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1.
Anticancer Res ; 41(9): 4249-4258, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34475044

RESUMO

BACKGROUND/AIM: Recent studies have indicated the clinical significance of tumor-associated macrophages (TAMs) in breast cancer; however, the detailed mechanisms of cell-cell interactions between TAMs and cancer cells remain unclear. MATERIALS AND METHODS: In vitro cell culture studies using human monocyte-derived macrophages and breast cancer cell lines were performed to test which cytokines would be involved in cell-cell interactions between cancer cells and macrophages. In addition, studies using human resected samples and animal breast cancer models were performed to examine the significance of TAMs in cancer development. RESULTS: Osteopontin, HB-EGF, and IL-6 were suggested to be macrophage-derived growth factors for breast cancer cells. FROUNT inhibitor significantly blocked TAM infiltration and subcutaneous tumor growth in an E0771 mouse breast cancer model. CONCLUSION: TAMs express growth factors, such as osteopontin, for cancer cells, and targeting of TAM infiltration might be a promising approach for anti-breast cancer therapy.


Assuntos
Neoplasias da Mama/patologia , Fator de Crescimento Semelhante a EGF de Ligação à Heparina/genética , Interleucina-6/genética , Macrófagos/citologia , Osteopontina/genética , Macrófagos Associados a Tumor/citologia , Animais , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Comunicação Celular , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Células Cultivadas , Técnicas de Cocultura , Feminino , Fator de Crescimento Semelhante a EGF de Ligação à Heparina/metabolismo , Humanos , Interleucina-6/metabolismo , Células MCF-7 , Macrófagos/metabolismo , Camundongos , Transplante de Neoplasias , Osteopontina/metabolismo , Microambiente Tumoral , Macrófagos Associados a Tumor/metabolismo , Macrófagos Associados a Tumor/patologia
2.
Biomolecules ; 11(7)2021 07 16.
Artigo em Inglês | MEDLINE | ID: mdl-34356671

RESUMO

Unprecedented advances in secondary prevention have greatly improved the prognosis of cardiovascular diseases (CVDs); however, CVDs remain a leading cause of death globally. These findings suggest the need to reconsider cardiovascular risk and optimal medical therapy. Numerous studies have shown that inflammation, pro-thrombotic factors, and gene mutations are focused not only on cardiovascular residual risk but also as the next therapeutic target for CVDs. Furthermore, recent clinical trials, such as the Canakinumab Anti-inflammatory Thrombosis Outcomes Study trial, showed the possibility of anti-inflammatory therapy for patients with CVDs. Osteopontin (OPN) is a matricellular protein that mediates diverse biological functions and is involved in a number of pathological states in CVDs. OPN has a two-faced phenotype that is dependent on the pathological state. Acute increases in OPN have protective roles, including wound healing, neovascularization, and amelioration of vascular calcification. By contrast, chronic increases in OPN predict poor prognosis of a major adverse cardiovascular event independent of conventional cardiovascular risk factors. Thus, OPN can be a therapeutic target for CVDs but is not clinically available. In this review, we discuss the role of OPN in the development of CVDs and its potential as a therapeutic target.


Assuntos
Doenças Cardiovasculares/metabolismo , Osteopontina/metabolismo , Injúria Renal Aguda/metabolismo , Animais , Biomarcadores/análise , Síndrome Cardiorrenal/metabolismo , Humanos , Camundongos , Osteopontina/análise , Osteopontina/genética
3.
Mol Med Rep ; 24(4)2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-34328202

RESUMO

A number of drugs and other triggers can cause acute liver injury (ALI) in clinical practice. Therefore, identifying a safe drug for the prevention of liver injury is important. The aim of the present study was to investigate the potential preventive effect and regulatory mechanism of urantide on carbon tetrachloride (CCl4)­induced ALI by investigating the expression of components of the MAPK signalling pathway and the urotensin II (UII)/urotensin receptor (UT) system. Liver oedema and severe fatty degeneration of the cytoplasm were observed in ALI model rats, and the serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels were found to be significantly increased. Compared with those in the ALI model group, ALT and AST levels and the liver index did not significantly increase in each group given the preventive administration of urantide, and the liver tissue morphology was correspondingly protected. Moreover, the gene and protein expression levels of UII, G protein­coupled receptor (GPR14) and the oxidative stress­sensitive cytokines, α­smooth muscle actin and osteopontin were decreased, indicating that the protein translation process was effectively maintained. However, the expression levels of MAPK signalling pathway­related proteins and genes were decreased. It was found that urantide could effectively block the MAPK signalling pathway by antagonizing the UII/UT system, thus protecting the livers of ALI model rats. Therefore, it was suggested that ALI may be associated with the MAPK signalling pathway, and effective inhibition of the MAPK signalling pathway may be critical in protecting the liver.


Assuntos
Doença Hepática Induzida por Substâncias e Drogas/metabolismo , Doença Hepática Induzida por Substâncias e Drogas/prevenção & controle , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Fragmentos de Peptídeos/farmacologia , Urotensinas/farmacologia , Actinas/genética , Actinas/metabolismo , Animais , Tetracloreto de Carbono/toxicidade , Modelos Animais de Doenças , MAP Quinases Reguladas por Sinal Extracelular/genética , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , MAP Quinase Quinase 4/genética , MAP Quinase Quinase 4/metabolismo , Masculino , Osteopontina/genética , Osteopontina/metabolismo , Fragmentos de Peptídeos/uso terapêutico , Ratos Sprague-Dawley , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Urotensinas/uso terapêutico , Proteínas Quinases p38 Ativadas por Mitógeno/genética , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
4.
Environ Toxicol Pharmacol ; 87: 103700, 2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-34237469

RESUMO

Glyphosate [N-(phosphonomethyl)glycine] is the active ingredient in widely used broad-spectrum herbicides. Even though the toxicity mechanism of this herbicide in vertebrates is poorly understood, evidence suggests that glyphosate is an endocrine disruptor capable of producing morphological anomalies as well as cardiotoxic and neurotoxic effects. We used the zebrafish model to assess the effects of early life glyphosate exposure on the development of cartilage and bone tissues and organismal responses. We found functional alterations, including a reduction in the cardiac rate, significant changes in the spontaneous tail movement pattern, and defects in craniofacial development. These effects were concomitant with alterations in the level of the estrogen receptor alpha osteopontin and bone sialoprotein. We also found that embryos exposed to glyphosate presented spine deformities as adults. These developmental alterations are likely induced by changes in protein levels related to bone and cartilage formation.


Assuntos
Osso e Ossos/efeitos dos fármacos , Anormalidades Craniofaciais/induzido quimicamente , Glicina/análogos & derivados , Herbicidas/toxicidade , Teratógenos/toxicidade , Animais , Osso e Ossos/anormalidades , Anormalidades Craniofaciais/metabolismo , Anormalidades Craniofaciais/veterinária , Embrião não Mamífero/anormalidades , Embrião não Mamífero/efeitos dos fármacos , Embrião não Mamífero/metabolismo , Receptor alfa de Estrogênio/metabolismo , Feminino , Proteínas de Peixes/metabolismo , Glicina/toxicidade , Frequência Cardíaca/efeitos dos fármacos , Locomoção/efeitos dos fármacos , Masculino , Osteopontina/metabolismo , Sialoglicoproteínas/metabolismo , Peixe-Zebra/anormalidades , Peixe-Zebra/metabolismo
5.
Int J Mol Sci ; 22(10)2021 May 17.
Artigo em Inglês | MEDLINE | ID: mdl-34067898

RESUMO

The aim of the study was to examine the efficacy of cold atmospheric plasma (CAP) on the mineralization and cell proliferation of murine dental cementoblasts. Cells were treated with CAP and enamel matrix derivates (EMD). Gene expression of alkaline phosphatase (ALP), bone gamma-carboxyglutamate protein (BGLAP), periostin (POSTN), osteopontin (OPN), osterix (OSX), collagen type I alpha 1 chain (COL1A1), dentin matrix acidic phosphoprotein (DMP)1, RUNX family transcription factor (RUNX)2, and marker of proliferation Ki-67 (KI67) was quantified by real-time PCR. Protein expression was analyzed by immunocytochemistry and ELISA. ALP activity was determined by ALP assay. Von Kossa and alizarin red staining were used to display mineralization. Cell viability was analyzed by XTT assay, and morphological characterization was performed by DAPI/phalloidin staining. Cell migration was quantified with an established scratch assay. CAP and EMD upregulated both mRNA and protein synthesis of ALP, POSTN, and OPN. Additionally, DMP1 and COL1A1 were upregulated at both gene and protein levels. In addition to upregulated RUNX2 mRNA levels, treated cells mineralized more intensively. Moreover, CAP treatment resulted in an upregulation of KI67, higher cell viability, and improved cell migration. Our study shows that CAP appears to have stimulatory effects on regeneration-associated cell functions in cementoblasts.


Assuntos
Cementogênese/efeitos dos fármacos , Cemento Dentário/metabolismo , Gases em Plasma/farmacologia , Animais , Calcificação Fisiológica/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/genética , Camundongos , Osteocalcina/metabolismo , Osteopontina/metabolismo , Gases em Plasma/metabolismo , Transcriptoma/genética
6.
Biomolecules ; 11(6)2021 05 30.
Artigo em Inglês | MEDLINE | ID: mdl-34070790

RESUMO

Since the original description in 1996, the interaction between the cytokine osteopontin (OPN) and the homing receptor CD44 has been extensively studied in cancer, inflammation, bone remodeling, and various other conditions. Alternative splicing and extensive posttranslational modifications by both binding partners, as well as the possibility for lateral recruitment of additional membrane receptors or soluble co-ligands into a complex have left the exact molecular requirements for high-affinity OPN-CD44 binding unresolved. We now report that there is a moderate engagement between the unmodified molecules, which results in curved double-reciprocal plots for OPN titration, suggesting the existence of two binding sites or two binding conformations. Structural constraint of OPN, by immobilization or by addition of heparin, is required for its strong ligation of CD44. Prior literature provides evidence that heparin binding to OPN prompts the unfolding of a core element in the protein. This conformational adjustment may be essential for efficient CD44 interaction. The integrin α9ß1 seems to compete with the OPN-CD44 engagement, while the integrin αVß3 reflects additive binding, suggesting that the CD44 contact sites on OPN are downstream of the RGD motif but overlap with the SVVYGLR domain. Hyaluronate has no effect, placing the relevant domain on CD44 downstream of the N-terminus.


Assuntos
Receptores de Hialuronatos/química , Osteopontina/química , Humanos , Receptores de Hialuronatos/metabolismo , Osteopontina/metabolismo , Ligação Proteica , Domínios Proteicos , Estrutura Quaternária de Proteína
7.
PLoS Pathog ; 17(6): e1009618, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-34106998

RESUMO

Subpopulations of B-lymphocytes traffic to different sites and organs to provide diverse and tissue-specific functions. Here, we provide evidence that epigenetic differences confer a neuroinvasive phenotype. An EBV+ B cell lymphoma cell line (M14) with low frequency trafficking to the CNS was neuroadapted to generate a highly neuroinvasive B-cell population (MUN14). MUN14 B cells efficiently infiltrated the CNS within one week and produced neurological pathologies. We compared the gene expression profiles of viral and cellular genes using RNA-Seq and identified one viral (EBNA1) and several cellular gene candidates, including secreted phosphoprotein 1/osteopontin (SPP1/OPN), neuron navigator 3 (NAV3), CXCR4, and germinal center-associated signaling and motility protein (GCSAM) that were selectively upregulated in MUN14. ATAC-Seq and ChIP-qPCR revealed that these gene expression changes correlated with epigenetic changes at gene regulatory elements. The neuroinvasive phenotype could be attenuated with a neutralizing antibody to OPN, confirming the functional role of this protein in trafficking EBV+ B cells to the CNS. These studies indicate that B-cell trafficking to the CNS can be acquired by epigenetic adaptations and provide a new model to study B-cell neuroinvasion associated CNS lymphoma and autoimmune disease of the CNS, including multiple sclerosis (MS).


Assuntos
Linfócitos B/patologia , Linfócitos B/virologia , Neoplasias do Sistema Nervoso Central/virologia , Epigênese Genética , Infecções por Vírus Epstein-Barr/patologia , Animais , Linfócitos B/metabolismo , Transformação Celular Viral/fisiologia , Neoplasias do Sistema Nervoso Central/metabolismo , Neoplasias do Sistema Nervoso Central/patologia , Infecções por Vírus Epstein-Barr/genética , Infecções por Vírus Epstein-Barr/metabolismo , Herpesvirus Humano 4 , Linfoma/metabolismo , Linfoma/patologia , Linfoma/virologia , Camundongos , Osteopontina/metabolismo
8.
Int J Nanomedicine ; 16: 3633-3648, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34079254

RESUMO

Purpose: The deposition of hydroxyapatite (HAp) crystals plays an important role in the development of vascular calcification (VC). This study aimed to demonstrate the effects of nanosized HAp (nHAp) on vascular smooth muscle cells (VSMCs) and VC progression. Methods: Transmission electron microscopy (TEM) was used to examine cellular uptake of nHAp. Cell viability was determined using CCK-8 assay kit. Mitochondrial impairment and reactive oxygen species were detected by TEM and fluorescence dye staining, respectively. Cell apoptosis was detected by Western blot analysis and Annexin V staining. Mouse model of VC was built via applying nHAp on the surface of abdominal aorta. Calcification was visualized by Alizarin red and von Kossa staining. Results: We found that nHAp could promote osteogenic transformation of VSMCs by elevating expression of runt-related factor 2 (Runx2), osteopontin (OPN) and alkaline phosphatase (ALP), impairing function and morphology of mitochondria and inducing apoptosis of VSMCs. More phosphorylation of c-Jun N-terminal protein kinase/c-JUN (JNK/c-JUN) in VSMCs was detected after mixing nHAp with VSMCs. HAp-induced osteogenic transformation of VSMCs was blocked by JNK inhibitor SP600125, resulted in decreased ALP activity, less Runx2 and OPN expressions. SP600125 also inhibited apoptosis of VSMCs. Application of nHAp to outside of aorta induced osteogenic transformation and apoptosis of VSMCs, and significant deposition of calcium on the vessel walls of mice, which can be effectively attenuated by SP600125. Conclusion: JNK/c-JUN signaling pathway is critical for nHAp-induced calcification, which could be a potential therapeutic target for controlling the progression of VC.


Assuntos
Apoptose/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Durapatita/farmacologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Músculo Liso Vascular/citologia , Nanoestruturas/química , Osteogênese/efeitos dos fármacos , Fosfatase Alcalina/metabolismo , Animais , Cálcio/metabolismo , Células Cultivadas , Durapatita/química , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Camundongos , Osteopontina/metabolismo , Espécies Reativas de Oxigênio/metabolismo
9.
Res Vet Sci ; 138: 167-177, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34153557

RESUMO

Adhesion molecules play an important role in urinary calculus formation. The expressions of adhesion molecules in renal tubular has been reported in some animals. However, the role of adhesion molecules in the process of sheep urinary calculus formation is still unclear. The magnesium ammonium phosphate (MAP) is the main component of sheep urinary calculus. In this paper, the sheep renal tubular epithelial cells (RTECs) were isolated and treated with MAP, the expressions of osteopontin (OPN), intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1) and apoptosis-related indicators caspase-3, Bcl-2 and Bax in RTECs were observed, the viability of RTECs was detected by Cell Counting Kit-8 (CCK-8). The levels of superoxide dismutase (SOD) and malondialdehyde (MDA), and the expressions of inflammatory factors Interleukin-6 (IL-6), Interleukin-1 (IL-1), Interleukin-17 (IL-17) and tumor necrosis factor-α (TNF-α) were measured by enzyme-linked immunosorbent (ELISA). The histopathological observation of kidney in urolithiasis sheep was made. The results showed that MAP could reduce the viability and SOD activity, enhance the activity of MDA significantly and promote the expressions of IL-1, IL-6, IL-17 and TNF-α of RTECs. By western blot and qPCR methods, the expressions of ICAM-1, VCAM-1 and OPN increased in 48 h. In addition, the expression of caspase-3 increased significantly and the ratio of Bcl-2/Bax reduced with exposure to MAP. The renal tissue structure was seriously damaged, the RTECs in urolithiasis sheep were degenerative and necrotic.


Assuntos
Apoptose , Moléculas de Adesão Celular/metabolismo , Sobrevivência Celular , Citocinas/imunologia , Células Epiteliais/fisiologia , Estresse Oxidativo , Estruvita/metabolismo , Animais , Células Cultivadas , Molécula 1 de Adesão Intercelular/metabolismo , Rim/fisiologia , Osteopontina/metabolismo , Carneiro Doméstico/metabolismo , Carneiro Doméstico/urina , Cálculos Urinários/metabolismo , Molécula 1 de Adesão de Célula Vascular/metabolismo
10.
Cancer Invest ; 39(6-7): 559-570, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34043476

RESUMO

The study evaluated whether SPP1/osteopontin (OPN) splice variants are differentially expressed in nonmelanoma skin cancer compared to normal skin. The absolute number of mRNA molecules of OPN-a predominated in normal skin and nonmelanoma skin cancer compared to OPN-b, OPN-c, and OPN-5. However, mRNAs of OPN-a, OPN-b, and OPN-c were expressed in higher levels in cutaneous squamous cell carcinomas (cSCCs) and basal cell carcinomas relative to normal skin. Additionally, OPN-5 expression was higher than OPN-b and OPN-c, and OPN-c, in normal skin and nonmelanoma skin cancer, respectively. Furthermore, we identified four OPN-5 splice variants, which were cloned and analyzed for protein expression.


Assuntos
Processamento Alternativo , Carcinoma Basocelular/genética , Carcinoma de Células Escamosas/genética , Osteopontina/metabolismo , Neoplasias Cutâneas/genética , Idoso , Idoso de 80 Anos ou mais , Carcinoma Basocelular/metabolismo , Carcinoma de Células Escamosas/metabolismo , Linhagem Celular Tumoral , Clonagem Molecular , Feminino , Regulação Neoplásica da Expressão Gênica , Variação Genética , Humanos , Masculino , Pessoa de Meia-Idade , Osteopontina/genética , Isoformas de RNA/metabolismo , Neoplasias Cutâneas/metabolismo , Regulação para Cima
11.
Elife ; 102021 05 19.
Artigo em Inglês | MEDLINE | ID: mdl-34009124

RESUMO

To study disease development, an inventory of an organ's cell types and understanding of physiologic function is paramount. Here, we performed single-cell RNA-sequencing to examine heterogeneity of murine pancreatic duct cells, pancreatobiliary cells, and intrapancreatic bile duct cells. We describe an epithelial-mesenchymal transitory axis in our three pancreatic duct subpopulations and identify osteopontin as a regulator of this fate decision as well as human duct cell dedifferentiation. Our results further identify functional heterogeneity within pancreatic duct subpopulations by elucidating a role for geminin in accumulation of DNA damage in the setting of chronic pancreatitis. Our findings implicate diverse functional roles for subpopulations of pancreatic duct cells in maintenance of duct cell identity and disease progression and establish a comprehensive road map of murine pancreatic duct cell, pancreatobiliary cell, and intrapancreatic bile duct cell homeostasis.


Assuntos
Perfilação da Expressão Gênica , Heterogeneidade Genética , Ductos Pancreáticos/citologia , Análise de Célula Única , Transcriptoma , Animais , Linhagem Celular , Separação Celular , Dano ao DNA , Bases de Dados Genéticas , Modelos Animais de Doenças , Transição Epitelial-Mesenquimal , Feminino , Geminina/genética , Geminina/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Morfogênese , Osteopontina/genética , Osteopontina/metabolismo , Ductos Pancreáticos/metabolismo , Pancreatite Crônica/genética , Pancreatite Crônica/metabolismo , Pancreatite Crônica/patologia , Fenótipo , RNA-Seq
12.
Toxicol Appl Pharmacol ; 422: 115559, 2021 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-33961903

RESUMO

The occurrence and development of silicosis is related to the interaction of multiple cells through signal transmission caused by silica dust. Including inflammatory changes reduced by macrophages and phenotypic transdifferentiation reduced by lung fibroblasts. As a communication medium between cells, exosomes have become a hot research topic. To explore the role of exosomal proteins in the occurrence and development of silicosis and the possible intervention targets, this study conducted proteomic analysis of macrophage-derived exosomes induced by silica, to identify specific proteins for intervention. In this study, we used proteomic analysis to screen exosomal protein profiles from the RAW264.7 macrophages exposed to silica. A total of 291 proteins were differentially expressed, of which 178 were upregulated and 113 were downregulated. By performing functional annotation and analysis of the differentially expressed proteins, we identified proteins SPP1, HMGB3, and HNRNPAB, which were consistent with the proteomics analysis. The involvement of SPP1 protein in fibrosis was studied further. Knocking down the expression of SPP1 in exosomes resulted in a decrease in fibrosis-related indicators. These results help to understand that exosomal protein can mediate cell communication and play a key role in the transition from fibroblasts to myofibroblasts. Further, this study also provided strategies and scientific basis for future studies on the intervention of silicosis.


Assuntos
Comunicação Celular , Transdiferenciação Celular , Exossomos/efeitos dos fármacos , Fibroblastos/metabolismo , Macrófagos/efeitos dos fármacos , Osteopontina/metabolismo , Dióxido de Silício/toxicidade , Silicose/metabolismo , Animais , Proliferação de Células , Exossomos/genética , Exossomos/metabolismo , Fibroblastos/patologia , Macrófagos/metabolismo , Camundongos , Miofibroblastos/metabolismo , Miofibroblastos/patologia , Células NIH 3T3 , Osteopontina/genética , Proteoma , Proteômica , Células RAW 264.7 , Silicose/genética , Silicose/patologia
13.
Biochemistry ; 60(17): 1347-1355, 2021 05 04.
Artigo em Inglês | MEDLINE | ID: mdl-33876640

RESUMO

Protein phosphorylation is an abundant post-translational modification (PTM) and an essential modulator of protein functionality in living cells. Intrinsically disordered proteins (IDPs) are particular targets of PTM protein kinases due to their involvement in fundamental protein interaction networks. Despite their dynamic nature, IDPs are far from having random-coil conformations but exhibit significant structural heterogeneity. Changes in the molecular environment, most prominently in the form of PTM via phosphorylation, can modulate these structural features. Therefore, how phosphorylation events can alter conformational ensembles of IDPs and their interactions with binding partners is of great interest. Here we study the effects of hyperphosphorylation on the IDP osteopontin (OPN), an extracellular target of the Fam20C kinase. We report a full characterization of the phosphorylation sites of OPN using a combined nuclear magnetic resonance/mass spectrometry approach and provide evidence for an increase in the local flexibility of highly phosphorylated regions and the ensuing overall structural elongation. Our study emphasizes the simultaneous importance of electrostatic and hydrophobic interactions in the formation of compact substates in IDPs and their relevance for molecular recognition events.


Assuntos
Osteopontina/química , Osteopontina/metabolismo , Humanos , Simulação de Dinâmica Molecular , Fosforilação , Conformação Proteica , Dobramento de Proteína
14.
Cells ; 10(3)2021 03 11.
Artigo em Inglês | MEDLINE | ID: mdl-33799608

RESUMO

Pulmonary fibrosis is the chronic-progressive replacement of healthy lung tissue by extracellular matrix, leading to the destruction of the alveolar architecture and ultimately death. Due to limited pathophysiological knowledge, causal therapies are still missing and consequently the prognosis is poor. Thus, there is an urgent clinical need for models to derive effective therapies. Polo-like kinase 2 (PLK2) is an emerging regulator of fibroblast function and fibrosis. We found a significant downregulation of PLK2 in four different entities of human pulmonary fibrosis. Therefore, we characterized the pulmonary phenotype of PLK2 knockout (KO) mice. Isolated pulmonary PLK2 KO fibroblasts displayed a pronounced myofibroblast phenotype reflected by increased expression of αSMA, reduced proliferation rates and enhanced ERK1/2 and SMAD2/3 phosphorylation. In PLK2 KO, the expression of the fibrotic cytokines osteopontin and IL18 was elevated compared to controls. Histological analysis of PLK2 KO lungs revealed early stage remodeling in terms of alveolar wall thickening, increased alveolar collagen deposition and myofibroblast foci. Our results prompt further investigation of PLK2 function in pulmonary fibrosis and suggest that the PLK2 KO model displays a genetic predisposition towards pulmonary fibrosis, which could be leveraged in future research on this topic.


Assuntos
Colágeno/metabolismo , Fibroblastos/enzimologia , Pulmão/enzimologia , Proteínas Serina-Treonina Quinases/deficiência , Fibrose Pulmonar/enzimologia , Adulto , Animais , Proliferação de Células , Células Cultivadas , Feminino , Fibroblastos/patologia , Deleção de Genes , Predisposição Genética para Doença , Humanos , Interleucina-18/genética , Interleucina-18/metabolismo , Pulmão/patologia , Masculino , Camundongos da Linhagem 129 , Camundongos Knockout , Pessoa de Meia-Idade , Miofibroblastos/enzimologia , Miofibroblastos/patologia , Osteopontina/genética , Osteopontina/metabolismo , Fenótipo , Proteínas Serina-Treonina Quinases/genética , Fibrose Pulmonar/genética , Fibrose Pulmonar/patologia , Transdução de Sinais
15.
Exp Cell Res ; 403(2): 112598, 2021 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-33865812

RESUMO

During orthodontic treatment, mechanical stretch serves a crucial function in osteogenic differentiation of periodontal ligament stem cells (PDLSCs). Up-regulated reactive oxygen species (ROS) level is a result of cyclic mechanical stretch in many cell types. Nuclear factor erythroid-2-related factor-2 (Nrf2) is a master regulator in various antioxidants expression. However, it is not known whether cyclic mechanical stretch could induce the ROS generation in PDLSCs and whether Nrf2 participated in this process. The present study was aimed to investigate the role of Nrf2 in PDLSCs under cyclic mechanical stretch. Our results showed that cyclic mechanical stretch increased ROS level and the nuclear accumulation of Nrf2 during osteoblast differentiation. Knocking down Nrf2 by siRNA transfection increased ROS formation and suppressed osteogenic differentiation in PDLSCs. T-BHQ, a Nrf2 activator, promoted the osteogenic differentiation in PDLSCs under cyclic mechanical stretch, and improved the microstructure of alveolar bone during orthodontic tooth movement in rats by employing micro-CT system. Taken together, Nrf2 activation was involved in osteogenic differentiation under cyclic mechanical stretch in PDLSCs. T-BHQ could promote the osteogenic differentiation in vitro and in vivo, suggesting a promising option for the remodeling of the alveolar bone during orthodontic tooth movement.


Assuntos
Fator 2 Relacionado a NF-E2/genética , Osteoblastos/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Células-Tronco/metabolismo , Adolescente , Animais , Diferenciação Celular , Sobrevivência Celular/efeitos dos fármacos , Criança , Colágeno Tipo I/genética , Colágeno Tipo I/metabolismo , Subunidade alfa 1 de Fator de Ligação ao Core/genética , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Feminino , Regulação da Expressão Gênica , Heme Oxigenase-1/genética , Heme Oxigenase-1/metabolismo , Humanos , Hidroquinonas/farmacologia , Masculino , Mecanotransdução Celular , NAD(P)H Desidrogenase (Quinona)/genética , NAD(P)H Desidrogenase (Quinona)/metabolismo , Fator 2 Relacionado a NF-E2/antagonistas & inibidores , Fator 2 Relacionado a NF-E2/metabolismo , Osteoblastos/citologia , Osteoblastos/efeitos dos fármacos , Osteogênese/genética , Osteopontina/genética , Osteopontina/metabolismo , Ligamento Periodontal/citologia , Ligamento Periodontal/efeitos dos fármacos , Ligamento Periodontal/metabolismo , Cultura Primária de Células , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Ratos , Ratos Wistar , Espécies Reativas de Oxigênio/antagonistas & inibidores , Células-Tronco/citologia , Células-Tronco/efeitos dos fármacos , Estresse Mecânico , Técnicas de Movimentação Dentária/métodos
16.
Indian J Pathol Microbiol ; 64(2): 339-342, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33851630

RESUMO

Background: Soft tissue keratocysts (SKC) are extremely rare and show similar microscopic morphology to keratocystic odontogenic tumor. The aim was to investigate immunohistochemical (IHC) features and origin of SKCs developing in buccal mucosa and lateral facial deep region. Material and Methods: Expression of CK19, CK10/13, Ki67, Cyclin D1 and Osteopontin (OPN) of 9 SKCS were investigated using IHC. Forty different types of cysts in jaw/soft tissue were used as control. Follow-up was performed. Results: CK10/13 positivity occurred more frequently and intensely in SKC and intraosseous parakeratinized odontogenic keratocysts (COKC). However, OPN positivity was observed only in COKC. Conclusion: This is the largest case series of SKCs; along with first attempt to investigate the expression of OPN on SKC. Given the microscopic and immunohistochemical features, we prefer the view that SKC is odontogenic origin but represents the soft tissue counterpart of COKC, since their expressions of OPN were extremely different.


Assuntos
Mucosa Bucal/patologia , Cistos Odontogênicos/diagnóstico , Cistos Odontogênicos/patologia , Tumores Odontogênicos/patologia , Adulto , Idoso , Ciclina D1/metabolismo , Feminino , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Osteopontina/metabolismo
17.
Nat Biomed Eng ; 5(8): 914-925, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-33782572

RESUMO

Cancer recurrence can arise owing to rare circulating cancer stem cells (CSCs) that are resistant to chemotherapies and radiotherapies. Here, we show that a double-network hydrogel can rapidly reprogramme differentiated cancer cells into CSCs. Spheroids expressing elevated levels of the stemness genes Sox2, Oct3/4 and Nanog formed within 24 h of seeding the gel with cells from any of six human cancer cell lines or with brain cancer cells resected from patients with glioblastoma. Human brain cancer cells cultured on the double-network hydrogel and intracranially injected in immunodeficient mice led to higher tumorigenicity than brain cancer cells cultured on single-network gels. We also show that the double-network gel induced the phosphorylation of tyrosine kinases, that gel-induced CSCs from primary brain cancer cells were eradicated by an inhibitor of the platelet-derived growth factor receptor, and that calcium channel receptors and the protein osteopontin were essential for the regulation of gel-mediated induction of stemness in brain cancer cells.


Assuntos
Reprogramação Celular , Hidrogéis/química , Células-Tronco Neoplásicas/citologia , Animais , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Diferenciação Celular , Receptores ErbB/metabolismo , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Glioblastoma/metabolismo , Glioblastoma/patologia , Humanos , Hidrogéis/farmacologia , Camundongos , Camundongos SCID , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/transplante , Osteopontina/genética , Osteopontina/metabolismo , Fosforilação/efeitos dos fármacos , Polímeros/química , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Células Tumorais Cultivadas
18.
J Biol Chem ; 296: 100436, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33610546

RESUMO

While details remain unclear, initiation of woven bone mineralization is believed to be mediated by collagen and potentially nucleated by bone sialoprotein (BSP). Interestingly, our recent publication showed that BSP and type XI collagen form complexes in mineralizing osteoblastic cultures. To learn more, we examined the protein composition of extracellular sites of de novo hydroxyapatite deposition which were enriched in BSP and Col11a1 containing an alternatively spliced "6b" exonal sequence. An alternate splice variant "6a" sequence was not similarly co-localized. BSP and Col11a1 co-purify upon ion-exchange chromatography or immunoprecipitation. Binding of the Col11a1 "6b" exonal sequence to bone sialoprotein was demonstrated with overlapping peptides. Peptide 3, containing three unique lysine-triplet sequences, displayed the greatest binding to osteoblastic cultures; peptides containing fewer lysine triplet motifs or derived from the "6a" exon yielded dramatically lower binding. Similar results were obtained with 6-carboxyfluorescein (FAM)-conjugated peptides and western blots containing extracts from osteoblastic cultures. Mass spectroscopic mapping demonstrated that FAM-peptide 3 bound to 90 kDa BSP and its 18 to 60 kDa fragments, as well as to 110 kDa nucleolin. In osteoblastic cultures, FAM-peptide 3 localized to biomineralization foci (site of BSP) and to nucleoli (site of nucleolin). In bone sections, biotin-labeled peptide 3 bound to sites of new bone formation which were co-labeled with anti-BSP antibodies. These results establish the fluorescent peptide 3 conjugate as the first nonantibody-based method to identify BSP on western blots and in/on cells. Further examination of the "6b" splice variant interactions will likely reveal new insights into bone mineralization during development.


Assuntos
Calcificação Fisiológica/fisiologia , Colágeno Tipo XI/metabolismo , Osteopontina/metabolismo , Animais , Osso e Ossos/metabolismo , Calcificação Fisiológica/genética , Colágeno/metabolismo , Colágeno Tipo XI/genética , Fluoresceínas/química , Sialoproteína de Ligação à Integrina/metabolismo , Masculino , Osteoblastos/metabolismo , Osteopontina/genética , Peptídeos/metabolismo , Fosfoproteínas/metabolismo , Proteínas de Ligação a RNA/metabolismo , Ratos , Sialoglicoproteínas/metabolismo
19.
Biol Chem ; 402(2): 195-206, 2021 01 27.
Artigo em Inglês | MEDLINE | ID: mdl-33544472

RESUMO

Glioblastoma multiforme (GBM) is the most aggressive type of brain cancer with a median survival of only 15 months. To complement standard treatments including surgery, radiation and chemotherapy, it is essential to understand the contribution of the GBM tumor microenvironment. Brain macrophages and microglia particularly contribute to tumor angiogenesis, a major hallmark of GBM. ADAM8, a metalloprotease-disintegrin strongly expressed in tumor cells and associated immune cells of GBMs, is related to angiogenesis and correlates with poor clinical prognosis. However, the specific contribution of ADAM8 to GBM tumorigenesis remains elusive. Knockdown of ADAM8 in U87 glioma cells led to significantly decreased angiogenesis and tumor volumes of these cells after stereotactic injection into striate body of mice. We found that the angiogenic potential of ADAM8 in GBM cells and in primary macrophages is mediated by the regulation of osteopontin (OPN), an important inducer of tumor angiogenesis. By in vitro cell signaling analyses, we demonstrate that ADAM8 regulates OPN via JAK/STAT3 pathway in U87 cells and in primary macrophages. As ADAM8 is a dispensable protease for physiological homeostasis, we conclude that ADAM8 could be a tractable target to modulate angiogenesis in GBM with minor side-effects.


Assuntos
Proteínas ADAM/metabolismo , Neoplasias Encefálicas/metabolismo , Glioblastoma/metabolismo , Proteínas de Membrana/metabolismo , Neovascularização Patológica/metabolismo , Osteopontina/metabolismo , Proteínas ADAM/deficiência , Proteínas ADAM/genética , Animais , Neoplasias Encefálicas/patologia , Proliferação de Células , Células Cultivadas , Glioblastoma/patologia , Humanos , Proteínas de Membrana/deficiência , Proteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neoplasias Experimentais/metabolismo , Neoplasias Experimentais/patologia , Neovascularização Patológica/patologia
20.
FASEB J ; 35(3): e21405, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33559950

RESUMO

Microglia activation and release of pro-inflammatory cytokines have been closely linked to glaucoma. However, the mechanisms that initiate these pathways remain unclear. Here, we investigated the role of a pro-inflammatory cytokine--osteopontin (OPN), in retinal microglia activation process along with the underlying mechanisms in glaucoma. A rat chronic ocular hypertension (COH) model was established presenting an increase in retinal OPN level and activation of microglia. Primary microglia cells were isolated and cultured under a pressure culture system showing heightened expressions of microglia-derived OPN with changes in inflammatory factors (TNF-α, IL-1ß, and IL-6). OPN and OPN neutralizing antibody (Anti-OPN) interventions were both applied systems for comparison, and cross-referenced with OPN knockdown in vitro. JAK/STAT, NF-κB, ERK1/2, and p38 MAPK, recognized as the primary signaling pathways related to microglia activation, were then screened on whether they can facilitate OPN to act on microglia and their impact on specific inhibitors. Thereafter, retrograde labeling of retinal ganglion cells (RGCs) and flash visual evoked potentials (F-VEP) were used to investigate neuron protection in context of each blockade. Results suggest that OPN is able to enhance the proliferation and activation of retinal microglia in experimental glaucoma which may play a role in the glaucomatous optic neuropathy, and contribute to the eventual RGCs loss and vision function impairment. Such effect may be mediated through the regulation of p38 MAPK signaling pathway.


Assuntos
Glaucoma/tratamento farmacológico , Hipertensão Ocular/etiologia , Osteopontina/farmacologia , Células Ganglionares da Retina/efeitos dos fármacos , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Animais , Citocinas/metabolismo , Modelos Animais de Doenças , Potenciais Evocados Visuais/efeitos dos fármacos , Potenciais Evocados Visuais/fisiologia , Glaucoma/metabolismo , Masculino , Microglia/efeitos dos fármacos , Microglia/metabolismo , Hipertensão Ocular/metabolismo , Osteopontina/metabolismo , Ratos Sprague-Dawley , Retina/metabolismo , Células Ganglionares da Retina/metabolismo , Fator de Necrose Tumoral alfa/metabolismo
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