共查询到10条相似文献，搜索用时 31 毫秒
1.
YASUYOSHI TORII KIYOTAKA HITOMI YASUKO YAMAGISHI NORIHIRO TSUKAGOSHI 《Cell biology international》1996,20(7):459-464
MC3T3-E1 cells grown with ascorbic acid express sequentially osteoblastic marker proteins such as alkaline phosphatase (ALPase) and then form a mineralized extracellular matrix (ECM) as a consequence of osteoblastic differentiation. To explore the functional roles of ALPase in the process of osteoblastic maturation, an inducible expression vector for antisense ALPase RNA was constructed and stably transfected into MC3T3-E1 cells. The expression of antisense ALPase RNA in the differentiated MC3T3-E1 transfectants reduced markedly the ALPase activity, which resulted in a significant decrease in the deposition of minerals upon prolonged culture. These findings demonstrated directly that ALPase participated in the mineralizationof ECM. 相似文献
2.
Beck GR 《Journal of cellular biochemistry》2003,90(2):234-243
The spatial and temporal coordination of the many events required for osteogenic cells to create a mineralized matrix are only partially understood. The complexity of this process, and the nature of the final product, demand that these cells have mechanisms to carefully monitor events in the extracellular environment and have the ability to respond through cellular and molecular changes. The generation of inorganic phosphate during the process of differentiation may be one such signal. In addition to the requirement of inorganic phosphate as a component of hydroxyapatite mineral, Ca(10)(PO(4))(6)(OH)(2), a number of studies have also suggested it is required in the events preceding mineralization. However, contrasting results, physiological relevance, and the lack of a clear mechanism(s) have created some debate as to the significance of elevated phosphate in the differentiation process. More recently, a number of studies have begun to shed light on possible cellular and molecular consequences of elevated intracellular inorganic phosphate. These results suggest a model in which the generation of inorganic phosphate during osteoblast differentiation may in and of itself represent a signal capable of facilitating the temporal coordination of expression and regulation of multiple factors necessary for mineralization. The regulation of protein function and gene expression by elevated inorganic phosphate during osteoblast differentiation may represent a mechanism by which mineralizing cells monitor and respond to the changing extracellular environment. 相似文献
3.
A fibronectin fragment, which spans from the ninth to tenth type III domain, showed > 95% biological activity in promoting osteoblast adhesion, proliferation and differentiation level compared with those of native fibronectin (FN) molecule. This suggests that the FN type III ninth-tenth domain fragment serves a good substitute for native FN in biomimetic application for osteogenesis. 相似文献
4.
Pulsed electromagnetic field (PEMF) devices have been used clinically to promote the healing of surgically resistant fractures in vivo. However, there is a sparsity of data on how the timing of an applied PEMF effects the osteogenic cells that would be present within the fracture gap. The purpose of this study was to examine the response of osteoblast-like cells to a PEMF stimulus, mimicking that of a clinically available device, using four protocols for the timing of the stimulus. The PEMF signal consisted of a 5 ms pulse burst (containing 20 pulses) repeated at 15 Hz. Cultures of a human osteosarcoma cell line, SaOS-2, were exposed to the four timing protocols, each conducted over 3 days. Protocol one stimulated the cells for 8 h each day, protocol two stimulated the cells for 24 h on the first day, protocol three stimulated the cells for 24 h on the second day, and protocol four stimulated the cells for 24 h on the third day. Cells were seeded with either 25,000 or 50,000 cells/well (24-well cell culture plates). All assays showed reduced proliferation and increased differentiation (alkaline phosphatase activity) in the PEMF stimulated cultures compared with the control cultures, except for protocol four alkaline phosphatase measurements. No clear trend was observed between the four protocols; however this may be due to cell density. The results indicated that an osteoblast-like cell line is responsive to a 15 Hz PEMF stimulus, which will stimulate the cell line to into an increasing state of maturity. 相似文献
5.
Three placental alkaline phosphatases purified to homogeneity, i.e., the F, I, and S variants, were investigated for catalytic and stability properties. All three forms of the enzyme were found to have almost identical pH optima (10.7–10.8), similar sensitivity to the uncompetitive inhibitors L-phenylalanine (70%) and L-leucine (30%), and identical Km values against p-nitrophenylphosphate, -glycerophosphate, and -naphthylphosphate. Significant differences among the three types were observed in thermal stability. The F variant was found to be most stable and the I variant most labile at 79 C. At 70 C all three forms were stable.This investigation was supported by grants from the Swedish Medical Research Council (Projects No. 4217 and 03X-2725), from the Medical Faculty, University of Umeå, and Jubileumsklinikens i Umeå forskningsfond. 相似文献
6.
Role of metallothionein isoforms in bone formation processes in rat marrow mesenchymal stem cells in culture 总被引：1，自引：0，他引：1
Dohi Y Shimaoka H Ikeuchi M Ohgushi H Yonemasu K Minami T 《Biological trace element research》2005,104(1):57-69
Temporal changes in mRNAs for metallothionein (MT) isoforms in subcultures of rat marrow mesenchymal stem cells (MSCs) after
treatment with dexamethasone were investigated. Both MT-1 and MT-2 mRNA expression in the cultured MSCs with dexamethasone
showed maximum levels at d 1, whereas ALP and osteocalcin mRNAs peaked at d 12. MT-3 mRNA was not detected in the cultured
MSCs at any time. The expression level of MT-2 mRNA at d 1 was 9.4-fold higher than that of MT-1 mRNA. Finally, osteoblast
differentiation and mineralization of MSCs at d 14 was inhibited by the addition of a common antisense oligonucleotide for
both MT-1 and MT-2 in the culture medium during the first 4 d. The results suggest that the large amounts of MT-2 are produced
in the early stage of subculture of MSCs, and this might regulate their differentiation. 相似文献
7.
Herrmann M Umanskaya N Wildemann B Colaianni G Widmann T Zallone A Herrmann W 《Journal of cellular and molecular medicine》2008,12(4):1205-1210
Homocysteine (HCY) has recently been linked to fragility fractures. Moreover, HCY activates osteoclasts. Little is known about the effect of HCY on activity of human osteoblasts (OBs). We hypothesized that HCY decreases the activity of OBs. Osteoblasts obtained from tra-becular human bone specimens of eight donors were cultured with conditioned medium. Culture medium was adjusted to 0, 100, 500, 1000 and 2000 muM HCY. After 14 days alkaline phosphatase (AP) activity, pro-collagen type I N-terminal peptide (PINP) and osteocalcin (OC) secretion in the supernatant were measured. After 20 days the formation of mineralized matrix was analyzed. HCY-stimulated AP activity gradually (100 muM HCY: 118%, P= 0.006; 500 muM HCY: 125%, P < 0.001). At 1000 and 2000 muM HCY the increase of AP activity was reversible (1000 muM HCY: 106%, P= 0.317; 2000 muM HCY: 102%, P < 0.737). The PINP secretion was also stimulated by HCY reaching a maximum of 260 +/- 154 mug/l at 500 mumol/l versus 205 +/- 94 mu,g/l in controls. After 20 days of culture the formation of bone matrix was increased at 100 and 500 muM HCY. OC secretion was not significantly changed. The results of the present study consistently demonstrate a moderate stimulation of primary human OB activity by increasing concentrations of HCY. However, the magnitude of this effect seems to be less pronounced than recent observations on primary human osteoclasts, suggesting a dysbalance between OBs and osteoclasts in favour of osteoclasts. 相似文献
8.
Dina Abdallah Najwa Skafi Eva Hamade Mathieu Borel Sophie Reibel Nicolas Vitale Alaeddine El Jamal Carole Bougault Norbert Laroche Laurence Vico Bassam Badran Nader Hussein David Magne Rene Buchet Leyre Brizuela Saida Mebarek 《Journal of cellular biochemistry》2019,120(4):5923-5935
Mammalian phospholipase D (PLD) mostly hydrolyzes phosphatidylcholine producing phosphatidic acid. PLD activity was previously detected in different osteoblastic cell models, and was increased by several growth factors involved in bone homeostasis. To confirm possible actions of PLD isoforms during mineralization process, we analyzed their effects in osteoblastic cell models and during bone formation. PLD1 expression, along with PLD activity, increased during differentiation of primary osteoblasts and Saos-2 cells, and peaked at the onset of mineralization. Subsequently, both PLD1 expression and PLD activity decreased, suggesting that PLD1 function is regulated during osteoblast maturation. In contrast, PLD2 expression was not significantly affected during differentiation of osteoblasts. Overexpression of PLD1 in Saos-2 cells improved their mineralization potential. PLD inhibitor Halopemide or PLD1-selective inhibitor, led to a decrease in mineralization in both cell types. On the contrary, the selective inhibitor of PLD2, did not affect the mineralization process. Moreover, primary osteoblasts isolated from PLD1 knockout (KO) mice were significantly less efficient in mineralization as compared with those isolated from wild type (WT) or PLD2 KO mice. In contrast, bone formation, as monitored by high-resolution microcomputed tomography analysis, was not impaired in PLD1 KO nor in PLD2 KO mice, indicating that the lack of PLD1 or that of PLD2 did not affect the bone structure in adult mice. Taken together, our findings indicate that PLD activity, especially which of PLD1 isoform, may enhance the mineralization process in osteoblastic cells. Nonetheless, the lack of PLD1 or PLD2 do not seem to significantly affect bone formation in adult mice. 相似文献
9.
Fragments of cancellous and cortical bone from human maxilla and mandible were cultured by the explant technique. Cells isolated by trypsinization of primary cultures were characterized as osteoblasts on the basis of intracellular alkaline phosphatase activity, the constituents of the extracellular matrix, and response to human parathormone (PTH). In culture, the osteoblasts often gave rise to superposed clumps of large cells whose cytoplasm contained endoplasmic reticulum, numerous mitochondria, vacuoles, and a dense network of intermediate filaments, often at the level of the plasma membrane. In the presence of vitamin C and 1,25-dihydroxyvitamin D3, the osteoblasts produced an extracellular matrix composed of collagen type I and various non-collagenous proteins, including osteocalcin. Biochemical test results were comparable to those reported for osteoblasts of other origins (rat calvaria, human iliac crest), and namely elevated intracellular alkaline phosphatase activity and cAMP accumulation in response to stimulation by human PTH (1–34). Osteoblasts isolated in this manner were cultured in the presence of pure titanium disks to determine the effects of exposure to this metal. Electron microscopy revealed few significant differences in cell growth and specific enzyme activity compared to control osteoblasts grown on plastic dishes, reflecting the excellent biologic and biochemical relationship between the osteoblasts and pure titanium. This experimental system thus appears suitable for biocompatibility studies, and in particular, evaluation of dental implants. 相似文献
10.
The I variant of placental alkaline phosphatase was purified to homogeneity by means of DEAE-cellulose chromatography, isoelectric focusing, and gel filtration on AcA-34. The specific activity of the I variant was found to be 3.33 kat/mg. The enzyme is a dimer with an isoelectric point of 4.6 and a molecular weight of 120,000 as determined by sodium dodecylsulfate electrophoresis. The amino acid composition and other physicochemical properties of the I variant were compared with those of the more common F and S variants. The low activity associated with the I variant is apparently not due to a low specific activity, but to decreased molecular stability. The behavior in the ultracentrifuge and other observations suggest that the I variant differs from the F and S variants in surface charge distribution.This investigation was supported by grants from the Swedish Medical Research Council (projects No. 4217 and No. 03X-2725) and from the Medical Faculty, University of Umeå. 相似文献