in a regulatory pathway of trophoblast migration, invasion and EMT during early pregnancy, and may have a role in recurrent miscarriage pathogenesis, which can serve as a possible target for novel therapeutic development.