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MC3T3-E1 Osteoblasts (mc3t3-e1 + osteoblast)
Selected AbstractsDevelopment of Electrospun Three-arm Star Poly(, -caprolactone) Meshes for Tissue Engineering ApplicationsMACROMOLECULAR BIOSCIENCE, Issue 8 2010Dario Puppi Abstract We have developed three-dimensional electrospun microfibrous meshes of a novel star branched three-arm poly(, -caprolactone) (*PCL) as potential scaffolds for tissue engineering applications. The processing conditions required to obtain uniform fibers were optimized by studying their influence on fiber morphology and size. Polymer molecular weight and solution feed rate influenced both the mesh microstructure and the tensile properties of the developed mats. Electrospun samples were also tested for their mechanical properties in wet conditions, showing higher yield strength and strain in comparison to that observed in dry conditions. Cell culture experiments employing MC3T3-E1 osteoblast like cells showed good cell viability adhesion and collagen production on the *PCL scaffolds. [source] Surface Structures and Osteoblast Activity on Biomedical Polytetrafluoroethylene Treated by Long-Pulse, High-Frequency Oxygen Plasma Immersion Ion ImplantationADVANCED ENGINEERING MATERIALS, Issue 5 2010Liping Tong Abstract Polytetrafluoroethylene (PTFE) is a biologically safe polymer used widely in clinical medicine including oral and orthopedic surgery. However, the high bio-inertness of PTFE has hampered wider applications in the biomedical fields. In this work, we extend the treatment time in long-pulse, high-frequency oxygen plasma immersion ion implantation of PTFE and a more superhydrophobic surface with a water contact angle of 160° is created. X-ray photoelectron spectroscopy (XPS) and atomic force microscopy (AFM) reveal that the optimized long-pulse, high-frequency oxygen plasma immersion ion implantation process induces a rougher surface and to a lesser extent alters the surface oxygen concentration on the PTFE. Our data, especially long-term contact angles, suggest that the superhydrophobility stems from surface roughness alteration. Furthermore, the activity of MC3T3-E1 osteoblasts cultured on the treated surfaces is promoted in terms of quantities and morphology. [source] The notch-responsive transcription factor Hes-1 attenuates osteocalcin promoter activity in osteoblastic cellsJOURNAL OF CELLULAR BIOCHEMISTRY, Issue 3 2009Ying Zhang Abstract Notch signaling plays a key role in osteoblast differentiation. A major transcriptional downstream regulator of this pathway is the helix,loop,helix (HLH) transcription factor Hairy/Enhancer of Split 1 (Hes-1). Here we investigated the function of Hes-1 in osteoblastic cells. Endogenous Hes-1 gene expression decreases during progression of bone cell phenotype development in MC3T3-E1 osteoblasts suggesting that it is a negative regulator of osteoblast differentiation. Forced expression of Hes-1 inhibits osteocalcin (OC) mRNA levels, and luciferase assays indicate that Hes-1 directly represses OC promoter activity. In vitro and in vivo protein/DNA interaction assays reveal that recombinant Hes-1 binds specifically to an E-box in the proximal promoter of the OC gene. Deletion of the Hes-1 WRPW domain (MHes-1) that recruits the co-repressor Groucho abrogates repression of OC promoter activity by Hes-1, but also blocks Hes-1 binding to the promoter. The latter result suggests that exogenous Hes-1 may be recruited to the OC promoter by both protein/DNA and protein/protein interactions. We conclude that the Notch-responsive Hes-1 protein is capable of repressing OC gene transcription in osteoblastic cells through an E-box in the proximal promoter. Hes-1 may contribute to osteoblast growth and differentiation by controlling basal bone-specific transcription directly through interactions with transcriptional regulators that are known to bind to the OC gene promoter. J. Cell. Biochem. 108: 651,659, 2009. © 2009 Wiley-Liss, Inc. [source] Characterization of the upstream mouse Cbfa1/Runx2 promoter,JOURNAL OF CELLULAR BIOCHEMISTRY, Issue 4 2001Z. S. Xiao Abstract Cbfa1 (or Runx2/AML-3/PEPB2,) is a transcriptional activator of osteoblastic differentiation. To investigate the regulation of Cbfa1 expression, we isolated and characterized a portion of the 5,-flanking region of the Cbfa1 gene containing its "bone-related" or P1 promoter and exon 1. We identified additional coding sequence in exon 1 and splice donor sites that potentially give rise to a novel Cbfa1 isoform containing an 18 amino acid insert. In addition, primer extension mapping identified in the Cbfa1 promoter a minor mRNA start site located ,0.8 kb 5, upstream of the ATG encoding the MASN/p57 isoform and ,0.4 kb upstream of the previously reported start site. A luciferase reporter construct containing 1.4 kb of the mouse Cbfa1 promoter was analyzed in Ros 17/2.8 and MC3T3-E1 osteoblast cell lines that express high levels of Cbfa1 transcripts. The activity of this construct was also examined in non-osteoblastic Cos-7 and NIH3T3 cells that do not express Cbfa1 and mesenchymal-derived cell lines, including CH3T101/2, C2C12, and L929 cells, that express low levels of mature Cbfa1 transcripts. The 1.4 kb 5, flanking sequence of the Cbfa1 gene directed high levels of transcriptional activity in Ros 17/2.8 and MC3T3-E1 osteoblasts compared to non-osteoblasts Cos-7 cells, but this construct also exhibited high levels of expression in C310T1/2, L929, and C2C12 cells as well as NIH3T3 cells. In addition, Cbfa1 mRNA expression, but not the activity of the Cbfa1 promoter, was upregulated in a dose-dependent manner in pluripotent mesenchymal C2C12 by bone morphogenetic protein-2 (BMP-2). These data indicate that Cbfa1 is expressed in osteogenic as well as non-osteogenic cells and that the regulation of Cbfa1 expression is complex, possibly involving both transcriptional and post-transcriptional mechanisms. Additional studies are needed to further characterize important regulatory elements and to identify additional regions of the promoter and/or post-transcriptional events responsible for the cell-type restricted regulation of Cbfa1 expression. J. Cell. Biochem. 82: 647,659, 2001. © 2001 Wiley-Liss, Inc. [source] Impaired cell cycle regulation of the osteoblast-related heterodimeric transcription factor Runx2-Cbf, in osteosarcoma cellsJOURNAL OF CELLULAR PHYSIOLOGY, Issue 3 2009Inga A. San Martin Bone formation and osteoblast differentiation require the functional expression of the Runx2/Cbf, heterodimeric transcription factor complex. Runx2 is also a suppressor of proliferation in osteoblasts by attenuating cell cycle progression in G1. Runx2 levels are modulated during the cell cycle, which are maximal in G1 and minimal beyond the G1/S phase transition (S, G2, and M phases). It is not known whether Cbf, gene expression is cell cycle controlled in preosteoblasts nor how Runx2 or Cbf, are regulated during the cell cycle in bone cancer cells. We investigated Runx2 and Cbf, gene expression during cell cycle progression in MC3T3-E1 osteoblasts, as well as ROS17/2.8 and SaOS-2 osteosarcoma cells. Runx2 protein levels are reduced as expected in MC3T3-E1 cells arrested in late G1 (by mimosine) or M phase (by nocodazole), but not in cell cycle arrested osteosarcoma cells. Cbf, protein levels are cell cycle independent in both osteoblasts and osteosarcoma cells. In synchronized MC3T3-E1 osteoblasts progressing from late G1 or mitosis, Runx2 levels but not Cbf, levels are cell cycle regulated. However, both factors are constitutively elevated throughout the cell cycle in osteosarcoma cells. Proteasome inhibition by MG132 stabilizes Runx2 protein levels in late G1 and S in MC3T3-E1 cells, but not in ROS17/2.8 and SaOS-2 osteosarcoma cells. Thus, proteasomal degradation of Runx2 is deregulated in osteosarcoma cells. We propose that cell cycle control of Runx2 gene expression is impaired in osteosarcomas and that this deregulation may contribute to the pathogenesis of osteosarcoma. J. Cell. Physiol. 221: 560,571, 2009. © 2009 Wiley-Liss, Inc. [source] Pulsating fluid flow modulates gene expression of proteins involved in Wnt signaling pathways in osteocytesJOURNAL OF ORTHOPAEDIC RESEARCH, Issue 10 2009Ana Santos Abstract Strain-derived flow of interstitial fluid activates signal transduction pathways in osteocytes that regulate bone mechanical adaptation. Wnts are involved in this process, but whether mechanical loading modulates Wnt signaling in osteocytes is unclear. We assessed whether mechanical stimulation by pulsating fluid flow (PFF) leads to functional Wnt production, and whether nitric oxide (NO) is important for activation of the canonical Wnt signaling pathway in MLO-Y4 osteocytes. MC3T3-E1 osteoblasts were studied as a positive control for the MLO-Y4 osteocyte response to mechanical loading. MLO-Y4 osteocytes and MC3T3-E1 osteoblasts were submitted to 1-h PFF (0.7,±,0.3 Pa, 5 Hz), and postincubated (PI) without PFF for 0.5,3 h. Gene expression of proteins related to the Wnt canonical and noncanonical pathways were studied using real-time polymerase chain reaction (PCR). In MLO-Y4 osteocytes, PFF upregulated gene expression of Wnt3a, c-jun, connexin 43, and CD44 at 1,3-h PI. In MC3T3-E1 osteoblasts, PFF downregulated gene expression of Wnt5a and c-jun at 0.5,3-h PI. In MLO-Y4 osteocytes, gene expression of PFF-induced Wnt target genes was suppressed by the Wnt antagonist sFRP4, suggesting that loading activates the Wnt canonical pathway through functional Wnt production. The NO inhibitor L-NAME suppressed the effect of PFF on gene expression of Wnt target genes, suggesting that NO might play a role in PFF-induced Wnt production. The response to PFF differed in MC3T3-E1 osteoblasts. Because Wnt signaling is important for bone mass regulation, osteocytes might orchestrate loading-induced bone remodeling through, among others, Wnts. © 2009 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 27:1280,1287, 2009 [source] Osteoblast Adhesion and Proliferation on Poly(3-octylthiophene) Thin FilmsMACROMOLECULAR BIOSCIENCE, Issue 3 2010Charlene Rincón Abstract In this study we assessed the suitability of semiconducting P3OT thin films (30,nm) to sustain attachment, spreading, and proliferation of MC3T3-E1 osteoblasts. Cell area correlated with surface wettability: area was larger on the more hydrophilic surface (TCPS) and lower on the more hydrophobic surface (P3OT). Cells were rounder, characterized by higher circularity values, on TCPS and Si compared to P3OT. P3OT proliferation rate at 24,h fell twofold after 48,h, then recovered at 72,h to a value significantly higher than that on TCPS. Presoaking experiments showed no evidence of cytotoxic effects or leachants from P3OT. Overall, we conclude that P3OT is a viable substrate for osteoblast attachment and proliferation. [source] | |||||||||||||