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1.
J Immunol Res ; 2022: 9166370, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35340587

RESUMO

Tumor necrosis factor-α (TNF-α) lies at the apex of signal transduction cascades that results in induced destruction of joints in rheumatoid arthritis. It is therefore of great medicinal interest to modulate the cellular responses to TNF-α. Ebosin, a novel exopolysaccharide derived from Streptomyces sp, has been demonstrated to have remarkable therapeutic actions on collagen-induced arthritis in rats, while it also suppressed the production of IL-1ß, TNF-α, and IL-6 at both mRNA and protein levels in cultured fibroblast-like synoviocytes. In order to further understand the potential mechanisms involved in the anti-inflammatory effects of ebosin at molecular level, we investigated the impact of it on the activation of MAPK and NF-κB pathways following TNF-α induced in fibroblast-like synoviocytes (FLS). The results showed that the phosphorylation levels of TNF-α-induced p38, JNK1, JNK2, IKKα, IKKß, and IκB, as well as NF-κB nuclear translocation, were reduced significantly in FLS cells in response to ebosin. Furthermore, we proved that ebosin decreased the level of NF-κB in the nucleus and blocked the DNA-binding ability of NF-κB using electrophoresis mobility gel shift assay. Besides, low levels of matrix metalloproteinases (MMP-1 and MMP-3) and chemokines (interleukin-8 and RANTES) were found in TNF-α-stimulated fibroblast-like synoviocytes treated with ebosin. These results indicate that ebosin can suppress a range of activities in both MAPK and NF-κB pathways induced by TNF-α in rat fibroblast-like synoviocytes, which provides a rationale for examining the use of ebosin as a potential therapeutic candidate for rheumatic arthritis.


Assuntos
NF-kappa B , Sinoviócitos , Animais , Fibroblastos , NF-kappa B/metabolismo , Polissacarídeos Bacterianos , Ratos , Sinoviócitos/metabolismo , Fator de Necrose Tumoral alfa/metabolismo
2.
In Vitro Cell Dev Biol Anim ; 58(2): 169-178, 2022 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-35194763

RESUMO

Cell adhesion to extracellular matrix proteins mediates resistance to radio- and chemotherapy by activating integrin signaling. In addition, mutual and cooperative interactions between integrin and growth factor receptor signaling contribute to the cellular radiation response. Here, we investigate to which extend the crosstalk between ß1 integrins and growth factor receptor signaling determines the cellular radiation response of fibroblasts by assessing clonogenic survival and cell cycling. By utilizing growth factor signaling competent and either ß1 integrin wildtype GD25ß1A fibroblasts or ß1 integrin mutant, signaling incompetent GD25ß1B fibroblasts, we show basal clonogenic survival to depend on growth factor receptor but not integrin signaling. Our data further suggest the cooperation between ß1 integrins and growth factor receptors to be critical for enhancing the radiation-induced G2/M cell cycle block leading to improved clonogenic radiation survival. By pharmacological inhibition of EGFR and PI3K, we additionally show that the essential contribution of EGFR signaling to radiogenic G2/M cell cycle arrest depends on the co-activation of the ß1 integrin signaling axis, but occurs independent of PI3K. Taken together, elucidation of the signaling circuitry underlying the EGFR/ß1 integrin crosstalk may support the development of advanced molecular targeted therapies for radiation oncology.


Assuntos
Integrina beta1 , Transdução de Sinais , Animais , Ciclo Celular , Fibroblastos/metabolismo , Integrina beta1/genética , Integrina beta1/metabolismo , Receptores de Fatores de Crescimento/metabolismo
3.
Anticancer Res ; 42(5): 2469-2477, 2022 May.
Artigo em Inglês | MEDLINE | ID: mdl-35489744

RESUMO

BACKGROUND/AIM: This study aimed to establish a setup for ultra-high-dose-rate (FLASH) carbon-ion irradiation, and to conduct the first human cell experiments using FLASH carbon ions. MATERIALS AND METHODS: A system for FLASH carbon-ion irradiation (1-3 Gy at 13 or 50 keV/µm) was developed. The growth and senescence of HFL1 lung fibroblasts were assessed by crystal violet staining assays and senescence-associated ß-galactosidase staining, respectively. Survival of HSGc-C5 cancer cells was assessed by clonogenic assays. RESULTS: The dose rates of carbon ions ranged from 96-195 Gy/s, meeting the definition of FLASH. With both 13 and 50 keV/µm beams, no FLASH sparing effect was observed on the growth suppression and senescence of HFL1 cells, nor on the survival of HSGc-C5 cells. CONCLUSION: We successfully conducted the first human cell experiments with FLASH carbon ions. No FLASH effect was observed under the conditions examined.


Assuntos
Carbono , Radioterapia com Íons Pesados , Fibroblastos/efeitos da radiação , Humanos , Íons
4.
Clin. transl. oncol. (Print) ; 24(7): 1354-1364, julio 2022.
Artigo em Inglês | IBECS | ID: ibc-203834

RESUMO

BackgroundGastric cancer (GC) is a malignancy that belongs to one of the most common leading causes of cancer death. Cancer-associated fibroblasts (CAFs) promote the GC cells’ malignant behavior. It is still unknown how GC converts normal fibroblasts (NFs) to CAFs.MethodsGC cells were co-cultured with NFs. Bioinformatics was used to analyze the genes and signaling pathways that were changed in fibroblast. RT-PCR, western blot, and Elisa assays were used to detect the expression of cytokines in fibroblast and condition medium. Western blot and immunofluorescence demonstrated activation of relevant pathways in CAFs-like cells. Transwell, scrape, colony formation, and CCK-8 assays were performed to reveal the feedback effect of CAFs-like cells on GC cells.ResultsGC promoted the conversion of NFs to CAFs by secreting Interleukin 17A (IL-17). It included both morphological and molecular marker changes. This process was achieved by activating the nuclear factor-κB (NF-κB) pathway. On the other hand, CAFs cells could secrete C-X-C Motif Chemokine Ligand 8 (IL-8, IL-8), which promoted the malignant phenotype of GC cells. In this way, a feedback loop of mutual influence was constructed in the GC and tumor microenvironment (TME).ConclusionsOur research proved a novel model of GC-educated NFs. GC-IL-17-fibroblast-IL-8-GC axis might be a potential pathway of the interaction between GC and TME.


Assuntos
Humanos , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Citocinas/metabolismo , Interleucina-17 , Interleucina-8 , Neoplasias Gástricas/patologia , Retroalimentação , Fibroblastos/metabolismo , Microambiente Tumoral
5.
Front Endocrinol (Lausanne) ; 13: 900791, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35707463

RESUMO

Periostin is a matricellular protein that is ubiquitously expressed in normal human tissues and is involved in pathologic mechanism of chronic inflammatory and fibrotic disease. In this study we investigate periostin in the pathogenesis of Graves' orbitopathy (GO) using human orbital adipose tissue obtained from surgery and primary cultured orbital fibroblasts in vitro. POSTN (gene encoding periostin) expression in Graves' orbital tissues and healthy control tissues was studied, and the role of periostin in GO pathologic mechanism was examined through small-interfering RNA (siRNA)-mediated silencing. POSTN gene expression was significantly higher in Graves' orbital tissues than healthy control tissues in real-time PCR results, and immunohistochemical staining revealed higher expression of periostin in Graves' orbital tissues than normal tissues. Silencing periostin using siRNA transfection significantly attenuated TGF-ß-induced profibrotic protein production and phosphorylated p38 and SMAD protein production. Knockdown of periostin inhibited interleukin-1 ß -induced proinflammatory cytokines production as well as phosphorylation of NF-κB and Ak signaling protein. Adipocyte differentiation was also suppressed in periostin-targeting siRNA transfected GO cells. We hypothesize that periostin contributes to the pathogenic process of inflammation, fibrosis and adipogenesis of GO. Our study provides in vitro evidence that periostin may be a novel potential therapeutic target for the treatment of GO.


Assuntos
Oftalmopatia de Graves , Adipogenia , Citocinas/metabolismo , Fibroblastos/metabolismo , Fibrose , Oftalmopatia de Graves/tratamento farmacológico , Humanos , Inflamação/metabolismo , RNA Interferente Pequeno/genética
6.
Front Immunol ; 13: 882718, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35707536

RESUMO

Over the past few decades, basic studies aimed at curing patients with cancer have been constantly evolving. A myriad of mechanistic studies on physiological changes and related factors in tumor growth and metastasis have been reported. Recently, several studies have been considerate to how tumors adapt to unfavorable environments, such as glucose deprivation, oxidative stress, hypoxic conditions, and immune responses. Tumors attempt to adapt to unfavorable environments with genetic or non-genetic changes, the alteration of metabolic signals, or the reconfiguration of their environment through migration to other organs. One of the distinct features in solid tumors is heterogeneity because their environments vary due to the characteristics of colony growth. For this reason, researchers are paying attention to the communication between growing tumors and neighboring environments, including stromal cells, immune cells, fibroblasts, and secreted molecules, such as proteins and RNAs. During cancer survival and progression, tumor cells undergo phenotype and molecular changes collectively referred to as cellular plasticity, which result from microenvironment signals, genetics and epigenetic alterations thereby contributing to tumor heterogeneity and therapy response. In this review, we herein discuss the adaptation process of tumors to adverse environments via communication with neighboring cells for overcoming unfavorable growth conditions. Understanding the physiology of these tumors and their communication with the tumor environment can help to develop promising tumor treatment strategies.


Assuntos
Neoplasias , Microambiente Tumoral , Fibroblastos/metabolismo , Humanos , Imunidade , Neoplasias/terapia , Células Estromais/metabolismo
7.
Mol Med ; 28(1): 64, 2022 06 11.
Artigo em Inglês | MEDLINE | ID: mdl-35690741

RESUMO

BACKGROUND: Hypoxia is one of the important characteristics of synovial microenvironment in rheumatoid arthritis (RA), and plays an important role in synovial hyperplasia. In terms of cell survival, fibroblast-like synovial cells (FLSs) are relatively affected by hypoxia. In contrast, fibroblast-like synovial cells from patients with RA (RA-FLSs) are particularly resistant to hypoxia-induced cell death. The purpose of this study was to evaluate whether fibroblast-like synovial cells in patients with osteoarthritis (OA-FLSs) and RA-FLSs have the same adaptation to hypoxia. METHODS: CCK-8, flow cytometry and BrdU were used to detect the proliferation of OA-FLSs and RA-FLSs under different oxygen concentrations. Apoptosis was detected by AV/PI, TUNEL and Western blot, mitophagy was observed by electron microscope, laser confocal microscope and Western blot, the state of mitochondria was detected by ROS and mitochondrial membrane potential by flow cytometry, BNIP3 and HIF-1α were detected by Western blot and RT-qPCR. The silencing of BNIP3 was achieved by stealth RNA system technology. RESULTS: After hypoxia, the survival rate of OA-FLSs decreased, while the proliferation activity of RA-FLSs further increased. Hypoxia induced an increase in apoptosis and inhibition of mitophagy in OA-FLSs, but not in RA-FLSs. Hypoxia led to a more lasting adaptive response. RA-FLSs displayed a more significant increase in the expression of genes transcriptionally regulated by HIF-1α. Interestingly, they showed higher BNIP3 expression than OA-FLSs, and showed stronger mitophagy and proliferation activities. BNIP3 siRNA experiment confirmed the potential role of BNIP3 in the survival of RA-FLSs. Inhibition of BNIP3 resulted in the decrease of cell proliferation, mitophagy and the increase of apoptosis. CONCLUSION: In summary, RA-FLSs maintained intracellular redox balance through mitophagy to promote cell survival under hypoxia. The mitophagy of OA-FLSs was too little to maintain the redox balance of mitochondria, resulting in apoptosis. The difference of mitophagy between OA-FLSs and RA-FLSs under hypoxia is mediated by the level of BNIP3 expression.


Assuntos
Artrite Reumatoide , Osteoartrite , Sinoviócitos , Artrite Reumatoide/genética , Proliferação de Células/genética , Células Cultivadas , Fibroblastos/metabolismo , Humanos , Hipóxia/metabolismo , Proteínas de Membrana/genética , Osteoartrite/genética , Osteoartrite/metabolismo , Proteínas Proto-Oncogênicas , Membrana Sinovial/metabolismo , Sinoviócitos/metabolismo
8.
Nat Commun ; 13(1): 3409, 2022 Jun 14.
Artigo em Inglês | MEDLINE | ID: mdl-35701396

RESUMO

Fibroblasts, the principal cell type of connective tissue, secrete extracellular matrix components during tissue development, homeostasis, repair and disease. Despite this crucial role, the identification and distinction of fibroblasts from other cell types are challenging and laden with caveats. Rapid progress in single-cell transcriptomics now yields detailed molecular portraits of fibroblasts and other cell types in our bodies, which complement and enrich classical histological and immunological descriptions, improve cell class definitions and guide further studies on the functional heterogeneity of cell subtypes and states, origins and fates in physiological and pathological processes. In this review, we summarize and discuss recent advances in the understanding of fibroblast identification and heterogeneity and how they discriminate from other cell types.


Assuntos
Tecido Conjuntivo , Fibroblastos , Fibroblastos/metabolismo
9.
Nat Cell Biol ; 24(6): 812-814, 2022 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-35654838
10.
Nat Cell Biol ; 24(6): 940-953, 2022 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-35654839

RESUMO

Bidirectional signalling between the tumour and stroma shapes tumour aggressiveness and metastasis. ATF4 is a major effector of the Integrated Stress Response, a homeostatic mechanism that couples cell growth and survival to bioenergetic demands. Using conditional knockout ATF4 mice, we show that global, or fibroblast-specific loss of host ATF4, results in deficient vascularization and a pronounced growth delay of syngeneic melanoma and pancreatic tumours. Single-cell transcriptomics of tumours grown in Atf4Δ/Δ mice uncovered a reduction in activation markers in perivascular cancer-associated fibroblasts (CAFs). Atf4Δ/Δ fibroblasts displayed significant defects in collagen biosynthesis and deposition and a reduced ability to support angiogenesis. Mechanistically, ATF4 regulates the expression of the Col1a1 gene and levels of glycine and proline, the major amino acids of collagen. Analyses of human melanoma and pancreatic tumours revealed a strong correlation between ATF4 and collagen levels. Our findings establish stromal ATF4 as a key driver of CAF functionality, malignant progression and metastasis.


Assuntos
Fibroblastos Associados a Câncer , Melanoma , Neoplasias Pancreáticas , Animais , Fibroblastos Associados a Câncer/metabolismo , Colágeno/metabolismo , Fibroblastos/metabolismo , Regulação Neoplásica da Expressão Gênica , Melanoma/genética , Camundongos , Camundongos Knockout , Neovascularização Patológica/metabolismo , Neoplasias Pancreáticas/patologia
11.
Stem Cell Res ; 62: 102826, 2022 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-35667217

RESUMO

An induced pluripotent stem cell (hiPSC) line (MPIi008-A) was generated from fibroblasts of a 1-year-old male patient with Denys-Drash syndrome using lentiviral delivery of reprogramming factors OCT4, SOX2, KLF4 and c-MYC. The MPIi008-A iPSC line exhibited typical iPSC morphology and normal karyotype, expressed pluripotent stem cell markers, and showed developmental potential to differentiate into derivatives of all three germ layers in vivo. The hiPSC line harbours a heterozygous missense mutation (R394L) in exon 9 of the WT1 gene.


Assuntos
Síndrome de Denys-Drash , Células-Tronco Pluripotentes Induzidas , Células-Tronco Pluripotentes , Diferenciação Celular , Síndrome de Denys-Drash/metabolismo , Fibroblastos/metabolismo , Heterozigoto , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Lactente , Masculino , Mutação
12.
Nature ; 606(7913): 396-405, 2022 06.
Artigo em Inglês | MEDLINE | ID: mdl-35650435

RESUMO

Disseminated cancer cells from primary tumours can seed in distal tissues, but may take several years to form overt metastases, a phenomenon that is termed tumour dormancy. Despite its importance in metastasis and residual disease, few studies have been able to successfully characterize dormancy within melanoma. Here we show that the aged lung microenvironment facilitates a permissive niche for efficient outgrowth of dormant disseminated cancer cells-in contrast to the aged skin, in which age-related changes suppress melanoma growth but drive dissemination. These microenvironmental complexities can be explained by the phenotype switching model, which argues that melanoma cells switch between a proliferative cell state and a slower-cycling, invasive state1-3. It was previously shown that dermal fibroblasts promote phenotype switching in melanoma during ageing4-8. We now identify WNT5A as an activator of dormancy in melanoma disseminated cancer cells within the lung, which initially enables the efficient dissemination and seeding of melanoma cells in metastatic niches. Age-induced reprogramming of lung fibroblasts increases their secretion of the soluble WNT antagonist sFRP1, which inhibits WNT5A in melanoma cells and thereby enables efficient metastatic outgrowth. We also identify the tyrosine kinase receptors AXL and MER as promoting a dormancy-to-reactivation axis within melanoma cells. Overall, we find that age-induced changes in distal metastatic microenvironments promote the efficient reactivation of dormant melanoma cells in the lung.


Assuntos
Envelhecimento , Pulmão , Melanoma , Metástase Neoplásica , Células Estromais , Microambiente Tumoral , Idoso , Envelhecimento/patologia , Fibroblastos/patologia , Humanos , Pulmão/patologia , Melanoma/patologia , Invasividade Neoplásica/patologia , Metástase Neoplásica/patologia , Neoplasia Residual , Proteínas Proto-Oncogênicas , Receptores Proteína Tirosina Quinases , Pele/patologia , Células Estromais/patologia , Proteína Wnt-5a , c-Mer Tirosina Quinase
13.
Cytokine ; 156: 155921, 2022 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-35667282

RESUMO

Systemic sclerosis (SSc) is an autoimmune prototypical connective tissues disease that results in alterations in vasculature, inflammation and fibrosis of the skin. Interleukin-1 family cytokines has been implicated in the disease including IL-1. IL-36α is an IL-1 family member that is clearly implicated in psoriatic skin disease but its role in systemic sclerosis disease is not clear. The aim of this work is to evaluate the levels and role of IL-36α in systemic sclerosis. Early diffuse SSc patients sera was isolated along with healthy controls and IL-36 levels quantified by ELISA. In vitro analysis was also undertaken with primary dermal fibroblasts with recombinant IL-36α and keratinocyte cells were also incubated with IL-36α. Cytokines were measured by ELISA. Serum IL-36 was significantly elevated compared to healthy controls. Elevated neutrophil elastase was also elevated in the matched sera. IL-36 was not directly pro-fibrotic in dermal fibroblasts but did induce pro-inflammatory cytokines that were dependant on the MAPK pathway for their release. IL-36α also led to release of CCL20 and CCL2 in keratinocytes which may potentiate fibrosis. IL-36α is elevated in SSc serum and whilst not directly pro-fibrotic it may through keratinocytes, potentiate fibrosis through keratinocyte-immune fibroblast cross-talk.


Assuntos
Interleucina-1/sangue , Esclerodermia Difusa , Escleroderma Sistêmico , Citocinas/metabolismo , Fibroblastos/metabolismo , Fibrose , Humanos , Interleucina-1/metabolismo , Interleucinas/metabolismo , Esclerodermia Difusa/metabolismo , Esclerodermia Difusa/patologia , Escleroderma Sistêmico/metabolismo , Pele/metabolismo
14.
Pharmazie ; 77(5): 132-136, 2022 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-35655385

RESUMO

Various chemical reagents containing inhibitors of mitochondrial activity, antioxidants, nuclear factor-kappa B (NF-kB) inhibitor, mammalian target of rapamycin (mTOR) inhibitor and other clinical therapeutics were screened in order to identify those that selectively decrease the viability of senescent human lung fibroblasts. Cell viability was measured using the CCK-8 assay. The results showed that pravastatin, a drug for hyperlipidemia, decreased the viability of senescent cells but not non-senescent cells. The effect of pravastatin on senescent cells is thought to be due to the inhibition of cell proliferation, rather than cell death. The effect of pravastatin was further investigated using the glucose metabolism assay, which showed that glucose consumption was inhibited both in non-senescent and senescent cells and intracellular nicotinamide adenine dinucleotide (NAD) was decreased in senescent cells. Changes to the mRNA expression levels of senescence-associated genes in response to pravastatin treatment were quantified by real-time-qPCR. There were no significant changes in the relative mRNA expression levels of IL-1ß, p16, p21, and p53 in pravastatin-treated non-senescent cells, whereas the expression of IL-1ß and p16 were increased by pravastatin only in senescent cells. The results of this study suggest that pravastatin does not induce senolysis, but rather selectively inhibits the proliferation of senescent cells and that cellular senescence is enhanced by decreasing intracellular NAD and promoting IL-1ß production.


Assuntos
NAD , Pravastatina , Fibroblastos/metabolismo , Humanos , Pulmão , NAD/metabolismo , NAD/farmacologia , Pravastatina/farmacologia , RNA Mensageiro/metabolismo
15.
Int J Mol Sci ; 23(11)2022 May 24.
Artigo em Inglês | MEDLINE | ID: mdl-35682566

RESUMO

Erythropoietin (Epo) is a crucial hormone regulating red blood cell number and consequently the hematocrit. Epo is mainly produced in the kidney by interstitial fibroblast-like cells. Previously, we have shown that in cultures of the immortalized mouse renal fibroblast-like cell line FAIK F3-5, sphingosine 1-phosphate (S1P), by activating S1P1 and S1P3 receptors, can stabilize hypoxia-inducible factor (HIF)-2α and upregulate Epo mRNA and protein synthesis. In this study, we have addressed the role of intracellular iS1P derived from sphingosine kinases (Sphk) 1 and 2 on Epo synthesis in F3-5 cells and in mouse primary cultures of renal fibroblasts. We show that stable knockdown of Sphk2 in F3-5 cells increases HIF-2α protein and Epo mRNA and protein levels, while Sphk1 knockdown leads to a reduction of hypoxia-stimulated HIF-2α and Epo protein. A similar effect was obtained using primary cultures of renal fibroblasts isolated from wildtype mice, Sphk1-/-, or Sphk2-/- mice. Furthermore, selective Sphk2 inhibitors mimicked the effect of genetic Sphk2 depletion and also upregulated HIF-2α and Epo protein levels. The combined blockade of Sphk1 and Sphk2, using Sphk2-/- renal fibroblasts treated with the Sphk1 inhibitor PF543, resulted in reduced HIF-2α and Epo compared to the untreated Sphk2-/- cells. Exogenous sphingosine (Sph) enhanced HIF-2α and Epo, and this was abolished by the combined treatment with the selective S1P1 and S1P3 antagonists NIBR-0213 and TY52156, suggesting that Sph was taken up by cells and converted to iS1P and exported to then act in an autocrine manner through S1P1 and S1P3. The upregulation of HIF-2α and Epo synthesis by Sphk2 knockdown was confirmed in the human hepatoma cell line Hep3B, which is well-established to upregulate Epo production under hypoxia. In summary, these data show that sphingolipids have diverse effects on Epo synthesis. While accumulation of intracellular Sph reduces Epo synthesis, iS1P will be exported to act through S1P1+3 to enhance Epo synthesis. Furthermore, these data suggest that selective inhibition of Sphk2 is an attractive new option to enhance Epo synthesis and thereby to reduce anemia development in chronic kidney disease.


Assuntos
Eritropoetina , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Esfingosina , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Epoetina alfa , Eritropoetina/genética , Eritropoetina/metabolismo , Fibroblastos/metabolismo , Hipóxia , Rim/metabolismo , Camundongos , RNA Mensageiro/genética , Esfingosina/metabolismo
16.
Int J Mol Sci ; 23(12)2022 Jun 18.
Artigo em Inglês | MEDLINE | ID: mdl-35743247

RESUMO

Osteoarthritis (OA) is characterized by the infiltration and adhesion of monocytes into the inflamed joint synovium. Interleukin (IL)-17 is a critical inflammatory mediator that participates in the progression of OA, although the mechanisms linking IL-17 and monocyte infiltration are not well understood. Our analysis of synovial tissue samples retrieved from the Gene Expression Omnibus (GEO) dataset exhibited higher monocyte marker (CD11b) and vascular cell adhesion molecule 1 (VCAM-1) levels in OA samples than in normal, healthy samples. The stimulation of human OA synovial fibroblasts (OASFs) with IL-17 increased VCAM-1 production and subsequently enhanced monocyte adhesion. IL-17 affected VCAM-1-dependent monocyte adhesion by reducing miR-5701 expression through the protein kinase C (PKC)-α and c-Jun N-terminal kinase (JNK) signaling cascades. Our findings improve our understanding about the effect of IL-17 on OA progression and, in particular, VCAM-1 production and monocyte adhesion, which may help with the design of more effective OA treatments.


Assuntos
MicroRNAs , Osteoartrite , Fibroblastos/metabolismo , Humanos , Interleucina-17/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , Monócitos/metabolismo , Osteoartrite/genética , Osteoartrite/metabolismo , Membrana Sinovial/metabolismo , Molécula 1 de Adesão de Célula Vascular/genética , Molécula 1 de Adesão de Célula Vascular/metabolismo
17.
Int J Mol Sci ; 23(12)2022 Jun 18.
Artigo em Inglês | MEDLINE | ID: mdl-35743248

RESUMO

Skin disorders are widespread around the world, affecting people of all ages, and oxidative stress represents one of the main causes of alteration in the normal physiological parameters of skin cells. In this work, we combined a natural protein, fibroin, with antioxidant compounds extracted in water from pomegranate waste. We demonstrate the effective and facile fabrication of bioactive and eco-sustainable films of potential interest for skin repair. The blended films are visually transparent (around 90%); flexible; stable in physiological conditions and in the presence of trypsin for 12 days; able to release the bioactive compounds in a controlled manner; based on Fickian diffusion; and biocompatible towards the main skin cells, keratinocytes and fibroblasts. Furthermore, reactive oxygen species (ROS) production tests demonstrated the high capacity of our films to reduce the oxidative stress induced in cells, which is responsible for various skin diseases.


Assuntos
Fibroínas , Romã (Fruta) , Fibroblastos , Humanos , Queratinócitos , Seda
18.
Int J Mol Sci ; 23(12)2022 Jun 20.
Artigo em Inglês | MEDLINE | ID: mdl-35743290

RESUMO

Uncontrolled proliferative diseases, such as fibrosis or cancer, can be fatal. We previously found that a compound containing the chromone scaffold (CS), ONG41008, had potent antifibrogenic effects associated with EMT or cell-cycle control resembling tumorigenesis. We investigated the effects of ONG41008 on tumor cells and compared these effects with those in pathogenic myofibroblasts. Stimulation of A549 (lung carcinoma epithelial cells) or PANC1 (pancreatic ductal carcinoma cells) with ONG41008 resulted in robust cellular senescence, indicating that dysregulated cell proliferation is common to fibrotic cells and tumor cells. The senescence was followed by multinucleation, a manifestation of mitotic slippage. There was significant upregulation of expression and rapid nuclear translocation of p-TP53 and p16 in the treated cancer cells, which thereafter died after 72 h confirmed by 6 day live imaging. ONG41008 exhibited a comparable senogenic potential to that of dasatinib. Interestingly, ONG41008 was only able to activate caspase-3, 7 in comparison with quercetin and fisetin, also containing CS in PANC1. ONG41008 did not seem to be essentially toxic to normal human lung fibroblasts or primary prostate epithelial cells, suggesting ONG41008 can distinguish the intracellular microenvironment between normal cells and aged or diseased cells. This effect might occur as a result of the increased NAD/NADH ratio, because ONG41008 restored this important metabolic ratio in cancer cells. Taken together, this is the first study to demonstrate that a small molecule can arrest uncontrolled proliferation during fibrogenesis or tumorigenesis via both senogenic and senolytic potential. ONG41008 could be a potential drug for a broad range of fibrotic or tumorigenic diseases.


Assuntos
Senescência Celular , Fibroblastos , Idoso , Carcinogênese/metabolismo , Dasatinibe/farmacologia , Fibroblastos/metabolismo , Humanos , Masculino , Quercetina/farmacologia , Microambiente Tumoral
19.
Int J Mol Sci ; 23(12)2022 Jun 20.
Artigo em Inglês | MEDLINE | ID: mdl-35743299

RESUMO

Wound healing pathologies are an increasing problem in ageing societies. Chronic, non-healing wounds, which cause high morbidity and severely reduce the quality of life of affected individuals, are frequently observed in aged individuals and people suffering from diseases affected by the Western lifestyle, such as diabetes. Causal treatments that support proper wound healing are still scarce. Here, we performed expression proteomics to study the effects of the small molecule TOP-N53 on primary human skin fibroblasts and keratinocytes. TOP-N53 is a dual-acting nitric oxide donor and phosphodiesterase-5 inhibitor increasing cGMP levels to support proper wound healing. In contrast to keratinocytes, which did not exhibit global proteome alterations, TOP-N53 had profound effects on the proteome of skin fibroblasts. In fibroblasts, TOP-N53 activated the cytoprotective, lysosomal degradation pathway autophagy and induced the expression of the selective autophagy receptor p62/SQSTM1. Thus, activation of autophagy might in part be responsible for beneficial effects of TOP-N53.


Assuntos
Doadores de Óxido Nítrico , Inibidores da Fosfodiesterase 5 , Idoso , Autofagia , Fibroblastos/metabolismo , Humanos , Queratinócitos/metabolismo , Óxido Nítrico/metabolismo , Doadores de Óxido Nítrico/metabolismo , Doadores de Óxido Nítrico/farmacologia , Inibidores da Fosfodiesterase 5/farmacologia , Proteoma/metabolismo , Qualidade de Vida , Pele/metabolismo
20.
Int J Mol Sci ; 23(12)2022 Jun 20.
Artigo em Inglês | MEDLINE | ID: mdl-35743318

RESUMO

Breast cancer-associated fibroblasts (BCAFs), the most abundant non-cancer stromal cells of the breast tumor microenvironment (TME), dramatically sustain breast cancer (BC) progression by interacting with BC cells. BCAFs, as well as myofibroblasts, display an up regulation of activation and inflammation markers represented by α-smooth muscle actin (α-SMA) and cyclooxygenase 2 (COX-2). BCAF aggregates have been identified in the peripheral blood of metastatic BC patients. We generated an in vitro stromal model consisting of human primary BCAFs grown as monolayers or 3D cell aggregates, namely spheroids and reverted BCAFs, obtained from BCAF spheroids reverted to 2D cell adhesion growth after 216 h of 3D culture. We firstly evaluated the state of activation and inflammation and the mesenchymal status of the BCAF monolayers, BCAF spheroids and reverted BCAFs. Then, we analyzed the MCF-7 cell viability and migration following treatment with conditioned media from the different BCAF cultures. After 216 h of 3D culture, the BCAFs acquired an inactivated phenotype, associated with a significant reduction in α-SMA and COX-2 protein expression. The deactivation of the BCAF spheroids at 216 h was further confirmed by the cytostatic effect exerted by their conditioned medium on MCF-7 cells. Interestingly, the reverted BCAFs also retained a less activated phenotype as indicated by α-SMA protein expression reduction. Furthermore, the reverted BCAFs exhibited a reduced pro-tumor phenotype as indicated by the anti-migratory effect exerted by their conditioned medium on MCF-7 cells. The deactivation of BCAFs without drug treatment is possible and leads to a reduced capability of BCAFs to sustain BC progression in vitro. Consequently, this study could be a starting point to develop new therapeutic strategies targeting BCAFs and their interactions with cancer cells.


Assuntos
Neoplasias da Mama , Fibroblastos Associados a Câncer , Neoplasias da Mama/metabolismo , Fibroblastos Associados a Câncer/metabolismo , Linhagem Celular Tumoral , Meios de Cultivo Condicionados/metabolismo , Ciclo-Oxigenase 2/genética , Ciclo-Oxigenase 2/metabolismo , Feminino , Fibroblastos/metabolismo , Humanos , Inflamação/patologia , Células Estromais/metabolismo , Microambiente Tumoral
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