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1.
Hua Xi Kou Qiang Yi Xue Za Zhi ; 38(3): 314-318, 2020 Jun 01.
Artigo em Chinês | MEDLINE | ID: mdl-32573141

RESUMO

Tubular dentin is of great significance in the process of tooth tissue and tooth regeneration, because it is not only the structural feature of primary dentin, but also can affect the tooth sensory function, affect the differentiation of dental pulp cells and provide strong mechanical support for teeth. Scaffold is one of the three elements of tissue engineering dentin regeneration. Most experiments on dentin regeneration involve the study of the microstructure and mechanical properties of the scaffold. The microstructure and mechanical characteristics of scaffold materials have important effects on the differentiation and adhesion of odontoblast, it can directly affect the tissue structure of regenerated dentin.


Assuntos
Polpa Dentária , Tecidos Suporte , Diferenciação Celular , Dentina , Odontoblastos , Regeneração , Engenharia Tecidual
2.
Hua Xi Kou Qiang Yi Xue Za Zhi ; 38(3): 330-337, 2020 Jun 01.
Artigo em Chinês | MEDLINE | ID: mdl-32573144

RESUMO

Human periodontal ligament-derived cells serve as an important source of seeding cells in periodontal regenerative medicine, and their osteogenic potential is closely related to alveolar bone repair and periodontal regeneration. Non-coding RNA (ncRNA), such as microRNA, long non-coding RNA, and circular RNA, play important roles in the regu-lation of osteogenic genes in human periodontal ligament-derived cells. In this review, we summarize the target genes, path-ways, and functions of the ncRNA network during osteogenic differentiation of periodontal ligament-derived cells.


Assuntos
MicroRNAs , Ligamento Periodontal , Diferenciação Celular , Células Cultivadas , Humanos , Osteogênese
3.
Zhongguo Zhong Yao Za Zhi ; 45(7): 1676-1683, 2020 Apr.
Artigo em Chinês | MEDLINE | ID: mdl-32489049

RESUMO

The objective of this study was to investigate the inhibitory effect of scutellarin on the differentiation of colonic cancer stem cells in vitro and in vivo and to explore its underlying hedgehog signaling-based mechanism. The effect of scutellarin on the growth in vitro of HT-29 cells-derived cancer stem-like cells(HT-29 CSC) was observed with 3 D cell culture. The effect of scutellarin on the transformation of HT-29 CSC cells was assessed by soft agar colony formation assay. Fetal calf serum was used to induce differentiation of stem cells and observe the effect of scutellarin on HT-29 CSC cells differentiation in vitro. The effects of scutellarin on mRNA expressions of Lgr5, c-Myc, CK20 and Nanog in HT-29 CSC cells were determined by quantitative Real-time polymerase chain reaction(qRT-PCR). The effects of scutellarin on protein expressions of c-Myc, Gli1 and Lgr5 in HT-29 CSC cells were examined by Western blot. After subcutaneous implantation of HT-29 CSC cells in nude mice, the effect of scutellarin on the mouse body weight and the growth of HT-29 CSC-derived tumor were explored. qRT-PCR was used for evaluating the effect of scutellarin on mRNA levels of CD133, Lgr5, Gli1, Ptch1, c-Myc, Ki-67, CK20 and Nanog in tumor. Western blot and immunohistochemistry analysis were used to detect the effect of scutellarin on protein expressions of c-Myc, Gli1, Lgr5, CD133 and Ki-67 in tumor. The in vitro experiments showed that scutellarin inhibited the growth, transformation and differentiation of HT-29 CSC cells, significantly down-regulated the mRNA levels of Lgr5, c-Myc, CK20 and Nanog in HT-29 CSC cells as well as the protein expression levels of c-Myc, Gli1 and Lgr5 in HT-29 CSC cells. Additionally, animal experiments showed that scutellarin significantly inhibited the growth of subcutaneous xenografts in nude mice, and down-regulated the mRNA expressions of CD133, Lgr5, Gli1, Ptch1, c-Myc, Ki-67, CK20 and Nanog as well as the protein levels of c-Myc, Gli1, Lgr5, CD133 and Ki-67 of xenografts in nude mice. Taken together, scutellarin could inhibit the differentiation of colo-nic cancer stem cells in vitro and in vivo, potentially by down regulation of hedgehog signaling pathway activity.


Assuntos
Células-Tronco Neoplásicas , Animais , Apigenina , Diferenciação Celular , Linhagem Celular Tumoral , Proliferação de Células , Glucuronatos , Proteínas Hedgehog , Humanos , Camundongos , Camundongos Nus
4.
J Oral Sci ; 62(3): 298-302, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32581177

RESUMO

In this study, a Porphyromonas gingivalis (P.g.)-infected mouse periodontitis model was used to investigate the effect of omega-3 fatty acid intake on differentiation and maturation of cultured osteoclast. Four-week-old C57BL/6JJcl mice were divided into four groups according to the diets they were fed from the beginning of the experiment (i.e., food containing omega-3 or omega-6 fatty acids) and whether they were orally administered P.g. Thirty-three days after beginning the experiment, bone marrow cells were sampled from the femoral bone of mice from each group and differentiated into osteoclasts; the effects of the ingestion of different fatty acids were subsequently investigated. There was no statistical interaction between the different fatty acids and P.g. infection on the number of osteoclasts (P = 0.6). However, the fatty acid type affected the number of osteoclasts in mice (P = 0.0013), with the omega-3 groups demonstrating lower osteoclast numbers than the omega-6 groups. Furthermore, the addition of resolvin E1 (RvE1), which is an omega-3 fatty acid-derived lipid mediator, suppressed the differentiation of mouse cultured osteoclasts (P < 0.0001). Therefore, the ingestion of omega-3 fatty acids may suppress osteoclast differentiation while inhibiting bone resorption and tissue destruction due to periodontitis.


Assuntos
Perda do Osso Alveolar , Ácidos Graxos Ômega-3 , Animais , Diferenciação Celular , Camundongos , Camundongos Endogâmicos C57BL , Osteoclastos , Porphyromonas gingivalis
5.
Braz Dent J ; 31(2): 122-126, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32556010

RESUMO

Although periodontitis is one of the commonest infectious inflammatory diseases in humans, the mechanisms involved with its immunopathology remain ill understood. Numerous molecules may induce inflammation and lead to bone resorption, secondary to activation of monocytes into osteoclasts. TACE (TNF-α converting enzyme) and DC-STAMP (dendritic cell-specific transmembrane protein) appear to play a role on bone resorption since TACE induces the release of sRANKL (soluble receptor activator of nuclear factor kappa-ß ligand) whereas DC-STAMP is a key factor in osteoclast induction. The present study evaluated the levels of TACE and DC-STAMP in patients with and without periodontitis. Twenty individuals were selected: 10 periodontally healthy participants undergoing gingivectomy for esthetic reasons and 10 diagnosed with periodontitis. Protein levels of such molecules in gingival tissue were established using Western blotting. Protein levels of both TACE and DC-STAMP were higher in the periodontitis group than in the control group (p<0.05; Student t-test). In conclusion, TACE and DC-STAMP protein levels are elevated in patients with periodontitis, favoring progression of bone resorption.


Assuntos
Proteína ADAM17 , Proteínas Adaptadoras de Transdução de Sinal , Reabsorção Óssea , Proteínas de Membrana , Periodontite , Proteína ADAM17/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Diferenciação Celular , Humanos , Proteínas de Membrana/metabolismo , Osteoclastos
6.
Chin J Dent Res ; 23(2): 119-130, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32548603

RESUMO

OBJECTIVE: To investigate and characterise the differences between the open chromatin regions of oral and epidermal keratinocytes. METHODS: Human immortalised oral epithelial cell lines (HIOECs) were used as the standard model for oral keratinocytes, and primary normal human epidermal keratinocytes (NHEKs) were chosen as the model for epidermal keratinocytes. Assay for transposase accessible chromatin using sequencing (ATAC-seq) and H3K27ac chromatin immunoprecipitation sequencing (ChIP-seq) were used to evaluate the dynamic changes in open chromatin regions and active enhancers during oral keratinocyte differentiation. In silico prediction and dual-luciferase assays were used to evaluate the enriched motifs and maintain enhancer activity in specific enriched HIOECs. Integration and comparison of HIOEC ATAC-seq with NHEK ATAC-seq were used to identify oral keratinocyte-enriched open chromatin regions along with key motifs governing differential enhancer activity. The genomic regulatory elements and GWAS overlap algorithm was used to compare the annotation rate of HIOEC-overlapped craniofacial enhancers with other craniofacial enhancers for orofacial cleft-associated variants. RESULTS: During the differentiation of HIOECs, 14933 open chromatin regions became more accessible. Grainyhead-like (GRHL) and Krüppel-like factor (KLF) motifs were overrepresented in maintaining HIOEC-specific activity. Compared with NHEKs, 16161 open chromatin regions were uniquely accessible in HIOECs. Within these regions, the C/EBP motif governed HIOEC-specific enhancer regulating SOX2 and PITX2, which enhanced oral keratinocyte wound healing. When intersected with human craniofacial super-enhancers, open chromatin regions in HIOECS can better annotate the common variants associated with orofacial cleft. CONCLUSION: The intrinsic differences between the open chromatin regions of human oral and epidermal keratinocytes are directly maintained by a set of transcription factors.


Assuntos
Cromatina , Fenda Labial , Diferenciação Celular , Humanos , Queratinócitos , Fatores de Transcrição/genética
7.
Chin J Dent Res ; 23(2): 143-150, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32548605

RESUMO

OBJECTIVE: To compare the biological characteristics of dental pulp stem cells (DPSCs) and inflamed dental pulp derived stem cells (I-DPSCs) in vitro and their regeneration potential in Beagle immature premolars. METHODS: Pulpitis was induced in the premolars of one beagle dog by opening the pulp chamber for 2 weeks, and inflammation was histologically confirmed. DPSCs and I-DPSCs were isolated from normal and inflamed dental pulp, and cell morphology, expression of mesenchymal stem cell markers, clone formation ability, cell proliferation and osteogenic/odontogenic differentiation potential were compared. The dental pulp of 20 roots from 10 immature premolars was extracted and divided into two groups. DPSCs or I-DPSCs with scaffolds were transplanted into the root canals. The roots were extracted after 3 months, and pulp regeneration was evaluated by histological analysis. The data were statistically analysed using one-way ANOVA and a Student t test. RESULTS: Histological analyses showed lymphocyte infiltration and elevated TNF-α expression, which confirmed the diagnosis of pulpitis. I-DPSCs showed similar morphology, marker gene expression and clone formation ability but greater proliferation ability and osteogenic/odontogenic differentiation potential. Pulp-like tissue formation and bone- and dentine-like tissue deposition were observed in both DPSC- and I-DPSC-transplanted roots. CONCLUSION: DPSCs derived from inflammatory dental pulp tissue have similar biological characteristics to those from normal dental pulp and could mediate pulp and dentine regeneration in immature premolars.


Assuntos
Polpa Dentária , Regeneração , Animais , Dente Pré-Molar , Diferenciação Celular , Proliferação de Células , Células Cultivadas , Cães , Humanos , Células-Tronco
8.
Anticancer Res ; 40(6): 3039-3047, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32487597

RESUMO

Over the past few decades, longevity without disease has become an important topic worldwide. However, as life expectancy increases, the number of patients with cancer is also increasing. Tumor progression is related to interactions between tumor cells and mesenchymal stem cells (MSCs) in the tumor microenvironment. MSCs are multipotent stromal cells known to be present in a variety of locations in the body, including bones, cartilage, fat, muscles, and dental pulp. MSCs migrate toward inflamed areas during pathological immune responses. MSCs also migrate toward tumor stroma and participate in tumor progression. MSCs can contribute to tumor progression by interacting with tumor cells via paracrine signaling and differentiate into diverse cell types. This also enables MSCs to make direct contact with tumor cells in tumor stroma. Interactions between tumor cells and MSCs enhance tumorigenic and metastatic potential, in addition to stimulating epithelial to mesenchymal transition. Herein, we reviewed the research associated with the tumor-enhancing role of MSCs in tumor progression, from primary tumor growth to distant tumor metastasis.


Assuntos
Células-Tronco Mesenquimais/metabolismo , Diferenciação Celular , Progressão da Doença , Humanos , Microambiente Tumoral
9.
Shanghai Kou Qiang Yi Xue ; 29(1): 7-12, 2020 Feb.
Artigo em Chinês | MEDLINE | ID: mdl-32524113

RESUMO

PURPOSE: To analyze the effect of recombinant connective tissue growth factor(rCTGF) on proliferation and differentiation of human dental pulp cells(hDPCs). METHODS: Human dental pulp cells were cultured in vitro and treated with rCTGF at different concentrations (0, 1, 10, 100 ng/mL). The proliferation of dental pulp cells was detected by CCK8 assay. The formation of mineralized nodules was determined by alizarin red staining and half-quantitative alizarin Red S assay. qRT-PCR was utilized to detect the expression of odontogenic differentiation related genes DMP-1, DSPP and OC, and the phosphorylation level of ERK1/2 signaling pathway was detected by Western blot. The data were analyzed with SAS 9.3 software package. RESULTS: High concentration of rCTGF(100 ng/mL) could promote proliferation of dental pulp cells. After mineralization induction, 10 g/mL rCTGF had the best effect on promoting the formation of mineralized nodules in dental pulp cells, and calcium ion deposition was the most obvious(P<0.05). The expression of odontogenic differentiation related genes DMP-1 and DSPP was significantly up-regulated(P<0.05). Western blot results showed that hDPCs stimulated by 10 ng/mL rCTGF could increase the expression of p-ERK1/2. CONCLUSIONS: rCTGF may promote the proliferation and differentiation of human dental pulp cells through activating ERK1/2 signaling pathway.


Assuntos
Fator de Crescimento do Tecido Conjuntivo , Polpa Dentária , Fosfatase Alcalina , Diferenciação Celular , Proliferação de Células , Células Cultivadas , Humanos , Odontogênese
10.
Science ; 368(6490): 497-505, 2020 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-32355025

RESUMO

Androgen deprivation is the cornerstone of prostate cancer treatment. It results in involution of the normal gland to ~90% of its original size because of the loss of luminal cells. The prostate regenerates when androgen is restored, a process postulated to involve stem cells. Using single-cell RNA sequencing, we identified a rare luminal population in the mouse prostate that expresses stemlike genes (Sca1 + and Psca +) and a large population of differentiated cells (Nkx3.1 +, Pbsn +). In organoids and in mice, both populations contribute equally to prostate regeneration, partly through androgen-driven expression of growth factors (Nrg2, Rspo3) by mesenchymal cells acting in a paracrine fashion on luminal cells. Analysis of human prostate tissue revealed similar differentiated and stemlike luminal subpopulations that likewise acquire enhanced regenerative potential after androgen ablation. We propose that prostate regeneration is driven by nearly all persisting luminal cells, not just by rare stem cells.


Assuntos
Androgênios/metabolismo , Próstata/fisiologia , Próstata/cirurgia , Neoplasias da Próstata/cirurgia , Regeneração , Antagonistas de Androgênios/uso terapêutico , Proteína de Ligação a Androgênios/genética , Animais , Antígenos de Neoplasias/genética , Ataxina-1/genética , Diferenciação Celular/genética , Proteínas Ligadas por GPI/genética , Expressão Gênica , Proteínas de Homeodomínio/genética , Humanos , Masculino , Células-Tronco Mesenquimais/fisiologia , Camundongos , Proteínas de Neoplasias/genética , Fatores de Crescimento Neural/genética , Tamanho do Órgão , Organoides/metabolismo , Organoides/fisiologia , Próstata/metabolismo , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/metabolismo , Regeneração/genética , Análise de Sequência de RNA , Análise de Célula Única , Trombospondinas/genética , Fatores de Transcrição/genética
11.
Gene ; 752: 144758, 2020 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-32422235

RESUMO

Drugs targeting chromatin-modifying enzymes have entered clinical trials for myeloid malignancies, including INCB059872, a selective irreversible inhibitor of Lysine-Specific Demethylase 1 (LSD1). While initial studies of LSD1 inhibitors suggested these compounds may be used to induce differentiation of acute myeloid leukemia (AML), the mechanisms underlying this effect and dose-limiting toxicities are not well understood. Here, we used precision nuclear run-on sequencing (PRO-seq) and ChIP-seq in AML cell lines to probe for the earliest regulatory events associated with INCB059872 treatment. The changes in nascent transcription could be traced back to a loss of CoREST activity and activation of GFI1-regulated genes. INCB059872 is in phase I clinical trials, and we evaluated a pre-treatment bone marrow sample of a patient who showed a clinical response to INCB059872 while being treated with azacitidine. We used single-cell RNA-sequencing (scRNA-seq) to show that INCB059872 caused a shift in gene expression that was again associated with GFI1/GFI1B regulation. Finally, we treated mice with INCB059872 and performed scRNA-seq of lineage-negative bone marrow cells, which showed that INCB059872 triggered accumulation of megakaryocyte early progenitor cells with gene expression hallmarks of stem cells. Accumulation of these stem/progenitor cells may contribute to the thrombocytopenia observed in patients treated with LSD1 inhibitors.


Assuntos
Regulação Leucêmica da Expressão Gênica/efeitos dos fármacos , Histona Desmetilases/antagonistas & inibidores , Leucemia Mieloide Aguda/metabolismo , Animais , Antineoplásicos/farmacologia , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Feminino , Histona Desmetilases/genética , Histona Desmetilases/metabolismo , Humanos , Leucemia Mieloide Aguda/genética , Camundongos , Camundongos Endogâmicos C57BL , RNA-Seq , Análise de Célula Única/métodos , Células-Tronco/metabolismo , Células THP-1 , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Sequenciamento Completo do Exoma/métodos
12.
J Cell Mol Med ; 24(12): 6988-6999, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32374474

RESUMO

Outbreaks of infections with viruses like Sars-CoV-2, Ebola virus and Zika virus lead to major global health and economic problems because of limited treatment options. Therefore, new antiviral drug candidates are urgently needed. The promising new antiviral drug candidate silvestrol effectively inhibited replication of Corona-, Ebola-, Zika-, Picorna-, Hepatis E and Chikungunya viruses. Besides a direct impact on pathogens, modulation of the host immune system provides an additional facet to antiviral drug development because suitable immune modulation can boost innate defence mechanisms against the pathogens. In the present study, silvestrol down-regulated several pro- and anti-inflammatory cytokines (IL-6, IL-8, IL-10, CCL2, CCL18) and increased TNF-α during differentiation and activation of M1-macrophages, suggesting that the effects of silvestrol might cancel each other out. However, silvestrol amplified the anti-inflammatory potential of M2-macrophages by increasing expression of anti-inflammatory surface markers CD206, TREM2 and reducing release of pro-inflammatory IL-8 and CCL2. The differentiation of dendritic cells in the presence of silvestrol is characterized by down-regulation of several surface markers and cytokines indicating that differentiation is impaired by silvestrol. In conclusion, silvestrol influences the inflammatory status of immune cells depending on the cell type and activation status.


Assuntos
Antivirais/farmacologia , Betacoronavirus/efeitos dos fármacos , Citocinas/genética , Células Dendríticas/efeitos dos fármacos , Fatores Imunológicos/farmacologia , Macrófagos/efeitos dos fármacos , Triterpenos/farmacologia , Betacoronavirus/crescimento & desenvolvimento , Betacoronavirus/imunologia , Diferenciação Celular/efeitos dos fármacos , Vírus Chikungunya/efeitos dos fármacos , Vírus Chikungunya/crescimento & desenvolvimento , Vírus Chikungunya/imunologia , Citocinas/classificação , Citocinas/imunologia , Células Dendríticas/imunologia , Células Dendríticas/virologia , Ebolavirus/efeitos dos fármacos , Ebolavirus/crescimento & desenvolvimento , Ebolavirus/imunologia , Perfilação da Expressão Gênica , Regulação da Expressão Gênica/efeitos dos fármacos , Vírus da Hepatite E/efeitos dos fármacos , Vírus da Hepatite E/crescimento & desenvolvimento , Vírus da Hepatite E/imunologia , Humanos , Imunidade Inata/efeitos dos fármacos , Macrófagos/imunologia , Macrófagos/virologia , Especificidade de Órgãos , Picornaviridae/efeitos dos fármacos , Picornaviridae/crescimento & desenvolvimento , Picornaviridae/imunologia , Cultura Primária de Células , Transdução de Sinais , Zika virus/efeitos dos fármacos , Zika virus/crescimento & desenvolvimento , Zika virus/imunologia
13.
Biol Res ; 53(1): 22, 2020 May 19.
Artigo em Inglês | MEDLINE | ID: mdl-32430065

RESUMO

BACKGROUND: Duchenne muscular dystrophy (DMD) is a devastating genetic muscular disorder with no effective treatment that is caused by the loss of dystrophin. Human induced pluripotent stem cells (hiPSCs) offer a promising unlimited resource for cell-based therapies of muscular dystrophy. However, their clinical applications are hindered by inefficient myogenic differentiation, and moreover, the engraftment of non-transgene hiPSC-derived myogenic progenitors has not been examined in the mdx mouse model of DMD. METHODS: We investigated the muscle regenerative potential of myogenic progenitors derived from hiPSCs in mdx mice. The hiPSCs were transfected with enhanced green fluorescent protein (EGFP) vector and defined as EGFP hiPSCs. Myogenic differentiation was performed on EGFP hiPSCs with supplementary of basic fibroblast growth factor, forskolin, 6-bromoindirubin-3'-oxime as well as horse serum. EGFP hiPSCs-derived myogenic progenitors were engrafted into mdx mice via both intramuscular and intravenous injection. The restoration of dystrophin expression, the ratio of central nuclear myofibers, and the transplanted cells-derived satellite cells were accessed after intramuscular and systemic transplantation. RESULTS: We report that abundant myogenic progenitors can be generated from hiPSCs after treatment with these three small molecules, with consequent terminal differentiation giving rise to mature myotubes in vitro. Upon intramuscular or systemic transplantation into mdx mice, these myogenic progenitors engrafted and contributed to human-derived myofiber regeneration in host muscles, restored dystrophin expression, ameliorated pathological lesions, and seeded the satellite cell compartment in dystrophic muscles. CONCLUSIONS: This study demonstrates the muscle regeneration potential of myogenic progenitors derived from hiPSCs using non-transgenic induction methods. Engraftment of hiPSC-derived myogenic progenitors could be a potential future therapeutic strategy to treat DMD in a clinical setting.


Assuntos
Células-Tronco Pluripotentes Induzidas/transplante , Distrofia Muscular de Duchenne/terapia , Animais , Diferenciação Celular , Células Cultivadas , Modelos Animais de Doenças , Proteínas de Fluorescência Verde , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL
14.
Int J Nanomedicine ; 15: 2501-2513, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32368037

RESUMO

Purpose: The extracellular matrix (ECM) labyrinthine network secreted by mesenchymal stem cells (MSCs) provides a microenvironment that enhances cell adherence, proliferation, viability, and differentiation. The potential of graphene-based nanomaterials to mimic a tissue-specific ECM has been recognized in designing bone tissue engineering scaffolds. In this study, we investigated the expression of specific ECM proteins when human fat-derived adult MSCs adhered and underwent osteogenic differentiation in the presence of functionalized graphene nanoparticles. Methods: Graphene nanoparticles with 6-10% oxygen content were prepared and characterized by XPS, FTIR, AFM and Raman spectroscopy. Calcein-am and crystal violet staining were performed to evaluate viability and proliferation of human fat-derived MSCs on graphene nanoparticles. Alizarin red staining and quantitation were used to determine the effect of graphene nanoparticles on osteogenic differentiation. Finally, immunofluorescence assays were used to investigate the expression of ECM proteins during cell adhesion and osteogenic differentiation. Results: Our data show that in the presence of graphene, MSCs express specific integrin heterodimers and exhibit a distinct pattern of the corresponding bone-specific ECM proteins, primarily fibronectin, collagen I and vitronectin. Furthermore, MSCs undergo osteogenic differentiation spontaneously without any chemical induction, suggesting that the physicochemical properties of graphene nanoparticles might trigger the expression of bone-specific ECM. Conclusion: Understanding the cell-graphene interactions resulting in an osteogenic niche for MSCs will significantly improve the application of graphene nanoparticles in bone repair and regeneration.


Assuntos
Proteínas da Matriz Extracelular/metabolismo , Grafite/farmacologia , Células-Tronco Mesenquimais/metabolismo , Nanopartículas/química , Osteogênese/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Humanos , Integrinas/metabolismo , Células-Tronco Mesenquimais/efeitos dos fármacos , Oxigênio/química , Espectroscopia Fotoeletrônica , Multimerização Proteica
15.
Int J Nanomedicine ; 15: 2633-2646, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32368045

RESUMO

Objective: The aim of this study is to fabricate functional scaffolds to gene delivery bone morphogenetic protein-2 (BMP-2) plasmid for bone formation in bone tissue engineering. Methods: Dendriplexes (DPs) of generation 4 polyamidoamin (G4-PAMAM)/BMP-2 plasmid were prepared through microfluidic (MF) platform. The physiochemical properties and toxicity of DPs were evaluated by DLS, AFM, FESEM and MTT assay. In order to create a suitable environment for stem cell growth and differentiation, poly-l-lactic acid (PLLA) and poly-l-lactic acid/poly (ethylene oxide) (PLLA/PEO) scaffolds containing hydroxyapatite nanoparticles (HA) and DPs were fabricated by the electrospinning method. The osteogenic potency of the scaffolds on human adipose tissue-derived mesenchymal stem cells (hASCs) was investigated. Results: The results revealed that tuning the physical properties of DPs by adjusting flow parameters in microfluidic platform can easily improve the cell viability compared to conventional bulk mixing method. Also, the result showed that the presence of HA and DPs in PLLA/PEO scaffold enhanced alkaline phosphatase (ALP) activity and increased the amount of deposited Ca, as well as, related to osteogenesis gen markers. Conclusion: This study indicated that on using the MF platform in preparation of DPs and loading them along with HA in PLLA/PEO scaffold, the osteogenic differentiation of hASCs could be tuned.


Assuntos
Proteína Morfogenética Óssea 2/metabolismo , Osso e Ossos/fisiologia , Durapatita/química , Microfluídica , Nanofibras/química , Poliaminas/química , Engenharia Tecidual/métodos , Tecidos Suporte/química , Fosfatase Alcalina/metabolismo , Cálcio/metabolismo , Adesão Celular , Morte Celular , Diferenciação Celular , Proliferação de Células , Forma Celular , DNA/metabolismo , Dendrímeros/química , Humanos , Células-Tronco Mesenquimais/metabolismo , Nanopartículas/química , Tamanho da Partícula , Plasmídeos/metabolismo , Poliésteres/química , Resistência à Tração
16.
Int J Nanomedicine ; 15: 2669-2683, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32368048

RESUMO

Background: Zinc oxide nanoparticles (ZnO NPs) are one of the most widely used nanomaterials in a variety of fields such as industrial, pharmaceutical, and household applications. Increasing evidence suggests that ZnO NPs could elicit unignorable harmful effect to the cardiovascular system, but the potential deleterious effects to human cardiomyocytes remain to be elucidated. Human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) have been increasingly used as a promising in vitro model of cardiomyocyte in various fields such as drug cardiac safety evaluation. Herein, the present study was designed to elucidate the cardiac adverse effects of ZnO NPs and explore the possible underlying mechanism using hiPSC-CMs. Methods: ZnO NPs were characterized by transmission electron microscopy and dynamic light scattering. The cytotoxicity induced by ZnO NPs in hiPSC-CMs was evaluated by determination of cell viability and lactate dehydrogenase release. Cellular reactive oxygen species (ROS) and mitochondrial membrane potential were measured by high-content analysis (HCA). Mitochondrial biogenesis was assayed by detection of mtDNA copy number and PGC-1α pathway. Moreover, microelectrode array techniques were used to investigate cardiac electrophysiological alterations. Results: We demonstrated that ZnO NPs concentration- and time-dependently elicited cytotoxicity in hiPSC-CMs. The results from HCA revealed that ZnO NPs exposure at low-cytotoxic concentrations significantly promoted ROS generation and induced mitochondrial dysfunction. We further demonstrated that ZnO NPs could impair mitochondrial biogenesis and inhibit PGC-1α pathway. In addition, ZnO NPs at insignificantly cytotoxic concentrations were found to trigger cardiac electrophysiological alterations as evidenced by decreases of beat rate and spike amplitude. Conclusion: Our findings unveiled the potential harmful effects of ZnO NPs to human cardiomyocytes that involve mitochondrial biogenesis and the PGC-1α pathway that could affect cardiac electrophysiological function.


Assuntos
Coração/fisiopatologia , Células-Tronco Pluripotentes Induzidas/citologia , Miócitos Cardíacos/metabolismo , Nanopartículas/toxicidade , Biogênese de Organelas , Óxido de Zinco/toxicidade , Diferenciação Celular/efeitos dos fármacos , DNA Mitocondrial/genética , Coração/efeitos dos fármacos , Humanos , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Masculino , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Miócitos Cardíacos/efeitos dos fármacos , Nanopartículas/ultraestrutura , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo
17.
Life Sci ; 255: 117827, 2020 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-32450170

RESUMO

AIMS: Data suggest pharmacological treatment of depression with selective serotonin reuptake inhibitors (SSRI) may impair bone health. Our group has previously modeled compromised craniofacial healing after treatment with sertraline, a commonly prescribed SSRI, and hypothesized potential culprits: alterations in bone cells, collagen, and/or inflammation. Here we interrogate bone lineage cell alterations due to sertraline treatment as a potential cause of the noted compromised bone healing. MAIN METHODS: Murine pre-osteoblast, pre-osteoclast, osteoblast, and osteoclast cells were treated with clinically relevant concentrations of the SSRI. Studies focused on serotonin pathway targets, cell viability, apoptosis, differentiation, and the osteoblast/osteoclast feedback loop. KEY FINDINGS: All cells studied express neurotransmitters (e.g. serotonin transporter, SLC6A4, SSRI target) and G-protein-coupled receptors associated with the serotonin pathway. Osteoclasts presented the greatest native expression of Slc6a4 with all cell types exhibiting decreases in Slc6a4 expression after SSRI treatment. Pre-osteoclasts exhibited alteration to their differentiation pathway after treatment. Pre-osteoblasts and osteoclasts showed reduced apoptosis after treatment but showed no significant differences in functional assays. RANKL: OPG mRNA and protein ratios were decreased in the osteoblast lineage. Osteoclast lineage cells treated with sertraline demonstrated diminished TRAP positive cells when pre-exposed to sertraline prior to RANKL-induced differentiation. SIGNIFICANCE: These data suggest osteoclasts are a likely target of bone homeostasis disruption due to sertraline treatment, most potently through the osteoblast/clast feedback loop.


Assuntos
Osso e Ossos/efeitos dos fármacos , Osteoblastos/efeitos dos fármacos , Osteoclastos/efeitos dos fármacos , Inibidores de Captação de Serotonina/toxicidade , Células 3T3 , Animais , Apoptose/efeitos dos fármacos , Osso e Ossos/citologia , Diferenciação Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Camundongos , Osteoblastos/citologia , Osteoclastos/citologia , Ligante RANK/metabolismo , Células RAW 264.7 , RNA Mensageiro/metabolismo
18.
Cell Prolif ; 53(5): e12819, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32372504

RESUMO

OBJECTIVE: The objective of this study was to explore characteristics of bone marrow mesenchymal stromal cells (BM-MSCs) derived from patients with myelodysplastic syndrome (MDS) and multiple myeloma (MM). METHODS: BM-MSCs were recovered from 17 of MDS patients, 23 of MM patients and 9 healthy donors and were passaged until proliferation stopped. General characteristics and gene expression profiles of MSCs were analysed. In vitro, ex vivo coculture, immunohistochemistry and knockdown experiments were performed to verify gene expression changes. RESULTS: BM-MSCs failed to culture in 35.0% of patients and 50.0% of recovered BM-MSCs stopped to proliferate before passage 6. MDS- and MM-MSCs shared characteristics including decreased osteogenesis, increased angiogenesis and senescence-associated molecular pathways. In vitro and ex vivo experiments showed disease-specific changes such as neurogenic tendency in MDS-MSCs and cardiomyogenic tendency in MM-MSCs. Although the age of normal control was younger than patients and telomere length was shorter in patient's BM-MSCs, they were not different according to disease category nor degree of proliferation. Specifically, poorly proliferation BM-MSCs showed CDKN2A overexpression and CXCL12 downregulation. Immunohistochemistry of BM biopsy demonstrated that CDKN2A was intensely accumulation in perivascular BM-MSCs failed to culture. Interestingly, patient's BM-MSCs revealed improved proliferation activity after CDKN2A knockdown. CONCLUSION: These results collectively indicate that MDS-MSCs and MM-MSCs have common and different alterations at various degrees. Hence, it is necessary to evaluate their alteration status using representative markers such as CDKN2A expression.


Assuntos
Medula Óssea/patologia , Células-Tronco Mesenquimais/patologia , Mieloma Múltiplo/patologia , Síndromes Mielodisplásicas/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Medula Óssea/metabolismo , Células da Medula Óssea/metabolismo , Células da Medula Óssea/patologia , Diferenciação Celular/fisiologia , Proliferação de Células/fisiologia , Células Cultivadas , Quimiocina CXCL12/metabolismo , Feminino , Humanos , Masculino , Células-Tronco Mesenquimais/metabolismo , Pessoa de Meia-Idade , Mieloma Múltiplo/metabolismo , Síndromes Mielodisplásicas/metabolismo , Osteogênese/fisiologia , Adulto Jovem
19.
Cell Prolif ; 53(5): e12810, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32472648

RESUMO

OBJECTIVES: Gli1+ cells have received extensive attention in tissue homeostasis and injury mobilization. The aim of this study was to investigate whether Gli1+ cells respond to force and contribute to bone remodelling. MATERIALS AND METHODS: We established orthodontic tooth movement (OTM) model to assess the bone response for mechanical force. The transgenic mice were utilized to label and inhibit Gli1+ cells, respectively. Additionally, mice that conditional ablate Yes-associated protein (Yap) in Gli1+ cells were applied in the present study. The tooth movement and bone remodelling were analysed. RESULTS: We first found Gli1+ cells expressed in periodontal ligament (PDL). They were proliferated and differentiated into osteoblastic cells under tensile force. Next, both pharmacological and genetic Gli1 inhibition models were utilized to confirm that inhibition of Gli1+ cells led to arrest of bone remodelling. Furthermore, immunofluorescence staining identified classical mechanotransduction factor Yap expressed in Gli1+ cells and decreased after suppression of Gli1+ cells. Additionally, conditional ablation of Yap gene in Gli1+ cells inhibited the bone remodelling as well, suggesting Gli1+ cells are force-responsive cells. CONCLUSIONS: Our findings highlighted that Gli1+ cells in PDL directly respond to orthodontic force and further mediate bone remodelling, thus providing novel functional evidence in the mechanism of bone remodelling and first uncovering the mechanical responsive property of Gli1+ cells.


Assuntos
Remodelação Óssea/fisiologia , Osso e Ossos/metabolismo , Osso e Ossos/fisiologia , Proteína GLI1 em Dedos de Zinco/metabolismo , Animais , Diferenciação Celular/fisiologia , Mecanotransdução Celular/fisiologia , Camundongos , Camundongos Transgênicos , Osteoclastos/metabolismo , Osteoclastos/fisiologia , Ligamento Periodontal/metabolismo , Ligamento Periodontal/fisiologia , Estresse Mecânico , Técnicas de Movimentação Dentária/métodos
20.
Life Sci ; 253: 117728, 2020 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-32353430

RESUMO

AIMS: Studies indicate that the pattern of shear stress determines the direction of endothelial progenitor cells (EPCs) differentiation. However, the mechanism remains largely unknown. Herein, we try to identify the role of oscillatory shear stress (OSS) in the transdifferentiation of EPCs into mesenchymal cells and the mechanism involved. MATERIALS AND METHODS: OSS was applied to EPCs using the flow chamber system in vitro. Matrigel, Boyden chamber, and healing assay were used to observe the changes in EPCs function. Further, 2',7'-dichlorofluorescein diacetate (DCFH-DA) probe and/or western blot were performed to detect the expression of reactive oxygen species (ROS), p53 and PKCζ in EPCs. EPCs transduced with Lentivirus carrying Tp53 were implanted into the arterial vessel in the balloon injured rat model, and neointimal thickening was verified by HE staining. KEY FINDINGS: OSS enhanced the expression of mesenchymal cell markers alpha-smooth muscle actin (α-SMA) and smooth muscle 22 alpha (SM22α) on EPCs. In the meantime, OSS time-dependently decreased p53 expression in EPCs, which was partially abolished by treatment with ROS scavenger N-acetylcysteine (NAC) or protein kinase C zeta (PKCζ) inhibitor Go6983. Moreover, the p53 agonist tenovin-1 attenuated the changes of OSS-mediated the mesenchymal cell markers and EPCs function. Besides, we also found that transplanting EPCs transfected with LV-Tp53 significantly inhibited neointimal thickening and promoted reendothelialization in vivo. SIGNIFICANCE: This study demonstrates OSS-induced EPC transdifferentiation into mesenchymal cells and ROS/PKCζ/p53 pathway play an essential role in it. It may serve as a promising therapeutic target for cardiovascular disease in the future.


Assuntos
Células Progenitoras Endoteliais/citologia , Células-Tronco Mesenquimais/citologia , Espécies Reativas de Oxigênio/metabolismo , Estresse Mecânico , Acetanilidas/farmacologia , Animais , Diferenciação Celular/fisiologia , Transdiferenciação Celular/fisiologia , Células Cultivadas , Masculino , Proteína Quinase C/metabolismo , Ratos , Tioureia/análogos & derivados , Tioureia/farmacologia , Fatores de Tempo , Proteína Supressora de Tumor p53/metabolismo
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