Morphological analysis of osteoclastogenesis induced by RANKL in mouse bone marrow cell cultures

Gardner, Colin

doi:10.1016/j.cellbi.2006.12.008

Cited by 17 publications

(23 citation statements)

References 28 publications

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“…This coincides with the results that plating cell density affects the size and morphology of in-vitrodifferentiated osteoclasts (Gardner, 2007). A cell-cell adhesion molecule E-cadherin is reported to be involved in the fusion of mononuclear precursor cells during osteoclastogenesis (Mbalaviele et al, 1995).…”

Section: Introductionsupporting

confidence: 89%

The transient appearance of zipper-like actin superstructures during the fusion of osteoclasts

Takito

Nakamura

Yoda

et al. 2012

Journal of Cell Science

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SummaryMultinucleated osteoclasts are responsible for bone resorption. Hypermultinucleated osteoclasts are often observed in some bone-related diseases such as Paget's disease and cherubism. The cellular mechanics controlling the size of osteoclasts is poorly understood. We introduced EGFP-actin into RAW 264.7 cells to monitor actin dynamics during osteoclast differentiation. Before their terminal differentiation into osteoclasts, syncytia displayed two main types of actin assembly, podosome clusters and clusters of zipper-like structures. The zipper-like structures morphologically resembled the adhesion zippers found at the initial stage of cell-cell adhesion in keratinocytes. In the zipper-like structure, Arp3 and cortactin overlapped with the distribution of dense F-actin, whereas integrin b3, paxillin and vinculin were localized to the periphery of the structure. The structure was negative for WGA-lectin staining and biotin labeling. The zipper-like structure broke down and transformed into a large actin ring, called a podosome belt. Syncytia containing clusters of zipper-like structures had more nuclei than those with podosome clusters. Differentiated osteoclasts with a podosome belt also formed the zipper-like structure at the cell contact site during cell fusion. The breakdown of the cell contact site resulted in the fusion of the podosome belts following plasma membrane fusion. Additionally, osteoclasts in mouse calvariae formed the zipper-like structure in the sealing zone. Therefore, we propose that the zipper-like actin superstructures might be involved in cell-cell interaction to achieve efficient multinucleation of osteoclasts. Understanding of the zipper-like structure might lead to selective therapeutics for bone diseases caused by hypermultinucleated osteoclasts.

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Section: Introductionsupporting

confidence: 89%

The transient appearance of zipper-like actin superstructures during the fusion of osteoclasts

Takito

Nakamura

Yoda

et al. 2012

Journal of Cell Science

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“…5; see Gardner 2007). In TGFβ-stimulated wells, more cells appeared to be of the larger type (more nuclei and associated cytoplasm).…”

Section: Resultsmentioning

confidence: 95%

“…Few individual spread cells were present but small ring structures had formed, mostly within the cytoplasmic masses. Groups of nuclei were always associated with the ring structures, which appeared to be the same as those on the exterior of individual spread cells (called interior rings; Gardner 2007). PGE2 also increased the area of cytoplasmic masses, but to a lesser extent than TGFβ.…”

Section: Resultsmentioning

confidence: 98%

“…When the osteoclasts reach a critical size, they spread and develop an exterior ring structure. These mature osteoclasts are strongly adherent and are differentiated to perform bone resorption (Gardner 2007;Gardner et al 2007). …”

Section: Discussionmentioning

confidence: 99%

“…The plates were subsequently washed with physiological saline, centrifuged for 2 min at 2000 rpm, and left to dry by evaporation. Osteoclasts were counted in categories representing significant stages in their development (Gardner 2007); cells containing three to six nuclei (small), larger fused cells with more than six nuclei (intermediate) and cells with a spreading morphology and an external ring structure (spread cells). These latter cells represented mature, strongly adhering osteoclasts fully differentiated for resorbing bone.…”

Section: Osteoclast Countingmentioning

confidence: 99%

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Comparison of morphological effects of PGE2 and TGFβ on osteoclastogenesis induced by RANKL in mouse bone marrow cell cultures

Gardner¹

2007

Cell Tissue Res

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RANKL, in the presence of M-CSF, induces the development and fusion of TRAP+ osteoclasts in mouse bone marrow cultures at 3-5 days. Early during culture (day 3), most cells are small (up to six nuclei). At lower cell densities, these osteoclasts exhibit a rounded morphology with cytoplasm extending around the cells but, at higher densities, this changes to a stellate morphology with the cytoplasm being retracted around the nuclei with numerous localised cytoplasmic extensions. Under optimal conditions, osteoclast fusion results in conglomerates of many cells, which become large cytoplasmic masses on day 4. PGE2 and TGFbeta have both been shown to increase osteoclast development in this model and their effects on the morphology of osteoclasts during fusion and differentiation have been compared under all these conditions. PGE2 or TGFbeta increase osteoclast numbers and size and also the number of nuclei, indicating increased osteoclast development and fusion. TGFbeta increases the size of rounded osteoclasts (with respect to the number of nuclei) more than PGE2, suggesting that TGFbeta increases cytoplasmic extension. TGFbeta increases the size and number of nuclei in stellate cells but particularly increases the number and length of the cytoplasmic extensions, in contrast to PGE2. Fusion of these extensions with other osteoclasts results in large networks of interconnected cells. On day 4, spreading cells develop but these are still interconnected by cytoplasmic links, a phenomenon not seen in control wells or after treatment with PGE2. TGFbeta is more effective than PGE2 in increasing fusion in the formation of cell conglomerates and cytoplasmic masses. PGE2 decreases overall cell density resulting in additional indirect effects on osteoclast numbers and morphology. However, PGE2 particularly promotes the formation of large mature spreading osteoclasts later during culture.

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How Degradation of Calcium Phosphate Bone Substitute Materials is influenced by Phase Composition and Porosity

et al. 2011

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The chemical composition of calcium phosphate (CaP) materials for the regenerative therapy of large bone defects is similar to that of bone. Additionally, calcium phosphates show an excellent biocompatibility. Besides the support of defect healing calcium phosphate implants should be completely degraded within an adequate time period to be replaced by newly formed bone. Although degradation of CaP‐implants occurs mainly by dissolution of the material, it is important to characterize the osteoclastic resorption as well, which is involved in native bone remodeling. The degradation of bone substitutes made of calcium phosphate ceramics is influenced by various parameters, such as defect size and localization, the general health situation, and age of the patient, but also material properties are important. Especially, the calcium phosphate composition is crucial for the degradation behavior of a calcium phosphate material. Additionally, at the cellular level the micro‐ and macroporosity, including interconnecting pores, influences both, the dissolution and the osteoclastic resorption. In our study, three different calcium phosphate materials (hydroxyapatite, tricalcium phosphate, and a biphasic calcium phosphate) and two different geometries (dense 2D samples and porous 3D scaffolds) are compared regarding their dissolution and resorption behavior. The results show, that the dissolution of CaP‐ceramics, as examined by the incubation in a degradation solution, depends mainly on the calcium phosphate phase but also on the porosity of the implant. Regarding the resorption, cell proliferation and differentiation of a monocytic cell line as well as the formation of resorption lacunas are analyzed. Cell proliferation is comparable on all phase compositions. Cell differentiation and resorption, however, are influenced by the calcium phosphate phase composition and by the implant porosity as well. By understanding these two mechanisms of degradation, bone substitute materials and, as a result, the bone regeneration of large bone defects using CaP‐ceramics can be improved.

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Morphological analysis of osteoclastogenesis induced by RANKL in mouse bone marrow cell cultures

Cited by 17 publications

References 28 publications

The transient appearance of zipper-like actin superstructures during the fusion of osteoclasts

The transient appearance of zipper-like actin superstructures during the fusion of osteoclasts

Comparison of morphological effects of PGE2 and TGFβ on osteoclastogenesis induced by RANKL in mouse bone marrow cell cultures

How Degradation of Calcium Phosphate Bone Substitute Materials is influenced by Phase Composition and Porosity

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