Kossa Staining (kossa + staining)

Distribution by Scientific Domains

Kinds of Kossa Staining

  • von kossa staining

  • Selected Abstracts

    Augmentation of osseous phenotypes in vivo with a synthetic peptide

    Xinhua Lin
    Abstract The synthetic peptide B2A2-K-NS augmented the in vitro expression of osseous phenotypes when cells were stimulated with BMP-2, an osteoinductive growth factor. B2A2-K-NS significantly enhanced the effects of BMP-2-induced alkaline phosphatase activity and mineralization. In the absence of BMP-2, B2A2-K-NS did not have an effect on these endpoints. Based on these observations, in vivo studies were conducted to evaluate if B2A2-K-NS could augment osseous phenotypes in an osteoinductive environment in which BMP-2 should be present. In one study, human demineralized bone matrix (DBM) was used to generate an osteoinductive environment and the effects of B2A2-K-NS on ectopic mineralization of subcutaneous implants evaluated. In the second study, a noncritical sized defect in rabbit ulnas with inherent reparative capacity was used as the osteoinductive environment and was treated with or without B2A2-K-NS. In the DBM studies, B2A2-K-NS augmented mineralization as determined using a combination of radiographic analysis and von Kossa staining at 4 weeks postimplant. In the rabbit ulna model, B2A2-K-NS significantly increased the radiographic bone density in the defects compared to carrier-only or no-treatment controls after 6 weeks. Histological staining confirmed that B2A2-K-NS generated a pronounced bone repair response. The results are consistent with the hypothesis that B2A2-K-NS augments osseous phenotypes in an osteoinductive environment, and suggests that B2A2-K-NS may have clinical utility. 2006 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 25:531,539, 2007 [source]

    Retinoic acid is a potential negative regulator for differentiation of human periodontal ligament cells

    Natsuko Shibuya
    Background and objectives:, Retinoic acid (RA) exerts a wide variety of effects on development, cellular differentiation and homeostasis in various tissues. However, little is known about the effects of RA on the differentiation of periodontal ligament cells. In this study, we investigated whether RA can affect the dexamethasone-induced differentiation of periodontal ligament cells. Methods and results:, Human periodontal ligament cells were differentiated via culturing in the presence of dexamethasone, ascorbic acid, and ,-glycerophosphate for mineralized nodule formation, as characterized by von Kossa staining. Continuous treatment with all- trans -RA inhibited the mineralization in a dose-dependent manner, with complete inhibition over 1 m RA. Other RA analogs, 9- cis -RA and 13- cis -RA, were also effective. Furthermore, addition of RA for just the first 4 days completely inhibited the mineralization; however, as RA was added at later stages of culture, the inhibitory effect was diminished, suggesting that RA had a phase-dependent inhibition of mineralization. RA receptor (RAR)-, agonist (AM-580), but not retinoid X receptor agonist (methoprene acid), inhibited the mineralization, and reverse transcription,polymerase chain reaction analysis revealed that RAR-, was expressed on the cells, suggesting that RAR-, was involved in the inhibitory mechanism. This inhibition was accompanied by inhibition of alkaline phosphatase activity; however, neither expression of platelet-derived growth factor (PDGF) receptor-,, PDGF receptor-,, or epidermal growth factor (EGF) receptor, nor phosphorylation of extracellular signal-regulated kinases triggered by PDGF-ascorbic acid or PDGF-BB was changed, as assessed by flow cytometry or western blot analyses. Conclusions:, These findings suggest that RA is a potential negative regulator for differentiation of human periodontal ligament cells. [source]

    Growth hormone regulates osteogenic marker mRNA expression in human periodontal fibroblasts and alveolar bone-derived cells

    H. R. Haase
    Background:, Growth hormone (GH) is a potent regulator of bone formation. The proposed mechanism of GH action is through the stimulation of osteogenic precursor cell proliferation and, following clonal expansion of these cells, promotion of differentiation along the osteogenic lineage. Objectives:, We tested this hypothesis by studying the effects of GH on primary cell populations of human periodontal ligament cells (PLC) and alveolar bone cells (ABC), which contain a spectrum of osteogenic precursors. Methods:, The cell populations were assessed for mineralization potential after long-term culture in media containing ,-glycerophosphate and ascorbic acid, by the demonstration of mineral deposition by Von Kossa staining. The proliferative response of the cells to GH was determined over a 48-h period using a crystal violet dye-binding assay. The profile of the cells in terms of osteogenic marker expression was established using quantitative reverse transcriptase polymerase chain reaction (RT-PCR) for alkaline phosphatase (ALP), osteopontin, osteocalcin, bone sialoprotein (BSP), as well as the bone morphogenetic proteins BMP-2, BMP-4 and BMP-7. Results:, As expected, a variety of responses were observed ranging from no mineralization in the PLC populations to dense mineralized deposition observed in one GH-treated ABC population. Over a 48-h period GH was found to be non-mitogenic for all cell populations. Quantitative reverse transcriptase polymerase chain reaction (RT-PCR) BSP mRNA expression correlated well with mineralizing potential of the cells. The change in the mRNA expression of the osteogenic markers was determined following GH treatment of the cells over a 48-h period. GH caused an increase in ALP in most cell populations, and also in BMP expression in some cell populations. However a decrease in BSP, osteocalcin and osteopontin expression in the more highly differentiated cell populations was observed in response to GH. Conclusion:, The response of the cells indicates that while long-term treatment with GH may promote mineralization, short-term treatment does not promote proliferation of osteoblast precursors nor induce expression of late osteogenic markers. [source]

    Reduced chondrogenic and adipogenic activity of mesenchymal stem cells from patients with advanced osteoarthritis

    ARTHRITIS & RHEUMATISM, Issue 3 2002
    J. Mary Murphy
    Objective Mesenchymal stem cells (MSCs) are resident in the bone marrow throughout normal adult life and have the capacity to differentiate along a number of connective tissue pathways, among them bone, cartilage, and fat. To determine whether functionally normal MSC populations may be isolated from patients with advanced osteoarthritis (OA), we have compared cells from patients undergoing joint replacement with cells from normal donors. Cell populations were compared in terms of yield, proliferation, and capacity to differentiate. Methods MSCs were prepared from bone marrow aspirates obtained from the iliac crest or from the tibia/femur during joint surgery. In vitro chondrogenic activity was measured as glycosaminoglycan and type II collagen deposition in pellet cultures. Adipogenic activity was measured as the accumulation of Nile Red O-positive lipid vacuoles, and osteogenic activity was measured as calcium deposition and by von Kossa staining. Results Patient-derived MSCs formed colonies in primary culture that were characteristically spindle-shaped with normal morphology. The primary cell yield in 36 of 38 cell cultures from OA donors fell within the range found in cultures from normal donors. However, the proliferative capacity of patient-derived MSCs was significantly reduced. There was a significant reduction in in vitro chondrogenic and adipogenic activity in cultures of patient-derived cells compared with that in normal cultures. There was no significant difference in in vitro osteogenic activity. There was no decline in chondrogenic potential with age in cells obtained from individuals with no evidence of OA. Conclusion These results raise the possibility that the increase in bone density and loss of cartilage that are characteristic of OA may result from changes in the differentiation profile of the progenitor cells that contribute to the homeostatic maintenance of these tissues. [source]

    Adipose-derived stem cell: a better stem cell than BMSC

    Yanxia Zhu
    Abstract To further study the proliferation and multi-differentiation potentials of adipose-derived stem cells (ADSCs), the cells were isolated with improved methods and their growth curves were achieved with cck-8. Surface protein expression was analyzed by flow cytometry to characterize the cell phenotype. The multi-lineage potential of ADSCs was testified by differentiating cells with adipogenic, chondrogenic, osteogenic, and myogenic inducers. The results showed that about 5,,105 stem cells could be obtained from 400 to 600,mg adipose tissue. The ADSCs can be continuously cultured in vitro for up to 1 month without passage and they have several logarithmic growth phases during the culture period. Also, the flow cytometry analysis showed that ADSCs expressed high levels of stem cell-related antigens (CD13, CD29, CD44, CD105, and CD166), while did not express hematopoiesis-related antigens CD34 and CD45, and human leukocyte antigen HLA-DR was also negative. Moreover, stem cell-related transcription factors, Nanog, Oct-4, Sox-2, and Rex-1 were positively expressed in ADSCs. The expression of alkaline phosphatase (ALP) was detected in the early osteogenic induction and the calcified nodules were observed by von Kossa staining. Intracellular lipid droplets could be observed by Oil Red staining. Differentiated cardiomyocytes were observed by connexin43 fluorescent staining. In order to obtain more stem cells, we can subculture ADSCs every 14 days instead of the normal 5 days. ADSCs still keep strong proliferation ability, maintain their phenotypes, and have stronger multi-differentiation potential after 25 passages. Copyright 2008 John Wiley & Sons, Ltd. [source]