Growth Of MG-63 Research Articles

Fetuin A functionalisation of biodegradable PLLA-co-PEG nonwovens towards enhanced biomineralisation and osteoblastic growth behaviour.

Therapy for large-scale bone defects remains a major challenge in regenerative medicine. In this context, biodegradable electrospun nonwovens are a promising material to be applied as a temporary implantable scaffold as their fibre diameters are in the micro- and nanometre range and possess a high surface-to-volume ratio paired with high porosity. In this work, in vitro assessment of biodegradable PLLA-co-PEG nonwovens with fetuin A covalently anchored to the surface has been performed in terms of biomineralisation and the influence on MG-63 osteoblast cell metabolic activity, biosynthesis of type I collagen propeptide and inflammatory potential. Our finding was that covalent fetuin A funtionalisation of the nonwoven material leads to a distinct increase in calcium affinity, thus enhancing biomineralisation while maintaining the distinct fibre morphology of the nonwoven. The cell seeding experiments showed that the fetuin A functionalised and subsequently in vitro biomineralised PLLA-co-PEG nonwovens did not show negative effects on MG-63 growth. Fetuin A funtionalisation and enhanced biomineralisation supported cell attachment, leading to improved cell morphology, spreading and infiltration into the material. Furthermore, no signs of increase in the inflammatory potential of the material have been detected by flow cytometry experiments. Overall, this study provides a contribution towards the development of artificial scaffolds for guided bone regeneration with the potential to enhance osteoinduction and osteogenesis.

Silencing tumor-intrinsic HHLA2 potentiates the anti-tumoral effect of paclitaxel on MG63 cells: Another side of immune checkpoint

BackgroundOsteosarcoma is common type of bone cancer; however, the prognosis of patients with metastatic osteosarcoma is poor. As a new inhibitory immune checkpoint molecule, HHLA2 is upregulated in osteosarcoma. Herein, we studied the significance of tumor-intrinsic HHLA2 in MG-63 growth. Also, we examined the influence of combined therapy of HHLA2 knockdown with paclitaxel on the apoptosis, cell cycle, migration, and stemness of MG-63 cells. MethodsThe 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay was performed to study the half-maximal inhibitory concentration (IC50) of paclitaxel and the cytotoxicity of HHLA2-small interfering RNA (siRNA) on MG-63 cells. The apoptosis and cell cycle were analyzed using flow cytometry. The wound-healing and colony formation assays were conducted to investigate the effect of paclitaxel and HHLA2 knockdown on the migration and stemness of MG-63 cells, respectively. QRT-PCR was used to determine the Bax, caspase-3, and Bcl-2 mRNA expression levels. ResultsHHLA2 silencing has enhanced the chemosensitivity of MG-63 cells to paclitaxel. Besides, HHLA2 knockdown has increased the paclitaxel-induced cytotoxic effect on MG-63 cells. In terms of stimulating apoptosis, decreasing clonogenicity, halting the cell cycle at the sub G1 phase, and inhibiting migration, tumor-intrinsic HHLA2 silencing has increased these anti-tumor effects of paclitaxel on MG-63 cells. Besides, HHLA2 knockdown has potentiated paclitaxel-mediated Bcl-2 downregulation and paclitaxel-mediated caspase-3 and Bax upregulation in MG-63 cells. ConclusionTumor-intrinsic HHLA2 knockdown increases the anti-tumoral effect of paclitaxel on MG-63 cells and enhances the chemosensitivity of MG-63 cells to paclitaxel.

Strontium-Doped Hydroxyapatite Nanowires with a Hierarchical Interface: Enhancing Osteoinduction and Cellular Attachment

Strontium-doped hydroxyapatite nanowires with hierarchical interfaces were prepared in this study by a one-step hydrothermal method using Pluronic123 as a structural guide. The cytotoxicity and biocompatibility of the materials were detected by the MTT assay. The adhesion and growth of MG63 (osteosarcoma cells) on materials with different doping ratios were observed by scanning electron microscopy and fluorescence microscopy. Real-time polymerase chain reaction (RT-PCR) was used to detect the transcription levels of alkaline phosphatase (ALP), Runt-associated transcription factor 2 (Runx2), osteopontin (OPN), type I collagen (Coll-I) and osteocalcin (OCN). The protein expression levels of ALP, Runx2, OPN, Col-I and OCN were detected by western blotting (WB). The results showed that strontium-doped hydroxyapatite nanowires had good biocompatibility and osteogenic induction ability, and the strontium-doped hydroxyapatite nanowires with a 1% strontium content exhibited a better osteogenic induction performance. Therefore, it is expected to become a new, promising bone induction scaffold material.

Abstract B10: Modulation of mRNA translation regulation in highly metastatic ssteosarcoma cells increases their sensitivity to redox stress

Abstract Improvement in outcomes for pediatric patients with metastatic osteosarcoma still remains elusive despite the development of new multiagent combinations. This unmet clinical need underscores the need for novel approaches that examine the metastatic process itself in order to identify new molecular targets whose modulation may have antimetastatic activity. During lung metastasis progression, osteosarcoma cells (OS) have to quickly adapt to the hostile microenvironment of the lung. Elucidating exactly how highly aggressive metastatic OS cells adapt to the lung microenvironment is the subject of the current work. Our research utilizes a pair of clonally related human OS cell lines, MG63.3 and MG63, with highly or poorly metastatic in vivo phenotypes, respectively. These OS cells experience redox stress in the lung microenvironment, as evident by in situ immunoreactivity with 3-nitrotyrosine (3-NT)---a marker of oxidative damage. In cell culture studies where high and poorly metastatic OS cells are exposed to PABA/NO, a chemical inducer of redox stress which also causes 3-NT accumulation, highly metastatic MG63.3 cells show functional differences in their response to redox stress compared to poorly metastatic MG63 cells. MG63.3 cells show lower levels of 3-NT staining, lower levels of caspase 3/7 activity, and have higher growth rates compared to MG63 cells in the presence of PABA/NO. These results suggest that MG63.3 cells show a greater adaptability to redox stress compared to MG63 cells. It is known that regulation of mRNA translation is a mechanism by which cancer cells can quickly adapt to the changing extracellular milieu. Furthermore, it has been shown that MG63.3 cells are able to translate weak mRNAs more efficiently than MG63 cells under stressful conditions. Such weak mRNAs encode for proteins involved in growth and proliferation. When we examine the expression levels of eukaryotic initiation factors such as eIF4E, eIF4G, and eIF4A, all of which are part of the eIF4F cap-initiation complex, we find that certain factors (such as eIF4G1) are differentially upregulated (both at the transcript and protein level) in MG63.3 cells compared to MG63 cells. We hypothesize that inhibition of eIF4G binding to the eIF4F complex will reduce the ability of MG63.3 cell to translate mRNAs important for survival and proliferation during redox stress. To address this hypothesis, we tested whether a small-molecule inhibitor of eIF4G/eIF4E interactions, called 4EGI-1, can sensitize highly metastatic MG63.3 cells to PABA/NO. When exposed to noncytotoxic levels of 4EGI-1 (≤ 20 μM), we find that 4EGI-1 can sensitize MG63.3 cells with lower concentrations of PABA/NO compared to control groups. 4EGI-1 treatment, in the presence of PABA/NO, can cause MG63.3 cells to accumulate higher levels of 3-NT compared to control groups. Furthermore, using an Incucyte machine to assess cell proliferation over 5 days, we find that combination treatment (4EGI-1 and PABA/NO) can greatly inhibit the growth of MG63.3 cells. These results suggest that certain mRNA transcripts that are important in adapting to redox stress are dependent on the eIF4F cap-initiation complex. Efforts are currently under way to further characterize which gene transcripts are important to redox stress adaptation. In addition, by using the pulmonary metastasis assay, we will be assessing whether 4EGI-1 has antimetastatic activity. Collectively, the data presented in the current work suggest that the adaptive mechanisms that metastatic OS cells use to manage redox stress may be an attractive therapeutic target in the development of novel antimetastatic therapeutics. Citation Format: Michael M. Lizardo, Poul Sorensen. Modulation of mRNA translation regulation in highly metastatic ssteosarcoma cells increases their sensitivity to redox stress [abstract]. In: Proceedings of the AACR Conference on Advances in Sarcomas: From Basic Science to Clinical Translation; May 16-19, 2017; Philadelphia, PA. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(2_Suppl):Abstract nr B10.

Powder Mixed Electric Discharge Machining: An Innovative Surface Modification Technique to Enhance Fatigue Performance and Bioactivity of β-Ti Implant for Orthopedics Application

The development of surface modification technique has been the subject of the studies regarding the fatigue performance and biological characterization of the modified layers. In the present research work, powder mixed electric discharge machining (PMEDM) a novel nonconventional machining technique has been proposed for surface modification of β-Ti implant for orthopedics application. The surface topography and morphology like roughness, surface cracks, and recast layer thickness of each of the machined specimens were investigated using Mitutoyo surface roughness tester and field-emission scanning electron microscopy (FE-SEM), respectively. This study aims to investigate the effect of surface characteristics of PMEDM process on the fatigue performance and bioactivity of β-Ti implants and moreover a comparative analysis is made on the fatigue performance and biological activity of specimens machined with presently used machining methods like electric discharge machining (EDM) and mechanical polishing. The high cycle fatigue (HCF) performance of polished specimens was superior and had no adverse effect of microstructure on fatigue endurance. As expected, the fatigue behavior of β-Ti implant-based alloy, after undergoing EDM treatment, is poorly observed due to the microrough surface. The fatigue performance is dependent on microstructure and surface roughness of the specimens. Subsequent PMEDM process significantly improves the fatigue endurance of β-Ti implant-based alloy specimens. PMEDMed surface with micro-, sub-micro-, and nano-structured topography exhibited excellent bioactivity and improved biocompatibility. PMEDMed surface enabled better adhesion and growth of MG-63 when compared with the polished and EDMed substrate. Furthermore, the differentiation results indicated that a combination of nanoscale featured submicrorough PMEDMed surface promotes various osteoblast differentiation activities like alkaline phosphatase (ALP) activity, osteocalcin production, the local factor osteoprotegerin, which inhibits osteoclastogenesis.

Green tea polyphenol EGCG suppresses osteosarcoma cell growth through upregulating miR-1.

(-)-Epigallocatechin-3-gallate (EGCG), the most abundant and active polyphenol in green tea, has been demonstrated to have anticancer effects in a wide variety of human cancer. MicroRNAs (miRNAs) are a class of short noncoding RNAs and play important role in gene regulation and are critically involved in the pathogenesis and progression of human cancer. This study aims to investigate the effects of EGCG on osteosarcoma (OS) cells and elucidate the underlying mechanism. Cellular function assays revealed that EGCG inhibited cell proliferation, induced cell cycle arrest and promoted apoptosis of OS cells in vitro, and also inhibited the growth of transplanted tumors in vivo. By miRNA microarray and RT-qPCR analysis, miR-1 was found to be significantly upregulated in MG-63 and U-2OS treated by EGCG in dose- and time-dependent manners, and miR-1 downregulation by inhibitor mimics attenuated EGCG-induced inhibition on cell growth of OS cells. We also confirmed that miR-1 was also frequently decreased in clinical OS tumor tissues. Moreover, both EGCG and miR-1 mimic inhibited c-MET expression, and combination treatment with EGCG and c-MET inhibitor (crizotinib) had enhanced inhibitory effects on the growth of MG-63 and U-2OS cells. Taken together, these results suggest that EGCG has an anticancer effect on OS cells, at least partially, through regulating miR-1/c-MET interaction.

Processing and Characterization of Novel Biomimetic Nanoporous Bioceramic Surface on β-Ti Implant by Powder Mixed Electric Discharge Machining

Herein, a β-Ti-based implant was subjected to powder mixed electric discharge machining (PMEDM) for surface modification to produce a novel biomimetic nanoporous bioceramic surface. The microstructure, surface topography, and phase composition of the non-machined and machined (PMEDMed) surfaces were investigated using field-emission scanning electron microscopy, energy-dispersive x-ray spectroscopy, and x-ray diffraction. The microhardness of the surfaces was measured on a Vickers hardness tester. The corrosion resistance of the surfaces was evaluated via potentiodynamic polarization measurements in simulated body fluid. The application of PMEDM not only altered the surface chemistry, but also imparted the surface with a nanoporous topography or a natural bone-like surface structure. The characterization results confirmed that the alloyed layer mainly comprised bioceramic oxides and carbide phases (TiO2, Nb2O5, ZrO2, SiO2, TiC, NbC, SiC). The microhardness of PMEDMed surface was twofold higher than that of the base material (β-Ti alloy), primarily because of the formation of the hard carbide phases on the machined layer. Electrochemical analysis revealed that PMEDMed surface featured insulative and protective properties and thus displayed higher corrosion resistance ability when compared with the non-machined surface. This result was attributed to the formation of the bioceramic oxides on the machined surface. Additionally, the in vitro biocompatibility of the surfaces was evaluated using human osteoblastic cell line MG-63. PMEDMed surface with a micro-, sub-micro-, and nano-structured topography exhibited bioactivity and improved biocompatibility relative to β-Ti surface. Furthermore, PMEDMed surface enabled better adhesion and growth of MG-63 when compared with the non-machined substrate.

Inhibitory effects of curcumin on growth of human osteosarcoma cell line MG-63 cells

Objective To investigate the inhibitory effects of curcumin (Cur) on proliferation and apopotosis of human osteosarcoma cell line MG-63 cells.Methods The cultured MG-63 cells were treated with Cur (12.5,25.0,50.0,100.0 μmol/L) for 12 h.The morphologic changes of MG-63 cells were observed by inverted microscope.The survival rate of MG-63 cells was measured by using methyl thiazol tetrazolium (MTT) assay.Cell counting of kit-8 (CCK-8) assay was used to test the growth inhibitory rate.Annexin V/propidium iodide (PI) staining was applied to detect the apoptosis.The survival ratios and the adhesion capacity of MG-63 were detected except the lactate dehydrogenase (LDH) releasing.Results Different concentrations of Cur (12.5,25.0,50.0,100.0 μmol/L) observed in this study have significant effects on the survival ratio of MG-63.The Cur could significantly inhibit the growth of MG-63 in a dose and time dependent manner and induce G2/M Phase cell cycle arrest.After 12 h,cell inhibition rate of 47.23％,24 h after the cell was 65.23％.Cur 12.5,25.0,50.0,100.0 μmol/L group for 24 h,MG-63 cell apoptosis rate was (11.24 ± 0.07)％,(28.23 ± 0.35)％,(50.21 ± 1.71)％ and (41.22 ±1.85) ％,50.0 μmol/L curcumin could induce apoptosis obviously.Conclusion When Cur increased from 12.5 μmol/L to 25.0 μmol/L,its MG-63 cells at 24h growth inhibition rate (13.56 ± 1.88) ％ to (48.27 ±2.32)％,50.0 μmol/L group 24 h growth inhibition rate to 65.23％.50.0 μmol/L of Cur on the MG-63 cells apoptosis induced by the most obvious,for 24 h,apoptosis rate was (50.21 ± 1.71)％. Key words: Curcumin ; Osteosarcoma; Proliferation

A novel ruthenium(II) complex inhibits cell proliferation and induces apoptosis in MG-63 cells

Objective To investigate the effects of a novel ruthenium(Ⅱ) polypyridyl complex △-[Ru (phen)2MCMIP]2 + (△-1) on proliferation and apoptosis of human osteosarcoma cell line MG-63 in vitro.Methods Cell counting of kit-8 (CCK-8) assay was used to detect the proliferation of MG-63 cells after 24,48,72h treatment with △-1 under the concentrations of 0.00,12.50,25.00,50.00,100.00,150.00 μmol/L;Changes of apoptosis and cell cycle in MG-63 cells after 24 h treatment of 0.00,25.00,50.00,100.00 μmol/L A-1 were determined and analyzed by flow cytometry (FCM).Results Ruthenium (Ⅱ) polypyridyl complex A-1 could significantly inhibit the growth of MG-63 in a dose and time dependent manner; the IC50 of 24 h,48 h,72 h was 57.80μmol/L,45.27μmol/L,32.51μmol/L respectively; flow cytometry detection showed that A-1 induced 37.10％ of apoptosis,while only 1.06％ in control group,and arrested cell cycle at G0/G1 phase.Conclusion Ruthenium(Ⅱ) polypyridyl complex A-1 is able to inhibit proliferation and induce apoptosis in MG-63 cells and arrest cell cycle at G0/G1 phase. Key words: Ruthenium(Ⅱ) complex; MG-63 cells; Proliferation; Apoptosis; Cell cycle

Effects of Garlicin on Proliferation and Apoptosis of MG-63 Cell Line

Objective To investigate the effects of garlicin on proliferation and apopotosis of human osteosarcoma cell line MG-63.Methods Under different garlicin concentrations,the MG-63 cells were observed by inverted microscope;cell counting of kit-8(CCK-8) assay was used to detect the growth inhibitory rates;and apoptosis was determined by acridine orange/ethidium bromide(AO/EB) double staining and Annexin V-FITC staining flow cytometry(FCM).Cell Cycle was analyzed by FCM.Results Garlicin could significantly inhibit the growth of MG-63 in a dose and time dependent manner and induce G2/M phase cell cycle arrest.10 μg/ml garlicin could induce apoptosis obviously and arrest cell cycle at G2/M phase.Conclusion Garlicin was able to inhibit proliferation and induce apoptosis in MG-63 cells.

느릅나무 근피 추출물에 의한 인체 암세포 증식 및 DNA 합성 억제효과

인체 암세포계(MG-63 골육암 세포, HT-29 인체 결장암세포, K-562 백혈암세포)를 이용하여 느릅나무 근피 메탄올 추출물, 열탕 추출물 및 즙액에 의한 암세포 성장에 미치는 효과를 검토하였다. 느릅나무 근피 메탄올 추출물, 열탕 추출물 및 즙액은 낮은 농도에서부터 인체 골육암 세포의 증식을 억제시켰다. 인체 결장암세포와 백혈암세포에서도 느릅나무 근피 메탄올 추출물, 열탕 추출물 및 즙액은 낮은 농도인 1 mg/ml에서부터 활성을 나타내어 40% 이상으로 암세포 증식 억제효과를 나타내어 앞서의 MG-63 골육암 세포에서 보다 그 증식 억제효과가 높은 것으로 나타났다. 느릅나무 근피 메탄올 추출물, 열탕 추출물 및 즙액을 골육암과 결장암세포에 투여한 2일 후에 세포내의 DNA 합성에 미치는 영향을 측정한 결과, 농도 의존적으로 암세포의 DNA 합성을 저해하는 것을 살펴 볼 수가 있었다. 따라서 느릅나무 근피 추출물은 인체 암세포 증식을 크게 억제하였으며 열탕 추출물에서도 항암활성 효과를 보여 느릅나무 근피 유래 생리활성 물질은 열에 안정한 것으로 여겨진다. Growth and DNA synthesis inhibitory effects of extracts from root bark of Ulmus parvifolia on MG-63 human osteosarcoma cells, HT-29 human colon cancer cells and K-562 leukemia cancer cells were studied. The root bark extract of Ulmus parvifolia was extracted with methanol, hot water and juice. The methanol extract showed the highest inhibitory effect on growth of MG-63, HT-29 and K-562 cancer cells by >85%. The treatment of hot water and juice extracts from root bark of Ulmus parvifolia also inhibited growth of the above cancer cells with increasing concentration. DNA synthesis of MG-63 and HT-29 cancer cells was significantly inhibited by adding methanol, hot water and juice extracts from root bark of Ulmus parvifolia with increasing concentration, showing that the inhibitory effect of growth was more effective on HT-29 cancer cells. These results suggest that the methanol extract from root bark of Ulmus parvifolia may have specific active com-pounds on anticancer effect. The hot water extract also showed a strong inhibitory effect on growth of cancer cells, indicating that the active compounds may be stable to heat.