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1.
Exp Appl Acarol ; 78(2): 273-293, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31168751

RESUMO

Tetranychus urticae (Acari: Tetranychidae) is an extremely serious cassava (Manihot esculenta) pest. Building a genomic resource to investigate the molecular mechanisms of cassava responses to T. urticae is vital for characterizing cassava resistance to mites. Based on the tolerance of cassava varieties to mite infestation (focusing on mite development rate, fecundity and physiology), cassava variety SC8 was selected to analyze transcriptomic and proteomic changes after 5 days of T. urticae feeding. Transcriptomic analysis revealed 698 and 2140 genes with significant expression changes under low and high mite infestation, respectively. More defense-related genes were found in the enrichment pathways at high mite density than at low density. In addition, iTRAQ-labeled proteomic analysis revealed 191 proteins with significant expression changes under low mite infestation. Differentially expressed genes and proteins were mainly found in the following defense-related pathways: flavonoid biosynthesis, phenylpropanoid biosynthesis, and glutathione metabolism under low-density mite feeding and plant hormone signal transduction and plant-pathogen interaction pathways under high-density mite feeding. The plant hormone signal transduction network, involving ethylene, jasmonic acid, and salicylic acid transduction pathways, was explored in relation to the M. esculenta response to T. urticae. Correlation analysis of the transcriptome and proteome generated a Pearson correlation coefficients of R = 0.2953 (P < 0.01), which might have been due to post-transcriptional or post-translational regulation resulting in many genes being inconsistently expressed at both the transcript and protein levels. In summary, the M. esculenta transcriptome and proteome changed in response to T. urticae, providing insight into the general activation of plant defense pathways in response to mite infestation.


Assuntos
Cadeia Alimentar , Manihot/fisiologia , Proteínas de Plantas/análise , Proteoma , Tetranychidae/fisiologia , Transcriptoma , Animais , Antibiose , Manihot/genética , Transdução de Sinais
2.
BMC Cancer ; 19(1): 454, 2019 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-31092229

RESUMO

BACKGROUND: Major vault protein (MVP) is the major component of vault, a eukaryotic organelle involved in multiple cellular processes, and is important in multiple cellular processes and diseases including the drug resistance in cancer chemotherapies. However, the role of MVP in lung cancer remains unclear. METHODS: We examined MVP expression in 120 non-small cell lung cancer (NSCLC) tumors and matched normal tissues by immunohistochemistry. Its relationship with NSCLC prognosis was determined by investigating the patient cohort and analyzing the data from a published dataset consisting with more than 1900 lung cancer patients. We further performed shRNA-introduced knockdown of MVP in Lewis lung carcinoma (LLC) cells and examined its effects on the tumor formation in a xenograft mouse model and the tumor cell proliferation, apoptosis, and signal transduction in vitro. RESULTS: We found that MVP was up-regulated significantly in tumor tissues compared with the matched tumor-adjacent normal tissues. The increased expression of MVP in lung adenocarcinoma was associated with a better prognosis. Knockdown of MVP in LLC cells promoted xenografted lung cancer formation in mice, which was accompanied with accelerated tumor cell proliferation and suppressed cell apoptosis in vitro. Knockdown of MVP stimulated STAT3 phosphorylation, nuclear localization, and activation of JAK2 and RAF/MEK/ERK pathways in LLC cells. Administration of STAT3 inhibitor WP1066 could prevent MVP knockdown induced tumorigenesis. CONCLUSIONS: Our findings demonstrate that MVP may act as a lung tumor suppressor via inhibiting STAT3 pathway. MVP would be a potential target for novel therapies of lung adenocarcinoma.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/patologia , Neoplasias Pulmonares/patologia , Fator de Transcrição STAT3/metabolismo , Regulação para Cima , Partículas de Ribonucleoproteínas em Forma de Abóbada/metabolismo , Idoso , Animais , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Linhagem Celular Tumoral , Núcleo Celular/metabolismo , Proliferação de Células , Sobrevivência Celular , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Masculino , Camundongos , Pessoa de Meia-Idade , Transplante de Neoplasias , Fosforilação , Prognóstico , Transdução de Sinais , Análise de Sobrevida , Partículas de Ribonucleoproteínas em Forma de Abóbada/genética
3.
Nat Commun ; 10(1): 1801, 2019 04 17.
Artigo em Inglês | MEDLINE | ID: mdl-30996248

RESUMO

Macrophage-orchestrated, low-grade chronic inflammation plays a pivotal role in obesity and atherogenesis. However, the underlying regulatory mechanisms remain incompletely understood. Here, we identify major vault protein (MVP), the main component of unique cellular ribonucleoprotein particles, as a suppressor for NF-κB signaling in macrophages. Both global and myeloid-specific MVP gene knockout aggravates high-fat diet induced obesity, insulin resistance, hepatic steatosis and atherosclerosis in mice. The exacerbated metabolic disorders caused by MVP deficiency are accompanied with increased macrophage infiltration and heightened inflammatory responses in the microenvironments. In vitro studies reveal that MVP interacts with TRAF6 preventing its recruitment to IRAK1 and subsequent oligomerization and ubiquitination. Overexpression of MVP and its α-helical domain inhibits the activity of TRAF6 and suppresses macrophage inflammation. Our results demonstrate that macrophage MVP constitutes a key constraint of NF-κB signaling thereby suppressing metabolic diseases.


Assuntos
Aterosclerose/imunologia , Fígado Gorduroso/imunologia , Inflamação/imunologia , Macrófagos/imunologia , Obesidade/imunologia , Partículas de Ribonucleoproteínas em Forma de Abóbada/metabolismo , Tecido Adiposo/patologia , Animais , Aterosclerose/etiologia , Aterosclerose/metabolismo , Biópsia , Células da Medula Óssea , Dieta Hiperlipídica/efeitos adversos , Modelos Animais de Doenças , Fígado Gorduroso/etiologia , Fígado Gorduroso/metabolismo , Feminino , Técnicas de Inativação de Genes , Humanos , Quinase I-kappa B/metabolismo , Inflamação/etiologia , Quinases Associadas a Receptores de Interleucina-1/metabolismo , Macrófagos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout para ApoE , NF-kappa B/metabolismo , Obesidade/etiologia , Obesidade/metabolismo , Obesidade/patologia , Cultura Primária de Células , Transdução de Sinais/imunologia , Fator 6 Associado a Receptor de TNF/metabolismo , Ubiquitinação , Partículas de Ribonucleoproteínas em Forma de Abóbada/genética , Partículas de Ribonucleoproteínas em Forma de Abóbada/imunologia
4.
Mater Sci Eng C Mater Biol Appl ; 98: 30-41, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30813031

RESUMO

Superparamagnetic iron oxide nanoparticles (IONPs) are promising bioactive additives to fabricate magnetic scaffolds for bone tissue engineering. To date, there has been no report on osteoinductivity of IONP-incorporated calcium phosphate cement (IONP-CPC) scaffold on stem cells using an exterior static magnetic field (SMF). The objectives of this study were to: (1) develop a novel magnetic IONP-CPC construct for bone tissue engineering, and (2) investigate the effects of IONP-incorporation and SMF application on the proliferation, osteogenic differentiation and bone mineral synthesis of human dental pulp stem cells (hDPSCs) seeded on IONP-CPC scaffold for the first time. The novel magnetic IONP-CPC under SMF enhanced the cellular performance of hDPSCs, yielding greater alkaline phosphatase activities (about 3-fold), increased expressions of osteogenic marker genes, and more cell-synthesized bone minerals (about 2.5-fold), compared to CPC control and nonmagnetic IONP-CPC. In addition, IONP-CPC induced more active osteogenesis than CPC control in rat mandible defects. These results were consistent with the enhanced cellular performance by magnetic IONP in media under SMF. Moreover, nano-aggregates were detected inside the cells by transmission electron microscopy (TEM). Therefore, the enhanced cell performance was attributed to the physical forces generated by the magnetic field together with cell internalization of the released magnetic nanoparticles from IONP-CPC constructs.


Assuntos
Fosfatos de Cálcio/química , Compostos Férricos/química , Engenharia Tecidual/métodos , Animais , Osso e Ossos/citologia , Osso e Ossos/efeitos dos fármacos , Fosfatos de Cálcio/farmacologia , Diferenciação Celular/efeitos dos fármacos , Humanos , Campos Magnéticos , Nanopartículas Metálicas/química , Microscopia Eletrônica de Transmissão , Osteogênese/efeitos dos fármacos , Ratos
5.
Int J Mol Med ; 43(1): 382-392, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30431055

RESUMO

Odonto/osteogenic differentiation of stem cells from the apical papilla (SCAPs) is a key process in tooth root formation and development. However, the molecular mechanisms underlying this process remain largely unknown. In the present study, it was identified that guanine and nucleotide binding protein 3 (GNAI3) was at least in part responsible for the odonto/osteogenic differentiation of SCAPs. GNAI3 was markedly induced in mouse tooth root development in vivo and in human SCAPs mineralization in vitro. Notably, knockdown of GNAI3 by lentiviral vectors expressing short­hairpin RNAs against GNAI3 significantly inhibited the proliferation, cell cycle progression and migration of SCAPs, as well as odonto/osteogenic differentiation of SCAPs in vitro, suggesting that GNAI3 may play an essential role in tooth root development. The promotive role of GNAI3 in odonto/osteogenic differentiation was further confirmed by downregulation of odonto/osteogenic makers in GNAI3­deficient SCAPs. In addition, knockdown of GNAI3 effectively suppressed activity of c­Jun N­terminal kinase (JNK) and extracellular­signal regulated kinase (ERK) signaling pathways that was induced during SCAPs differentiation, suggesting that GNAI3 promotes SCAPs mineralization at least partially via JNK/ERK signaling. Taken together, the present results implicate GNAI3 as a critical regulator of odonto/osteogenic differentiation of SCAPs in tooth root development, and suggest a possible role of GNAI3 in regeneration processes in dentin or other tissues.


Assuntos
Diferenciação Celular , Papila Dentária/citologia , Subunidades alfa Gi-Go de Proteínas de Ligação ao GTP/metabolismo , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Sistema de Sinalização das MAP Quinases , Odontogênese , Osteogênese , Células-Tronco/enzimologia , Animais , Antracenos/farmacologia , Biomarcadores/metabolismo , Calcificação Fisiológica/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Técnicas de Silenciamento de Genes , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Masculino , Camundongos Endogâmicos C57BL , Odontogênese/efeitos dos fármacos , Osteogênese/efeitos dos fármacos , Células-Tronco/citologia , Células-Tronco/efeitos dos fármacos , Raiz Dentária/embriologia , Raiz Dentária/metabolismo , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/genética
6.
Artigo em Inglês | MEDLINE | ID: mdl-30499649

RESUMO

One of the key factors in tissue engineering and regenerative medicine is to optimize the interaction between seed cells and scaffolds, such that the cells can grow in naturally biomimetic conditions. Their similarity to macromolecules and many unique properties mean that functional nanoparticles have promising potential for the modification and improvement of traditional scaffolds to obtain excellent biocompatibility, tunable stiffness, physical sensing, and stimulus-response capabilities. In the present study, we report magnetic poly(lactic-co-glycolic acid)/polycaprolactone (PLGA/PCL) scaffolds that were fabricated using a combination of the electrospinning technique and layer-by-layer assembly of superparamagnetic iron oxide nanoparticles (IONPs). PLGA/PCL scaffolds assembled with gold nanoparticles were prepared using the same method for comparison. The results showed that the assembled film of nanoparticles on the surface greatly enhanced the hydrophilicity and increased the elastic modulus of the scaffold, which subsequently improved the osteogenesis of the stem cells. Furthermore, the magnetic property of the IONPs was proved to be the key factor in enhancing osteogenic differentiation, which explained the superior osteogenic capacity of the magnetic scaffolds compared with that of the gold nanoparticles assembled scaffold. These results demonstrated the importance of magnetic nanomaterials as a bioactive interface between cells and scaffolds, and will promote the design of biomaterials to improve tissue engineering and regenerative medicine efficacy.

7.
Exp Cell Res ; 372(2): 158-167, 2018 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-30268758

RESUMO

Trio, the Rho guanine nucleotide exchange factor (Rho-GEF), plays diverse roles in cell migration, cell axon guidance and cytoskeleton reorganization. Conserved during evolution, Trio encodes two guanine nucleotide exchange factor domains (GEFs) and activates small GTPases. The Rho-family small GTPases RhoA and Rac1, which are target molecules of Trio, have been described to engage in craniofacial development and tooth formation. However, the exact role of Trio in tooth development remains elusive. In this study, we generated Wnt1-cre;Triofl/fl mice to address the potential function of Trio in tooth development. Wnt1-cre;Triofl/fl mice showed short root deformity as well as decreased expression of odontogenic makers such as RUNX2, OSX, OCN, and OPN. In vitro, Trio was silenced in human stem cells of dental papilla (SCAPs). Compared with the control group, the proliferation and migration ability in the experimental group was disrupted. After knocking down Trio in SCAPs, the cells showed phenotypes of poor odontogenic differentiation and weak mineralized nodules. To study the underlying mechanism, we investigated the p38 MAPK pathway and found that loss of Trio blocked the cascade transduction of p38 MAPK signaling. In conclusion, we identified Trio as a novel coordinator in regulating root development and clarified its relevant molecular events.

8.
Exp Cell Res ; 371(2): 342-352, 2018 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-30144446

RESUMO

Mesenchymal stem cells (MSCs) have been widely studied in the field of regenerative medicine with the potential to solve osteoporosis. Paired box 2 (Pax2), as a transcription factor, is the master regulator of embryogenesis and oncogenesis. However, the function of Pax2 in osteogenesis is unknown. Here, we reported for the first time that the expression of Pax2 gradually increased during osteogenic differentiation of mouse MSCs, and osteoprogenitor cells. However, detected in osteoblastic cells of mouse tibia, the expression of Pax2 in the embryonic stage was higher than that in adulthood. In C3H/10/T1/2 cells and compact bone-derived mouse MSCs (mMSCs), Pax2 knock-down inhibited the proliferation of these cells, down-regulated the expression of osteogenic marker genes, as well as repressed the ALP activity and mineralization. In addition, Pax2 enhanced the transcriptional activity of Runx2, and activated the MAPK pathway genes (ERK, JNK and p38). Furthermore, knock-down of Pax2 repressed the mMSCs-mediated bone regeneration in an ectopic bone formation model. In conclusion, Pax2 promotes osteogenesis of mouse MSCs, suggesting that Pax2 has a role in the pathophysiology of bone related diseases, and has potential application in bone tissue regeneration.

9.
J Cancer ; 9(14): 2571-2579, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30026856

RESUMO

Purpose: The role of Rac1 in cancer survival has been widely studied. However, the prognostic and clinicopathological value of Rac1 remains inconclusive. We performed a meta-analysis to clarify the role of Rac1 in cancer survival as well as its association with clinicopathological features. Methods: Eligible studies were searched from PubMed, Cochrane Library, Embase, and Web of Science databases. The pooled hazard ratios (HRs) and odds ratios (ORs) with corresponding 95% confidence intervals (CIs) were used to detect the prognostic and clinicopathological role of Rac1. Results: A total of 14 studies including 1793 patients were enrolled in the present meta-analysis. Pooled HR for overall survival (OS) (HR=2.02, 95% CI: 1.70-2.39) and disease-free survival (DFS) (HR=2.64, 95% CI: 1.71-4.09) indicated a significant poor prognostic effect for Rac1. Positive Rac1 expression was found to be correlated with tumor stage, blood vessel invasion, and lymph metastasis, but not with histological differentiation. Sensitivity test showed no single study altered OS or DFS significantly. No publication bias was detected by Egger's test and Begg's funnel plot test. Conclusion: This meta-analysis indicated that Rac1 could be used as a potential marker to predict cancer prognosis. Additionally, Rac1 expression was associated with the malignancy-related phenotype.

10.
Cell Death Differ ; 25(11): 1996-2009, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-29523871

RESUMO

The role of GATA-binding protein 4 (GATA4) in neural crest cells (NCCs) is poorly defined. Here we showed that mouse NCCs lacking GATA4 exhibited developmental defects in craniofacial bone, teeth, and heart. The defects likely occurred due to decreased cell proliferation at the developmental stage. The in vitro results were consistent with the mouse model. The isobaric tags for relative and absolute quantitation assay revealed that BARX1 is one of the differentially expressed proteins after GATA4 knockdown in NCCs. On the basis of the results of dual-luciferase, electro-mobility shift, and chromatin immunoprecipitation assays, Barx1 expression is directly regulated by GATA4 in NCCs. In zebrafish, gata4 knockdown affects the development of NCCs derivatives. However, the phenotype in zebrafish could be partly rescued by co-injection of gata4 morpholino oligomers and barx1 mRNA. This study identified new downstream targets of GATA4 in NCCs and uncovered additional evidence of the complex regulatory functions of GATA4 in NCC development.

11.
Artif Cells Nanomed Biotechnol ; : 1-11, 2018 Jan 21.
Artigo em Inglês | MEDLINE | ID: mdl-29355052

RESUMO

Literature search revealed no systematic report on iron oxide nanoparticle-incorporating calcium phosphate cement scaffolds (IONP-CPC). The objectives of this study were to: (1) use γFe2O3 nanoparticles (γIONPs) and αFe2O3 nanoparticles (αIONPs) to develop novel IONP-CPC scaffolds, and (2) investigate human dental pulp stem cells (hDPSCs) seeding on IONP-CPC for bone tissue engineering for the first time. IONP-CPC scaffolds were fabricated. Physiochemical properties of IONP-CPC scaffolds were characterized. hDPSC seeding on scaffolds, cell proliferation, osteogenic differentiation and bone matrix mineral synthesis by cells were measured. Our data demonstrated that the osteogenic differentiation of hDPSCs was markedly enhanced via IONP incorporation into CPC. Substantial increases (about three folds) in ALP activity and osteogenic gene expressions were achieved over those without IONPs. Bone matrix mineral synthesis by the cells was increased by two- to three folds over that without IONPs. The enhanced cellular osteogenesis was attributed to: (1) the surface nanotopography of IONP-CPC scaffold, and (2) the cell internalization of IONPs released from IONP-CPC scaffold. Our results demonstrate that the novel CPC functionalized with IONPs is promising to promote osteoinduction and bone regeneration. In conclusion, it is highly promising to incorporate γIONPs and αIONPs into CPC scaffold for bone tissue engineering, yielding substantially better stem cell attachment, spreading and osteogenic differentiation, and much greater bone mineral synthesis by the seeded cells. Therefore, novel CPC scaffolds containing γIONPs and αIONPs are promising for dental, craniofacial and orthopaedic applications to substantially enhance bone regeneration.

12.
J Tissue Eng Regen Med ; 12(4): e2085-e2098, 2018 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-29327431

RESUMO

A main challenge for use of scaffolds in bone engineering involves non-invasive monitoring in vivo and enhanced bone regeneration. The tissue repair effect of superparamagnetic iron oxide nanoparticles (SPIONs) was demonstrated previously by our group. However, testing in vivo is needed to confirm in vitro results. Here, SPIONs loaded gelatin sponge (GS) was used as a scaffold (SPIONs-GS) and implanted in the incisor sockets of Sprague-Dawley rats. Incisor sockets filled with nothing and filled with GS served as controls. Rats were sacrificed at 2 and 4 weeks. A significant decrease in the signal intensity of T2-weighted magnetic resonance imaging (MRI) in the SPIONs-GS group was noted. Changes in image intensity of scaffolds (indicating scaffold degradation and interaction with host tissues) could be visually monitored over time. Microcomputed tomography showed that the SPIONs-GS group had more newly formed bone (64.44 ± 10.92 vs. 28.1 ± 4.49, p < .0001) and a better preserved alveolar ridge than blank control group at 4 weeks (0.962 ± 0.01 vs. 0.92 ± 0.01, p < .0001). Histology confirmed imaging results, showing good consistency in new bone formation and scaffold degradation. The number of SPIONs decreased rapidly with time due to quick degradation of GS, whereas the number of endocytic SPIONs in cells increased with time. These residual SPIONs, together with newly formed bone, could be detected by MRI at 4 weeks. Therefore, it was clear that SPIONs induced active osteogenesis. In conclusion, good visibility on MRI and enhanced regeneration of bone can be obtained by implanting SPIONs-GS in vivo without using an external magnetic field.

13.
Int J Mol Med ; 41(2): 729-738, 2018 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-29207140

RESUMO

Platelet-rich plasma (PRP) is used in the clinic as an autologous blood product to stimulate bone regeneration and chondrogenesis. Numerous studies have demonstrated that PRP affects bone remodeling by accelerating osteoblast formation. With the research perspective focusing on osteoclasts, the present study established a mouse model of mandibular advancement to examine the effect of PRP on osteoclast differentiation induced by modification of the dynamics of the temporomandibular joint (TMJ). The lower incisors of the mice were trimmed by 1 mm and the resultant change in mandibular position during the process of eating induced condylar adaptation to this change. PRP significantly increased the bone mass and decreased osteoclastic activity, in vitro as well as in vivo. Mechanistically, the reduced expression of receptor activator of nuclear factor-κB ligand (RANKL)­induced differentiation marker genes, including nuclear factor of activated T-cells, cytoplasmic 1, c-fos and tartrate-resistant acid phosphatase, and that of the resorptive activity marker genes such as cathepsin k, carbonic anhydrase 2 and matrix metalloproteinase 9, indicated that PRP suppresses RANKL-induced osteoclast differentiation. A microarray analysis revealed that several genes associated with the Wnt pathway were differentially expressed, which indicated the involvement of this pathway in osteoclast differentiation. Furthermore, the activation of the Wnt pathway was verified by reverse transcription-quantitative polymerase chain reaction and immunoblot analysis of Dickkopf-related protein 1 and ß-catenin. The results of the present study indicated that PRP inhibits osteoclast differentiation through activation of the Wnt pathway.


Assuntos
Remodelação Óssea/genética , Diferenciação Celular/genética , Plasma Rico em Plaquetas , Ligante RANK/genética , Animais , Reabsorção Óssea/genética , Reabsorção Óssea/patologia , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Regulação da Expressão Gênica no Desenvolvimento/genética , Peptídeos e Proteínas de Sinalização Intercelular/genética , Camundongos , Análise em Microsséries , Osteoblastos/metabolismo , Osteoclastos/metabolismo , Proteínas Proto-Oncogênicas c-fos/genética , Linfócitos T/efeitos dos fármacos , Fosfatase Ácida Resistente a Tartarato/genética , Via de Sinalização Wnt/genética , beta Catenina/genética
14.
Nanomedicine ; 14(1): 35-45, 2018 01.
Artigo em Inglês | MEDLINE | ID: mdl-28887211

RESUMO

In this study, a novel calcium phosphate cement containing gold nanoparticles (GNP-CPC) was developed. Its osteogenic induction ability on human dental pulp stem cells (hDPSCs) was investigated for the first time. The incorporation of GNPs improved hDPSCs behavior on CPC, including better cell adhesion (about 2-fold increase in cell spreading) and proliferation, and enhanced osteogenic differentiation (about 2-3-fold increase at 14 days). GNPs endow CPC with micro-nano-structure, thus improving surface properties for cell adhesion and subsequent behaviors. In addition, GNPs released from GNP-CPC were internalized by hDPSCs, as verified by transmission electron microscopy (TEM), thus enhancing cell functions. The culture media containing GNPs enhanced the cellular activities of hDPSCs. This result was consistent with and supported the osteogenic induction results of GNP-CPC. In conclusion, GNP-CPC significantly enhanced the osteogenic functions of hDPSCs. GNPs are promising to modify CPC with nanotopography and work as bioactive additives thus enhance bone regeneration.


Assuntos
Cimentos para Ossos/farmacologia , Fosfatos de Cálcio/química , Polpa Dentária/citologia , Ouro/química , Nanopartículas Metálicas/administração & dosagem , Osteogênese/efeitos dos fármacos , Células-Tronco/citologia , Cimentos para Ossos/química , Adesão Celular/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Polpa Dentária/efeitos dos fármacos , Polpa Dentária/metabolismo , Humanos , Nanopartículas Metálicas/química , Células-Tronco/efeitos dos fármacos , Células-Tronco/metabolismo , Engenharia Tecidual/métodos
15.
J Mol Histol ; 48(5-6): 389-401, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-28986711

RESUMO

During tooth root development, stem cells from apical papillae (SCAPs) are indispensable, and their abilities of proliferation, migration and odontoblast differentiation are linked to root formation. Leucine-rich repeat-containing GPCR 4 (LGR4) modulates the biological processes of proliferation and differentiation in multiple stem cells. In this study, we showed that LGR4 is expressed in all odontoblast cell lineage cells and Hertwig's epithelial root sheath (HERS) during the mouse root formation in vivo. In vitro we determined that LGR4 is involved in the Wnt/ß-catenin signaling pathway regulating proliferation and odonto/osteogenic differentiation of SCAPs. Quantitative reverse-transcription PCR (qRT-PCR) confirmed that LGR4 is expressed during odontogenic differentiation of SCAPs. CCK8 assays and in vitro scratch tests, together with cell cycle flow cytometric analysis, demonstrated that downregulation of LGR4 inhibited SCAPs proliferation, delayed migration and arrested cell cycle progression at the S and G2/M phases. ALP staining revealed that blockade of LGR4 decreased ALP activity. QRT-PCR and Western blot analysis demonstrated that LGR4 silencing reduced the expression of odonto/osteogenic markers (RUNX2, OSX, OPN, OCN and DSPP). Further Western blot and immunofluorescence studies clarified that inhibition of LGR4 disrupted ß-catenin stabilization. Taken together, downregulation of LGR4 gene expression inhibited SCAPs proliferation, migration and odonto/osteogenic differentiation by blocking the Wnt/ß-catenin signaling pathway. These results indicate that LGR4 might play a vital role in SCAPs proliferation and odontoblastic differentiation.


Assuntos
Diferenciação Celular , Papila Dentária/citologia , Osteogênese , Receptores Acoplados a Proteínas-G/metabolismo , Células-Tronco/citologia , Animais , Animais Recém-Nascidos , Diferenciação Celular/genética , Movimento Celular , Proliferação de Células , Forma Celular , Regulação para Baixo/genética , Inativação Gênica , Humanos , Camundongos , Morfogênese , Odontogênese/genética , Osteogênese/genética , Estabilidade Proteica , RNA Interferente Pequeno/metabolismo , Células-Tronco/metabolismo , Raiz Dentária/citologia , Raiz Dentária/crescimento & desenvolvimento , Raiz Dentária/metabolismo , beta Catenina/metabolismo
16.
Sci Rep ; 7(1): 1534, 2017 05 08.
Artigo em Inglês | MEDLINE | ID: mdl-28484278

RESUMO

Transcription factor GATA4 regulates cardiac and osteoblast differentiation. However, its role in tooth development is not clear. Therefore, we generated Wnt1-Cre;GATA4 fl/fl mice, with conditional inactivation of the GATA4 gene in the dental papilla mesenchymal cells. Phenotypic analysis showed short root deformity along with reduced expressions of odonto/osteogenic markers. Proliferation (but not apoptosis) of cells around the apical area of the root was attenuated. In vitro, we knocked down GATA4 expression in stem cells of dental apical papilla (SCAPs). Proliferation, migration and odonto/osteogenic differentiation of SCAPs were affected in the shGATA4 group. Overexpression of GATA4 in SCAPs increased mineralization. Based on our previous iTRAQ results, guanine nucleotide binding proteins 3 (GNAI3) is one of the distinct proteins after GATA4 deletion. G protein signaling is involved in bone development, remodeling, and disease. In this study, both GATA4 deletion in the mouse root and knock-down in human SCAPs decreased the expression of GNAI3. Dual-luciferase and ChIP assay confirmed the direct binding of GATA4 to the GNAI3 promoter, both in vitro and in vivo. GNAI3 knock-down significantly decreased the odonto/osteogenic differentiation ability of SCAPs. We thus establish the role of GATA4 as a novel regulator of root development and elucidate its downstream molecular events.

17.
J Mol Histol ; 48(3): 187-197, 2017 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-28393293

RESUMO

Osteoblasts play a major role in bone remodeling and are regulated by transcription factors. GATA4, a zinc finger transcription factor from the GATA family, has an unclear role in osteoblast differentiation. In this study, the role of GATA4 in osteoblast differentiation was studied both in vitro and in vivo by GATA4 knockdown. GATA4 expression increased during osteoblast differentiation. GATA4 knockdown in osteoblast precursor cells reduced alkaline phosphatase activity and decreased the formation of calcified nodule in an osteogenic-induced cell culture system. In vivo, micro-CT showed that local injection of lentivirus-delivered GATA4 shRNA caused reduced new bone formation during tooth movement. Histological analyses such as total collagen and Goldner's trichrome staining confirmed these results. In vivo immunohistochemical analysis showed reduced expression of osterix (OSX), osteopontin (OPN), and osteocalcin (OCN) in the shGATA4 group (P < 0.05). Consistently, both western blotting and quantitative reverse-transcription PCR proved that expression of osteogenesis-related genes, including OSX, OPN, and OCN, was significantly repressed in the shGATA4 group in vitro (P < 0.01). For further analysis of the pathways involved in this process, we examined the MAPK signaling pathway, and found knockdown of GATA4, downregulated p38 signaling pathways (P < 0.01). Collectively, these results imply GATA4 is a regulator of osteoblastic differentiation via the p38 signaling pathways.


Assuntos
Remodelação Óssea , Diferenciação Celular , Fator de Transcrição GATA4/fisiologia , Sistema de Sinalização das MAP Quinases , Osteoblastos/citologia , Animais , Células Cultivadas , Fator de Transcrição GATA4/genética , Imuno-Histoquímica , Masculino , Camundongos Endogâmicos C57BL , Osteoblastos/fisiologia , RNA Interferente Pequeno/administração & dosagem , RNA Interferente Pequeno/farmacologia
18.
J Mol Histol ; 48(3): 199-208, 2017 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-28409326

RESUMO

Corticision is a common technique to accelerate orthodontic tooth movement; however, not much is known about the underlying mechanisms. In this study, we investigated the mechanism of alveolar tissue remodeling after corticision in a rat model of tooth movement (TM) by analyzing the differential transcriptome. A total of 36 male rats were equally divided into TM and TM with corticision (TM+C) groups. Alveolar bone response was examined using micro-computed tomography (micro-CT). Osteoclasts and osteoblasts were quantified on tartrate-resistant acid phosphatase (TRAP) and Goldner's trichrome staining. The transcriptomes of alveolus around the left maxillary first molar were determined on RNA sequencing (RNA-Seq), and the expression of selected differentially expressed genes (DEGs) validated on quantitative reverse-transcriptase polymerase chain reaction (qRT-PCR). Immunohistochemical examination of alveolar tissue was performed to examine the expressions of correlative proteins of the selected signaling pathway in the TM and TM+C groups. The ratio of bone volume to total volume (BV/TV), and the trabecular number (Tb.N) were significantly decreased, while the movement distance and the trabecular separation (Tb.Sp) was significantly increased in the TM+C group. However, no significant between-group difference in trabecular thickness (Tb.Th) was observed. On histomorphometric analysis, a significant increase in the number of osteoclasts and increased bone resorption was observed in the TM+C group. A total of 399 DEGs were identified on RNA-SEq. Eleven selected genes were confirmed on qRT-PCR, which included components of the Ras signaling pathway. Four proteins of the Ras signaling pathway showed a higher expression in the TM+C group. Our findings indicate that corticision may speed up orthodontic tooth movement by accelerating osteoclastogenesis mediated via the Ras signaling pathway.


Assuntos
Remodelação Óssea/genética , Análise de Sequência de RNA , Técnicas de Movimentação Dentária , Transcriptoma/genética , Processo Alveolar , Animais , Regulação da Expressão Gênica , Masculino , Osteoclastos , Ratos , Microtomografia por Raio-X , Proteínas ras/metabolismo
19.
Cancer Res ; 77(7): 1586-1598, 2017 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-28202524

RESUMO

Mechanisms of cross-talk between tumor cells and tumor-associated macrophages (TAM), which drive metastasis, are not fully understood. Scavenger receptor A1 (SR-A1) expressed primarily in macrophages has been associated with lung tumorigenesis. In this study, we used population genetics, transcriptomics, and functional analyses to uncover how SR-A1 is involved in lung cancer and its prognosis. SR-A1 genetic variants were investigated for possible association with survival of advanced stage NSCLC patients in the Harvard Lung Cancer Study cohort. Two SNPs (rs17484273, rs1484751) in SR-A1 were associated significantly with poor overall survival in this cohort. Data from The Cancer Genome Atlas showed considerable downregulation of SR-A1 in lung tumor tissues. The association of SR-A1 with prognosis was validated in animal models in the context of lung cancer metastasis. Macrophages derived from mice genetically deficient for SR-A1 exhibited accelerated metastasis in a model of lung cancer. On the other hand, tumor cell seeding, migration, and invasion, as well as macrophage accumulation in lung cancer tissue, were enhanced in SR-A1-deficient mice. SR-A1 deletion upregulated serum amyloid A1 (SAA1) in macrophages via MAPK/IκB/NFκB signaling. SAA1 promoted tumor cell invasion and macrophage migration in vitro and in vivo, but these effects were blocked by administration of an anti-SAA1 antibody. Overall, our findings show how SR-A1 suppresses lung cancer metastasis by downregulating SAA1 production in TAMs. Cancer Res; 77(7); 1586-98. ©2017 AACR.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/patologia , Neoplasias Pulmonares/patologia , Macrófagos/fisiologia , Metástase Neoplásica/prevenção & controle , Receptores Depuradores Classe A/fisiologia , Proteína Amiloide A Sérica/fisiologia , Animais , Carcinoma Pulmonar de Células não Pequenas/mortalidade , Movimento Celular , Feminino , Humanos , Neoplasias Pulmonares/mortalidade , Camundongos , Invasividade Neoplásica , Polimorfismo de Nucleotídeo Único , Prognóstico , Receptores Depuradores Classe A/genética , Microambiente Tumoral
20.
Int J Biol Sci ; 12(10): 1155-1167, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-27766031

RESUMO

Osteoclasts originate from bone marrow monocyte/macrophage lineage cells, which are important for bone health. Class A scavenger receptor (SR-A) is a multifunctional molecule that functions during differentiation of monocyte into macrophages and osteoclasts. To further characterize the role of SR-A in osteoclasts, we used the murine tooth movement model (TM) and the murine anterior cruciate ligament transection model of osteoarthritis (ACLT OA). In these two models the bones involved are of different origin and have different properties. Bone resorption was decreased in SR-A-/- mice compared to SR-A+/+ mice. Further evaluation showed that the number of multinucleated osteoclasts in SR-A-/- mice, compared to SR-A+/+ mice, was significantly decreased both in vivo and in vitro. The levels of interleukin-6 (IL-6) produced by osteoclasts were reduced in SR-A-/- mice compared to SR-A+/+ mice. In the in vitro marrow-derived osteoclast formation assay and in both mouse models, osteoclastogenesis was restored to normal in SR-A-/- mice by administration of recombinant murine IL-6. Moreover, neutralization of IL-6 reduced the number of osteoclasts formed in SR-A+/+ mice of TM model. Both extracellular signal-regulated kinase (ERK) and c-Jun N-terminal protein kinase (JNK), but not p38, signaling pathways were downregulated in receptor activator of nuclear factor-κB ligand (RANKL)-stimulated SR-A-/- osteoclasts. Importantly, when treated with either ERK or JNK inhibitor, the numbers of osteoclasts generated from RANKL-induced bone marrow derived-macrophages of SR-A+/+ mice, and their IL-6 production, were significantly decreased. This suggests that SR-A activates the ERK and JNK signaling pathways, and promotes production of IL-6 by osteoclasts to further stimulate osteoclast formation.


Assuntos
MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Interleucina-6/metabolismo , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Osteoclastos/citologia , Osteogênese/fisiologia , Receptores Depuradores Classe A/metabolismo , Animais , Ensaio de Imunoadsorção Enzimática , MAP Quinases Reguladas por Sinal Extracelular/genética , Interleucina-6/genética , Proteínas Quinases JNK Ativadas por Mitógeno/genética , Macrófagos/citologia , Macrófagos/metabolismo , Camundongos , Osteoartrite/metabolismo , Osteoclastos/metabolismo , Osteogênese/genética , Ligante RANK/genética , Ligante RANK/metabolismo , Receptores Depuradores Classe A/genética , Transdução de Sinais/genética , Transdução de Sinais/fisiologia , Microtomografia por Raio-X
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