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1.
Medicine (Baltimore) ; 99(26): e20635, 2020 Jun 26.
Artigo em Inglês | MEDLINE | ID: mdl-32590738

RESUMO

BACKGROUND: Gastric cancer (GC) is the most prevailing digestive tract malignant tumor worldwide with high mortality and recurrence rates. However, its potential molecular mechanism and prognostic biomarkers are still not fully understood. We aim to screen novel prognostic biomarkers related to GC prognosis using comprehensive bioinformatic tools. METHODS: Four gene expression microarray data were downloaded from the Gene Expression Omnibus (GEO) database (GSE26942, GSE33335, GSE63089, and GSE79973). Differentially expressed genes (DEGs) between gastric carcinoma and normal gastric tissue samples were identified by an integrated bioinformatic analysis. Gene Ontology (GO) term enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were performed using statistical software R. STRING and Cytoscape software were employed to construct protein-protein interaction (PPI) networks. Hub genes with a high score of connectivity identified from the PPI network were identified. Prognostic values of hub genes were evaluated in GSE15459 dataset. Hub genes related to GC overall survival were further validated in GEPIA (Gene Expression Profiling Interactive Analysis) online tool. RESULTS: A total of 12 upregulated DEGs and 59 downregulated DEGs were identified when the 4 microarray data overlapped. Among them, 10 hub genes with a high score of connectivity were identified. High expression of ghrelin and obestatin prepropeptide (GHRL), BGN, TIMP metallopeptidase inhibitor 1, thrombospondin 2, secreted phosphoprotein 1, and low expression of CHGA were associated with a poor overall survival of gastric cancer (all log rank P < .05). After validation in GEPIA database, only GHRL was confirmed associated with a poor overall survival of gastric cancer (log rank P = .04). CONCLUSIONS: GHRL could be used as a novel biomarker for the prediction of a poor overall survival of gastric cancer, and could be a novel therapeutic target for gastric cancer treatment. However, future experimental studies are still required to validate these findings.


Assuntos
Grelina/metabolismo , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/mortalidade , Biomarcadores Tumorais/metabolismo , Estudos de Casos e Controles , Cromogranina A/metabolismo , Expressão Gênica , Humanos , Análise em Microsséries , Osteopontina/metabolismo , Prognóstico , Trombospondinas/metabolismo , Inibidor Tecidual de Metaloproteinase-1/metabolismo
2.
Cardiovasc Ther ; 2020: 6869856, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32042311

RESUMO

Objectives: To observe the effect of avß3 single-stranded (ss) DNA on proliferation and migration of vascular smooth muscle cells (VSMCs) and its potential mechanism. Background: Percutaneous transluminal coronary angioplasty (PTCA) is currently the preferred method for the treatment of coronary heart disease. However, vascular restenosis still occurs after PTCA treatment, severely affecting the clinical efficacy of PTCA. Integrin avß3 single-stranded (ss) DNA on proliferation and migration of vascular smooth muscle cells (VSMCs) and its potential mechanism. Methods: In this experiment, we used systematic evolution of ligands by exponential enrichment (SELEX) to screen out avß3 single-stranded (ss) DNA on proliferation and migration of vascular smooth muscle cells (VSMCs) and its potential mechanism. ß3 single-stranded (ss) DNA on proliferation and migration of vascular smooth muscle cells (VSMCs) and its potential mechanism. ß3 single-stranded (ss) DNA on proliferation and migration of vascular smooth muscle cells (VSMCs) and its potential mechanism. ß3 single-stranded (ss) DNA on proliferation and migration of vascular smooth muscle cells (VSMCs) and its potential mechanism. Results: In the present study, we found that avß3 single-stranded (ss) DNA on proliferation and migration of vascular smooth muscle cells (VSMCs) and its potential mechanism. P < 0.05). Avß3 single-stranded (ss) DNA on proliferation and migration of vascular smooth muscle cells (VSMCs) and its potential mechanism. P < 0.05). AvP < 0.05). Av. Conclusions: The findings suggest that avß3 ssDNA inhibited the proliferation and migration of VSMCs by suppressing the activation of Ras-PI3K/MAPK signaling.ß3 single-stranded (ss) DNA on proliferation and migration of vascular smooth muscle cells (VSMCs) and its potential mechanism.


Assuntos
Aptâmeros de Nucleotídeos/metabolismo , Movimento Celular , Proliferação de Células , DNA de Cadeia Simples/metabolismo , Integrina alfaVbeta3/metabolismo , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Músculo Liso Vascular/enzimologia , Miócitos de Músculo Liso/enzimologia , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas ras/metabolismo , Animais , Apoptose , Proteínas Reguladoras de Apoptose/genética , Proteínas Reguladoras de Apoptose/metabolismo , Aptâmeros de Nucleotídeos/genética , Células Cultivadas , DNA de Cadeia Simples/genética , Quinase 1 de Adesão Focal/genética , Quinase 1 de Adesão Focal/metabolismo , Regulação da Expressão Gênica , Integrina alfaVbeta3/genética , Proteínas Quinases Ativadas por Mitógeno/genética , Músculo Liso Vascular/patologia , Miócitos de Músculo Liso/patologia , Osteopontina/genética , Osteopontina/metabolismo , Fosfatidilinositol 3-Quinases/genética , Fosforilação , Ratos Sprague-Dawley , Transdução de Sinais , Proteínas ras/genética
3.
Sci Rep ; 10(1): 1451, 2020 01 29.
Artigo em Inglês | MEDLINE | ID: mdl-31996744

RESUMO

Breast cancer patients treated with tamoxifen may experience recurrence due to endocrine resistance, which highlights the need for additional predictive and prognostic biomarkers. The glyco-phosphoprotein osteopontin (OPN), encoded by the SPP1 gene, has previously shown to be associated with poor prognosis in breast cancer. However, studies on the predictive value of OPN are inconclusive. In the present study, we evaluated tissue SPP1 mRNA and OPN protein expression as markers of recurrence in estrogen receptor- positive (ER+) breast cancer tissue. Tamoxifen- treated patients with recurrence or non-recurrence were selected using a matched case-control design. SPP1 mRNA expression was analysed using qPCR (n = 100) and OPN protein by immunohistochemistry (n = 116) using different antibodies. Odds ratios were estimated with conditional logistic regression. The SPP1 expression increased the risk of recurrence with an odds ratio (OR) of 2.50 (95% confidence interval [CI]; 1.30-4.82), after adjustment for tumour grade, HER 2 status and other treatments to OR 3.62 (95% CI; 1.45-9.07). However, OPN protein expression was not associated with risk of recurrence or with SPP1-gene expression, suggesting SPP1 mRNA a stronger prognostic marker candidate compared to tumor tissue OPN protein.


Assuntos
Adenocarcinoma/tratamento farmacológico , Antineoplásicos/uso terapêutico , Biomarcadores Tumorais/metabolismo , Neoplasias da Mama/tratamento farmacológico , Osteopontina/metabolismo , Tamoxifeno/uso terapêutico , Adenocarcinoma/diagnóstico , Adulto , Idoso , Idoso de 80 Anos ou mais , Neoplasias da Mama/diagnóstico , Estudos de Casos e Controles , Resistencia a Medicamentos Antineoplásicos , Feminino , Humanos , Pessoa de Meia-Idade , Osteopontina/genética , Prognóstico , Receptores Estrogênicos/metabolismo , Recidiva
4.
Nat Commun ; 11(1): 84, 2020 01 03.
Artigo em Inglês | MEDLINE | ID: mdl-31901081

RESUMO

Areas of a junction between two types of epithelia are known to be cancer-prone in many organ systems. However, mechanisms for preferential malignant transformation at the junction areas remain insufficiently elucidated. Here we report that inactivation of tumor suppressor genes Trp53 and Rb1 in the gastric squamous-columnar junction (SCJ) epithelium results in preferential formation of metastatic poorly differentiated neoplasms, which are similar to human gastroesophageal carcinoma. Unlike transformation-resistant antral cells, SCJ cells contain a highly proliferative pool of immature Lgr5-CD44+ cells, which are prone to transformation in organoid assays, comprise early dysplastic lesions, and constitute up to 30% of all neoplastic cells. CD44 ligand osteopontin (OPN) is preferentially expressed in and promotes organoid formation ability and transformation of the SCJ glandular epithelium. OPN and CD44 overexpression correlate with the worst prognosis of human gastroesophageal carcinoma. Thus, detection and selective targeting of the active OPN-CD44 pathway may have direct clinical relevance.


Assuntos
Carcinoma de Células Escamosas/metabolismo , Junção Esofagogástrica/metabolismo , Receptores de Hialuronatos/metabolismo , Osteopontina/metabolismo , Receptores Acoplados a Proteínas-G/metabolismo , Neoplasias Gástricas/metabolismo , Idoso , Idoso de 80 Anos ou mais , Animais , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/mortalidade , Carcinoma de Células Escamosas/patologia , Transformação Celular Neoplásica , Estudos de Coortes , Junção Esofagogástrica/patologia , Feminino , Humanos , Receptores de Hialuronatos/genética , Masculino , Camundongos , Camundongos Knockout , Pessoa de Meia-Idade , Osteopontina/genética , Receptores Acoplados a Proteínas-G/genética , Proteína do Retinoblastoma/genética , Proteína do Retinoblastoma/metabolismo , Neoplasias Gástricas/genética , Neoplasias Gástricas/mortalidade , Neoplasias Gástricas/patologia , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo
5.
FASEB J ; 34(1): 1122-1135, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31914633

RESUMO

Osteopontin (OPN) is a phosphoglycoprotein secreted into the extracellular matrix upon liver injury, acting as a cytokine stimulates the deposition of fibrillary collagen in liver fibrogenesis. In livers of mice subjected to bile duct ligation (BDL) and in cultured activated hepatic stellate cells (HSCs), we show that OPN, besides being overexpressed, is substantially phosphorylated by family with sequence similarity 20, member C (Fam20C), formerly known as Golgi casein kinase (G-CK), which is exclusively resident in the Golgi apparatus. In both experimental models, Fam20C becomes overactive when associated with a 500-kDa multiprotein complex, as compared with the negligible activity in livers of sham-operated rats and in quiescent HSCs. Fam20C knockdown not only confirmed the role of Fam20C itself in OPN phosphorylation, but also revealed that phosphorylation was essential for OPN secretion. However, OPN acts as a fibrogenic factor independently of its phosphorylation state, as demonstrated by the increased expression of Collagen-I by HSCs incubated with either a phosphorylated or nonphosphorylated form of recombinant OPN. Collectively, our results confirm that OPN promotes liver fibrosis and highlight Fam20C as a novel factor driving this process by favoring OPN secretion from HSCs, opening new avenues for deciphering yet unidentified mechanisms underlying liver fibrogenesis.


Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Proteínas da Matriz Extracelular/metabolismo , Células Estreladas do Fígado/metabolismo , Fígado/patologia , Osteopontina/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Animais , Citocinas/metabolismo , Fígado/metabolismo , Cirrose Hepática/metabolismo , Masculino , Camundongos , Camundongos Knockout , Fosforilação , Ratos , Ratos Wistar , Transdução de Sinais
6.
Nat Commun ; 11(1): 63, 2020 01 02.
Artigo em Inglês | MEDLINE | ID: mdl-31896743

RESUMO

Each vestibular sensory epithelium in the inner ear is divided morphologically and physiologically into two zones, called the striola and extrastriola in otolith organ maculae, and the central and peripheral zones in semicircular canal cristae. We found that formation of striolar/central zones during embryogenesis requires Cytochrome P450 26b1 (Cyp26b1)-mediated degradation of retinoic acid (RA). In Cyp26b1 conditional knockout mice, formation of striolar/central zones is compromised, such that they resemble extrastriolar/peripheral zones in multiple features. Mutants have deficient vestibular evoked potential (VsEP) responses to jerk stimuli, head tremor and deficits in balance beam tests that are consistent with abnormal vestibular input, but normal vestibulo-ocular reflexes and apparently normal motor performance during swimming. Thus, degradation of RA during embryogenesis is required for formation of highly specialized regions of the vestibular sensory epithelia with specific functions in detecting head motions.


Assuntos
Membrana dos Otólitos/embriologia , Ácido Retinoico 4 Hidroxilase/metabolismo , Tretinoína/metabolismo , Animais , Potenciais Evocados/genética , Potenciais Evocados/fisiologia , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Cabeça/fisiopatologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Osteopontina/metabolismo , Membrana dos Otólitos/citologia , Membrana dos Otólitos/metabolismo , Retinal Desidrogenase/genética , Retinal Desidrogenase/metabolismo , Ácido Retinoico 4 Hidroxilase/genética , Sáculo e Utrículo/citologia , Sáculo e Utrículo/embriologia , Tremor/genética , Tremor/fisiopatologia , Testes de Função Vestibular , Vestíbulo do Labirinto/embriologia , Vestíbulo do Labirinto/metabolismo
7.
Cell Prolif ; 53(2): e12758, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31922317

RESUMO

OBJECTIVE: The aim of this study is to investigate the role and potential mechanism of p75NTR in mineralization in vivo using p75NTR-knockout mice and in vitro using ectomesenchymal stem cells (EMSCs). MATERIALS AND METHODS: Femur bone mass and daily incisor mineralization speed were assessed in an in vivo p75NTR-knockout mouse model. The molecular signatures alkaline phosphatase (ALP), collagen type 1 (Col1), melanoma-associated antigen (Mage)-D1, bone sialoprotein (BSP), osteocalcin (OCN), osteopontin (OPN), distal-less homeobox 1 (Dlx1) and Msh homeobox 1 (Msx1) were examined in vitro in EMSCs isolated from p75NTR+/+ and p75NTRExIII-/- mice. RESULTS: p75NTR-knockout mice were smaller in body size than heterozygous and wild-type mice. Micro-computed tomography and structural quantification showed that the osteogenic ability of p75NTRExIII -knockout mice was significantly decreased compared with that of wild-type mice (P < .05). Weaker ALP and alizarin red staining and reduced expression of ALP, Col1, Runx2, BSP, OCN and OPN were also observed in p75NTRExIII-/- EMSCs. Moreover, the distance between calcein fluorescence bands in p75NTRExIII -knockout mice was significantly smaller than that in wild type and heterozygous mice (P < .05), indicating the lower daily mineralization speed of incisors in p75NTRExIII -knockout mice. Further investigation revealed a positive correlation between p75NTR and Mage-D1, Dlx1, and Msx1. CONCLUSION: p75NTR not only promotes osteogenic differentiation and tissue mineralization, but also shows a possible relationship with the circadian rhythm of dental hard tissue formation.


Assuntos
Células-Tronco Mesenquimais/metabolismo , Receptores de Fator de Crescimento Neural/metabolismo , Fosfatase Alcalina/metabolismo , Animais , Calcificação Fisiológica/fisiologia , Diferenciação Celular/fisiologia , Células Cultivadas , Colágeno Tipo I/metabolismo , Feminino , Sialoproteína de Ligação à Integrina/metabolismo , Masculino , Camundongos , Camundongos Knockout , Osteocalcina/metabolismo , Osteogênese/fisiologia , Osteopontina/metabolismo
8.
Am J Physiol Cell Physiol ; 318(1): C73-C82, 2020 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-31577514

RESUMO

Our objective was to investigate the role of primary cilia in low-magnitude, high-frequency vibration (LMHFV) treatment of MC3T3-E1 osteoblasts (OBs). We used chloral hydrate (CH), which has a well-characterized function in chemically removing primary cilia, to elucidate the role of primary cilia in LMHFV-induced OB osteogenic responses through cell viability assay, Western blot analysis, real-time quantitative RT-PCR, and histochemical staining methods. We observed a significant, 30% decrease in the number of MC3T3-E1 OBs with primary cilia (reduced from 64.3 ± 5%) and an approximately 50% reduction in length of primary cilia (reduced from 3 ± 0.8 µm) after LMHFV stimulation. LMHFV stimulation upregulated protein expression of the bone matrix markers collagen 1 (COL-1), osteopontin (OPN), and osteoclacin(OCN) in MC3T3-E1 OBs, indicating that LMHFV induces osteogenesis. High-concentration or long-duration CH exposure resulted in inhibition of MC3T3-E1 OB survival. In addition, Western blot analysis and RT-PCR revealed that CH treatment prevented LMHFV-induced osteogenesis. Furthermore, decreased alkaline phosphate activity, reduced OB differentiation, mineralization, and maturation were observed in CH-pretreated and LMHFV-treated OBs. We showed that LMHFV induces morphological changes in primary cilia that may fine-tune their mechanosensitivity. In addition, we demonstrated the significant inhibition by CH of LMHFV-induced OB mineralization, maturation, and differentiation, which might reveal the critical role of primary cilia in the process.


Assuntos
Diferenciação Celular , Cílios/metabolismo , Mecanotransdução Celular , Osteoblastos/metabolismo , Osteogênese , Vibração , Células 3T3 , Animais , Diferenciação Celular/genética , Hidrato de Cloral/toxicidade , Cílios/efeitos dos fármacos , Cílios/patologia , 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 , Ciclo-Oxigenase 2/genética , Ciclo-Oxigenase 2/metabolismo , Regulação da Expressão Gênica , Camundongos , Osteoblastos/efeitos dos fármacos , Osteoblastos/patologia , Osteocalcina/metabolismo , Osteogênese/genética , Osteopontina/metabolismo , Fatores de Tempo
9.
J Periodontal Res ; 55(1): 141-151, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31539178

RESUMO

BACKGROUND AND OBJECTIVES: Strontium ranelate is a medication indicated for the treatment of osteoporosis that presents concomitant anti-resorptive and osteoanabolic dual biological activity. However, the effects of strontium ranelate on alveolar bone have been poorly explored. Furthermore, to date, there are no data on the effects of this medication on alveolar bone loss (BL) during conditions of estrogen deficiency. Therefore, the aim of this study was to evaluate the effects of strontium ranelate on ligature-induced periodontitis in estrogen-deficient and estrogen-sufficient rats. METHODS: Ninety-six rats were assigned to one of the following groups: sham-surgery + water (estrogen-sufficient; n = 24); ovariectomy + water (estrogen-deficient; n = 24), sham-surgery + strontium ranelate (ranelate/estrogen-sufficient; n = 24) and; ovariectomy + strontium ranelate (ranelate/estrogen-deficient; n = 24). The rats received strontium ranelate or water from the 14th day after ovariectomy until the end of the experiment. On the 21st day after ovariectomy, one first mandibular molar received a ligature, while the contralateral tooth was left unligated. Eight rats per group were killed at 10, 20, and 30 days after ligature placement. Bone loss (BL) and trabecular bone area (TBA) were analyzed in the furcation area of ligated and unligated teeth at all experimental times by histometry. Tartrate-resistant acid phosphatase (TRAP) positive cells and immunohistochemical staining for osteocalcin (OCN), osteopontin (OPN), osteoprotegerin (OPG), and receptor activator of NF-КB ligand (RANKL) were assessed in the ligated teeth at 30 days after ligature placement. RESULTS: At 10 and 30 days, ligated teeth of the estrogen-deficient group exhibited higher BL, when compared to all other groups (P < .05). At 10 days, TBAs were higher in the unligated teeth of strontium ranelate-treated groups, when compared to those of untreated groups (P < .05). At 30 days, the ligated teeth of the estrogen-deficient group exhibited lower TBA than the other groups (P < .05). There were no differences among groups regarding the number of TRAP-stained cells (P < .05). The strontium ranelate-treated groups exhibited lower expressions of OCN and RANKL than the untreated groups (P < .05). The estrogen-sufficient group presented higher staining for OPG than both treated and untreated estrogen-deficient groups (P < .05). CONCLUSIONS: Strontium ranelate prevented ligature-induced BL in an estrogen-deficiency condition and, to a certain extent, increased TBA in the presence and absence of periodontal collapse in states of estrogen deficiency and estrogen sufficiency. Furthermore, strontium ranelate also affected the expression of bone markers, appearing to have acted predominantly as an anti-resorptive agent.


Assuntos
Perda do Osso Alveolar/tratamento farmacológico , Estrogênios/deficiência , Periodontite/tratamento farmacológico , Tiofenos/farmacologia , Animais , Osteocalcina/metabolismo , Osteopontina/metabolismo , Osteoprotegerina/metabolismo , Ovariectomia , Ligante RANK/metabolismo , Ratos , Ratos Wistar
10.
J Endod ; 46(1): 89-96, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31740066

RESUMO

INTRODUCTION: Although dentin matrix protein 1 (DMP1) and osteopontin (OPN) act as substrates and signaling molecules for odontoblastlike cell differentiation after tooth injury, the mutual interaction between these proteins in the mechanism of odontoblastlike cell differentiation remains to be clarified. This study aimed to elucidate the role of DMP1 and OPN in regulating odontoblastlike cell differentiation after tooth injury. METHODS: A groove-shaped cavity was prepared on the mesial surface of the upper first molars in wild-type and Opn knockout (KO) mice. The demineralized paraffin sections were processed for immunohistochemistry for nestin and DMP1 and in situ hybridization for Dmp1. For the in vitro assay, the experiments of organ culture for evaluating dentin-pulp complex regeneration using small interfering RNA treatment were performed. RESULTS: Once preexisting odontoblasts died, nestin-positive newly differentiated odontoblastlike cells were arranged along the pulp-dentin border and began to express DMP1/Dmp1. In Opn KO mice, the expression of DMP1/Dmp1 was up-regulated compared with that of wild-type mice. The in vitro assay showed that the gene suppression of Dmp1 by small interfering RNA showed a tendency to decrease the differentiation rate of odontoblastlike cells from 70.1% to 52.2% in wild-type teeth. In addition, the suppression of Dmp1 in Opn KO teeth tended to lead to the inhibition of odontoblastlike cell differentiation. CONCLUSIONS: These results suggest that the expression of Dmp1 is up-regulated in Opn KO mice both in vivo and in vitro, and DMP1 compensates for the lack of OPN in regulating odontoblastlike cell differentiation after tooth injury.


Assuntos
Diferenciação Celular , Proteínas da Matriz Extracelular , Osteopontina/metabolismo , Traumatismos Dentários , Animais , Dentina , Proteínas da Matriz Extracelular/metabolismo , Camundongos , Odontoblastos , Fosfoproteínas
11.
Mater Sci Eng C Mater Biol Appl ; 107: 110348, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31761176

RESUMO

The differentiation of adult stem cells is usually performed in vitro, by exposing them to specific factors. Alternatively, one can use nanocarriers containing such factors, to be internalized by the cells. In this work we have reduce the size of those carriers to the nanoscale, developing bioactive silica nanoparticles with diameters under 100 nm, containing calcium and phosphate ions (SiNPs-CaP). These ions, once released inside adult stem cells, induce bone cell proliferation and differentiation, and stimulate the expression of bone-related proteins in a single dose administration. The SiNPs-CaP nanomaterials were prepared through a sol-gel approach, and the ions added with a post-synthesis functionalization method. The synthesized SiNPs-CaP have narrow size distribution, good colloidal stability, and show high levels of ion incorporation. Furthermore, the SiNPs-CaP have good cytocompatibility and promote the osteogenic differentiation of human bone marrow mesenchymal stem cells (hBMSC), with alkaline phosphatase, osteopontin and osteocalcin production levels comparable to the ones obtained in standard osteogenic medium. The novel bioactive SiNPs-CaP are synthesized in a simple and fast manner and show the ability to promote osteogenic differentiation after a single dose administration, independently from external osteogenic inducers, showing great potential as carriers in bone tissue engineering applications.


Assuntos
Cálcio/administração & dosagem , Células-Tronco Mesenquimais/citologia , Nanopartículas/química , Osteogênese/efeitos dos fármacos , Fosfatos/administração & dosagem , Fosfatase Alcalina/metabolismo , Cálcio/química , Cálcio/farmacocinética , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Liberação Controlada de Fármacos , Humanos , Células-Tronco Mesenquimais/efeitos dos fármacos , Nanopartículas/administração & dosagem , Osteocalcina/metabolismo , Osteogênese/fisiologia , Osteopontina/metabolismo , Tamanho da Partícula , Fosfatos/química , Fosfatos/farmacocinética , Dióxido de Silício/química
12.
J Ethnopharmacol ; 248: 112329, 2020 Feb 10.
Artigo em Inglês | MEDLINE | ID: mdl-31672526

RESUMO

ETHNOPHARMACOLOGICAL RELEVANCE: Mesenchymal stem cells (MSCs) are multipotent stem cells possessing regenerative potential. Symphytum officinale (SO) is a medicinal plant and in homoeopathic literature, believed to accelerate bone healing. AIM OF THE STUDY: This study aimed to determine if homoeopathic doses of SO could augment osteogenesis in MSCs as they differentiate into osteoblasts in vitro. MATERIALS AND METHODS: Bone marrow samples were obtained from patients who underwent bone grafting procedures (n = 15). MSCs were isolated, expanded and characterized by flow cytometry (CD90, CD105). Cytotoxicity of SO was evaluated by MTT assay. Osteogenic differentiation was induced in MSCs with ß-glycerophosphate, ascorbic acid and dexamethasone over 2 weeks. Different homoeopathic doses of SO (MT, 3C, 6C, 12C and 30C) were added to the basic differentiation medium (BDM) and efficiency of MSCs differentiating into osteoblasts were measured by evaluating expression of Osteocalcin using flow cytometry, and alkaline phosphatase activity using ELISA. Gene expression analyses for osteoblast markers (Runx-2, Osteopontin and Osteocalcin) were evaluated in differentiated osteoblasts using qPCR. RESULTS: Flow cytometry (CD90, CD105) detected MSCs isolated from bone marrow (93-98%). MTT assay showed that the selected doses of SO did not induce any cytotoxicity in MSCs (24 hours). The efficiency of osteogenic differentiation (2 weeks) for different doses of Symphytum officinale was determined by flow cytometry (n = 10) for osteoblast marker, Osteocalcin, and most doses of Symphytum officinale enhanced osteogenesis. Interestingly, gene expression analysis for Runx-2 (n = 10), Osteopontin (n = 10), Osteocalcin (n = 10) and alkaline phosphatase activity (n = 8) also showed increased osteogenesis with the addition of Symphytum officinale to BDM, specially mother tincture. CONCLUSIONS: Our findings suggest that homoeopathic dose (specially mother tincture) of Symphytum officinale has the potential to enhance osteogenesis.


Assuntos
Conservadores da Densidade Óssea/farmacologia , Diferenciação Celular/efeitos dos fármacos , Confrei , Homeopatia , Células-Tronco Mesenquimais/efeitos dos fármacos , Osteoblastos/efeitos dos fármacos , Osteogênese/efeitos dos fármacos , Extratos Vegetais/farmacologia , Fosfatase Alcalina/metabolismo , Conservadores da Densidade Óssea/isolamento & purificação , Diferenciação Celular/genética , Linhagem Celular , Confrei/química , Subunidade alfa 1 de Fator de Ligação ao Core/genética , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Relação Dose-Resposta a Droga , Regulação da Expressão Gênica , Humanos , Células-Tronco Mesenquimais/metabolismo , Osteoblastos/metabolismo , Osteocalcina/genética , Osteocalcina/metabolismo , Osteogênese/genética , Osteopontina/genética , Osteopontina/metabolismo , Fenótipo , Extratos Vegetais/isolamento & purificação
13.
Artif Cells Nanomed Biotechnol ; 48(1): 159-168, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31852298

RESUMO

Accumulating evidence links osteopontin (OPN), a pro-fibrogenic extracellular matrix protein, to the pathogenesis of non-alcoholic fatty liver disease (NAFLD). In this study, liver tissues isolated from non-alcoholic steatohepatitis (NASH) patients expressed higher OPN than those of controls. However, the exact mechanism(s) for this phenomenon is yet to be clarified. Autophagy is the natural, regulated degradation and recycling of a cell's dysfunctional components, in order to maintain homeostasis. Increasing evidence supports that autophagy can constitute an effective Defence mechanism against NAFLD conditions. Herein, we constructed NAFLD mice model by high-fat (HF) and methionine-choline-deficient (MCD) diet and found that OPN is upregulated in livers of NAFLD mice. Besides, secreted OPN inhibited autophagosome-lysosome fusion via binding with its receptors integrin αVß3 and αVß5 in HepG2 cells supplemented with free fatty acids (FFA) and the livers of NAFLD mice. Silencing of OPN attenuated autophagy impairment and reduced lipid accumulation, while supplementation of OPN exhibited the opposite effect. Furthermore, treatment with anti-OPN Ab significantly attenuated steatosis as well as autophagy impairment in the liver. Our findings indicated that OPN plays a vital role in the pathogenesis of the development of NAFLD via autophagy impairment, which might represent a potential new therapeutic target for the treatment of NAFLD.


Assuntos
Autofagia , Metabolismo dos Lipídeos , Hepatopatia Gordurosa não Alcoólica/metabolismo , Hepatopatia Gordurosa não Alcoólica/patologia , Osteopontina/metabolismo , Animais , Progressão da Doença , Regulação da Expressão Gênica , Humanos , Integrina alfaVbeta3/metabolismo , Fígado/metabolismo , Fígado/patologia , Masculino , Camundongos
14.
Eur J Pharmacol ; 866: 172822, 2020 Jan 05.
Artigo em Inglês | MEDLINE | ID: mdl-31760068

RESUMO

Brown adipose tissue (BAT) plays important roles in regulating energy homeostasis and combating obesity. Accordingly, increasing the abundance and/or activating BAT would be effective and promising approaches to combat obesity and obesity-relative diseases. Our previous data in vitro have shown that osteopontin (OPN) induces the brown adipogenesis in 3T3-L1 cells via a phosphatidylinositol 3 kinase (PI3K)-AKT pathway. However, it is currently unknown whether OPN exerts such an effect on animals in vivo. Therefore, in the study we sought to investigate the pro-browning effects of OPN and to explore its underlying mechanisms by transfecting with Ad-GFP-aP2-OPN-shRNA to specifically down-regulate the OPN of white adipose tissue (WAT) in mice. Our present results show that downregulation of OPN in WAT exacerbates obesity and inhibits WAT-browning. Moreover, immunohistochemical results also exhibit that the downregulation of OPN significantly diminishes the expression and sub-cellular localization of UCP-1, PRDM16 and PGC-1α. Besides, the western blotting results reveal that the expression levels of PI3K, AKT-pS473 and PPARγ markedly reduce. Consequently, we conclude that the downregulation of OPN inhibits the browning of WAT through inhibiting the expression of PPARγ mediated by the PI3K-AKT pathway. The findings suggest that OPN is involved in regulation of WAT-browning and regulating its expression would become a potential strategy to combat obesity and obesity-relative metabolic diseases.


Assuntos
Tecido Adiposo Marrom/citologia , Tecido Adiposo Branco/citologia , Regulação para Baixo , Osteopontina/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais , Células 3T3-L1 , Animais , Proteínas de Ligação a DNA/metabolismo , Células Endoteliais da Veia Umbilical Humana , Humanos , Fígado/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Fatores de Transcrição/metabolismo , Proteína Desacopladora 1/metabolismo
15.
Mater Sci Eng C Mater Biol Appl ; 107: 110228, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31761154

RESUMO

This study simulated the high-phosphorus (Pi) environment in patients with chronic kidney disease. Nano-hydroxyapatite (HAP) crystals were used to damage rat aortic smooth muscle cells (A7R5) pre-damaged with different concentrations of Pi solution to compare the differences in HAP-induced calcification in A7R5 cells before and after injury by high-Pi condition. After the A7R5 cells were damaged by high-Pi environment, the following were observed. HAP resulted in declined cell viability and lysosomal integrity, release of lactate dehydrogenase, and increased reactive oxygen species production. The ability of high-Pi damaged cells to internalize HAP crystals declined; crystal adhesion and calcium deposition on the cell surface and alkaline phosphatase activities increased. Osteopontin expression and level of Runt-related transcription factor 2 were increased, and HAP-induced osteogenic transformation was enhanced. High-Pi condition promoted the adhesion of A7R5 cells to nano-HAP crystals and inhibited HAP endocytosis, increasing the risk of calcification.


Assuntos
Cálcio/metabolismo , Diferenciação Celular/efeitos dos fármacos , Durapatita/química , Fósforo/metabolismo , Fosfatase Alcalina/metabolismo , Animais , Adesão Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Durapatita/metabolismo , Endocitose , Lisossomos/metabolismo , Músculo Liso Vascular/citologia , Músculo Liso Vascular/metabolismo , Osteopontina/metabolismo , Ratos , Espécies Reativas de Oxigênio/metabolismo
16.
J Ethnopharmacol ; 246: 112207, 2020 Jan 10.
Artigo em Inglês | MEDLINE | ID: mdl-31476440

RESUMO

ETHNOPHARMACOLOGICAL RELEVANCE: Liuwei Dihuang (LWDH) is a classic prescription that has been used as a traditional medicinal formula for more than 1000 years in China. In clinical, LWDF is used for treating functional decline associated with senile disease and menopausal syndrome. Studies have demonstrated that LWDH could significantly improve estrogen level and ER expression, and suspend the process of atherosclerosis. However, the under mechanism of how LWDH suppressing VSMCs phenotypic conversion and proliferation through ER is still unknown. AIM OF THE STUDY: This study was to reveal the under mechanism of how LWDH inhibits the phenotypic conversion of VSMCs. MATERIALS AND METHODS: 24 ApoE-/- mice were divided into 4 groups: sham group, model group, E2 group, and LWDH group, and 6 C57BN/L6 mice were used as control group. The primary VSMCs were divided into control group, model group, E2 group, LWDH group, LWDH + MPP group, and LWDH + PHTPP group with or without control siRNA, ERα siRNA, ERß siRNA, and myocardin siRNA. Oil red staining was used to evaluate the lipid deposition in the cardiac aorta. Serum chemistry analysis to test serum TG, TC, LDL, and HDL. Immunofluorescence staining was used to test α-SMA, osteopontin and F-actin. Immunohistochemical staining was performed to check out the myocardin in the cardiac aorta. The mRNA levels of α-SMA, osteopontin, ERα, ERß, SRC3 and myocardin were detected by Real Time-PCR, and the protein expression levels of them were detected by Western blotting. Co-immunoprecipitation was proceed to test the interaction between ERα and SRC3 and SRC3 and myocardin. Flow cytometry was used to check out the cell cycle. Wound healing assay and Transwell were managed to evaluate the migration capacity of VSMCs. RESULTS: In vivo administration of LWDH suppressed AS symptoms, decreases phenotypic marker of vascular endothelial cell, and increases phenotypic marker of VSMC in ovariectomized ApoE-/- female mice. Moreover, LWDH significantly increased the mRNA and protein expression levels of ERα, ERß, SRC3 and myocardin in the cardiac aorta of ovariectomized ApoE-/- female mice. In vitro, LWDH altered cell cycle and reduced the elevated cyclinD protein expression migration capacity and in the model VSMCs. In addition, LWDH inhibited phenotypic conversion and promoted the expression of ER, SRC3, and myocardin of the primary VSMC phenotypic conversion model. Inhibition of ERα almost completely eliminated the impacts of LWDH on α- SMA and osteopontin. Furthermore, LWDH promoted the interaction between ERα and SRC3 and up-regulated the co-activation of SRC3 and myocardin. CONCLUSIONS: LWDH could inhibit the phenotypic conversion of VSMCs in vitro and in vivo by increasing the activity of myocardin through up-regulating the expression of ERα and promoting the interaction between ERα and SRC3. Our research reveals the under mechanism of how LWDH inhibits the phenotypic conversion of VSMCs.


Assuntos
Aterosclerose/prevenção & controle , Medicamentos de Ervas Chinesas/farmacologia , Miócitos de Músculo Liso/efeitos dos fármacos , Proteínas Nucleares/metabolismo , Transativadores/metabolismo , Actinas/genética , Actinas/metabolismo , Animais , Aorta/metabolismo , Cápsulas , Células Cultivadas , Receptor alfa de Estrogênio/genética , Receptor alfa de Estrogênio/metabolismo , Receptor beta de Estrogênio/genética , Receptor beta de Estrogênio/metabolismo , Feminino , Menopausa/genética , Menopausa/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout para ApoE , Músculo Liso Vascular/citologia , Miócitos de Músculo Liso/metabolismo , Proteínas Nucleares/genética , Osteopontina/genética , Osteopontina/metabolismo , Fenótipo , Ratos Sprague-Dawley , Transativadores/genética , Regulação para Cima/efeitos dos fármacos
17.
J Dairy Sci ; 103(1): 42-51, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31733850

RESUMO

Osteopontin (OPN) is a multifunctional protein highly expressed in milk, where it is hypothesized to be involved in immunological signaling via the conserved Arg-Gly-Asp (RGD) integrin-binding sequence. Intervention studies have indicated beneficial effects of orally administered OPN in animal and human infants, but the mechanisms underlying these effects are not well described. To induce physiological effects, OPN must resist gastrointestinal transit in a bioactive form. In this study, we subjected bovine milk OPN to in vitro gastrointestinal transit, and characterized the generated fragments using monoclonal antibody and mass spectrometric analyses. We found that the fragment Trp27-Phe151 containing the integrin-binding RGD sequence resisted in vitro gastric digestion. This resistance was dependent on glycosylation of threonine residues near the integrin-binding sequence in both human and bovine milk OPN. Furthermore, the fragment Trp27-Phe151 retained the ability to interact with integrins in an RGD-dependent process. These results suggest a mechanism for how ingested milk OPN can induce physiological effects via integrin signaling in the intestine.


Assuntos
Reatores Biológicos , Bovinos/fisiologia , Trânsito Gastrointestinal , Integrinas/metabolismo , Leite/química , Osteopontina/farmacologia , Animais , Humanos , Integrinas/química , Osteopontina/química , Osteopontina/metabolismo , Ligação Proteica
18.
Mater Sci Eng C Mater Biol Appl ; 107: 110204, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31761242

RESUMO

Using a novel microwave-assisted hydrothermal (MW) process we created nano-scale anatase on micro-arc-oxidized (MAO) titanium surface. The morphology/crystallinity and surface potential of the anatase which altered by Ca/P ions concentrations and pH in the MW medium were characterized by atomic force microscopy, Kelvin probe force microscopy, Raman spectrometer, and x-ray photoelectron spectroscopy. The surface of MWDD (processed in DD water) and MWCP (in neutral pH medium with Ca/P ions) are covered with anatase spikes, which enhance their nano-roughness, hydrophilicity, and possess lower surface potential than other groups. The nano-precipitates on surface of MWCP9 and MWCP11 (processed in medium containing Ca/P ions at pH 9 or pH 11) were mainly amorphous anatase with less P ions. The protein adsorption of negatively charged bovine serum albumin was higher in MWDD and MWCP groups which possess lower surface potential. The adsorption of positively charged histones on the surface of MWCP11 was higher compared to the other groups. The osteogenic characterizations of D1 mice bone marrow mesenchymal stem cells co-cultured for 1, 7, and 14 days were measured by ALP and osteopontin assays. Although MW groups revealed comparable viability of D1 mice bone marrow mesenchymal stem cells to MAO group, their superior hydrophilicity and higher protein adsorption, thus regulating the differentiation of osteoprogenitor stem cells demonstrating higher ALP and osteopontin secretion after 7 days and 14 days. The nanoscale topography, crystallinity and surface potential change the hydrophilicity and protein adsorption on the MW treated titanium surface, thus regulating the differentiation of osteoprogenitor stem cells in vitro.


Assuntos
Histonas/química , Nanoestruturas/química , Titânio/química , Adsorção , Animais , Bovinos , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Interações Hidrofóbicas e Hidrofílicas , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/metabolismo , Camundongos , Micro-Ondas , Osteogênese/efeitos dos fármacos , Osteopontina/metabolismo , Oxirredução , Soroalbumina Bovina/química , Propriedades de Superfície , Titânio/farmacologia
19.
PLoS One ; 14(12): e0225598, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31805069

RESUMO

Methylsulfonylmethane (MSM) is a naturally occurring, sulfate-containing, organic compound. It has been shown to stimulate the differentiation of mesenchymal stem cells into osteoblast-like cells and bone formation. In this study, we investigated whether MSM influences the differentiation of stem cells from human exfoliated deciduous teeth (SHED) into osteoblast-like cells and their osteogenic potential. Here, we report that MSM induced osteogenic differentiation through the expression of osteogenic markers such as osterix, osteopontin, and RUNX2, at both mRNA and protein levels in SHED cells. An increase in the activity of alkaline phosphatase and mineralization confirmed the osteogenic potential of MSM. These MSM-induced effects were observed in cells grown in basal medium but not osteogenic medium. MSM induced transglutaminase-2 (TG2), which may be responsible for the cross-linking of extracellular matrix proteins (collagen or osteopontin), and the mineralization process. Inhibition of TG2 ensued a significant decrease in the differentiation of SHED cells and cross-linking of matrix proteins. A comparison of mineralization with the use of mineralized and demineralized bone particles in the presence of MSM revealed that mineralization is higher with mineralized bone particles than with demineralized bone particles. In conclusion, these results indicated that MSM could promote differentiation and osteogenic potential of SHED cells. This osteogenic property is more in the presence of mineralized bone particles. TG2 is a likely cue in the regulation of differentiation and mineral deposition of SHED cells in response to MSM.


Assuntos
Calcificação Fisiológica/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Dimetil Sulfóxido/farmacologia , Osteogênese/efeitos dos fármacos , Sulfonas/farmacologia , Biomarcadores/metabolismo , Células Cultivadas , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Proteínas de Ligação ao GTP/metabolismo , Humanos , Osteopontina/metabolismo , Fator de Transcrição Sp7/metabolismo , Células-Tronco , Transglutaminases/metabolismo
20.
PLoS One ; 14(12): e0226839, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31860680

RESUMO

Previous studies have suggested that treatment plans for segmental bone defects (SBDs) are affected by the bone defect sizes. If the selected treatment was not the most appropriate, it would not contribute to bone healing, but increase complications. The induced membrane technique (IM) and distraction osteogenesis (DO) have been proved to be effective in treating SBDs. However, the differences between the two in therapeutic effects on SBDs with different sizes are still unclear. Thus, we aimed to observe the effects of IM and DO on different sizes of SBDs and to further determine what method is more appropriate for what defect size. Rat models of 4-, 6-and 8-mm mid-diaphyseal defects using IM and DO techniques were established. X-rays, micro-CT, histological and immunohistochemical examinations were performed to assess bone repair. Faster bone formation rate, shorter treatment duration, higher expressions of OPN and OCN and higher parameters of bone properties including bone mineral density (BMD), bone volume/total tissue volume (BV/TV), mineral apposition rate (MAR) and mineral surface/bone surface (MS/BS) were found in 4-mm SBDs treated with DO than in those with IM treatment. However, the results were reversed and IM outperformed DO in bone repair capacity for 8-mm SBDs, while no significant difference emerges in the case of 6-mm SBDs. This study suggests that the therapeutic effects of IM and DO may be subjected to sizes of bone defects and the best treatment size of defects is different between the two. For small-sized SBDs, DO may be more suitable and efficient than IM, but IM has advantages over DO for over-sized SBDs, while DO and IM show similar bone repair capability in moderate-sized SBDs, which would offer a new insight into how to choose DO and IM for SBDs in clinical practice and provide references for further clinical research.


Assuntos
Modelos Animais de Doenças , Deformidades Congênitas das Extremidades Inferiores/cirurgia , Osteogênese por Distração/métodos , Animais , Densidade Óssea , Regeneração Óssea , Diáfises/cirurgia , Imuno-Histoquímica , Deformidades Congênitas das Extremidades Inferiores/diagnóstico por imagem , Masculino , Osteoblastos/metabolismo , Osteocalcina/imunologia , Osteocalcina/metabolismo , Osteogênese , Osteopontina/imunologia , Osteopontina/metabolismo , Ratos , Ratos Sprague-Dawley , Tíbia/diagnóstico por imagem , Tíbia/cirurgia , Microtomografia por Raio-X
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