Ex vivo real-time observation of Ca2+ signaling in living bone in response to shear stress applied on the bone surface

Ishihara, Yoshihito; Sugawara, Yasutake; Kamioka, Hiroshi; Kawanabe, Noriaki; Hayano, Satoru; Balam, Tarek A.; Naruse, Keiji; Yamashiro, Takashi

doi:10.1016/j.bone.2012.12.002

Cited by 49 publications

(36 citation statements)

References 59 publications

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“…1C, D). This is consistent with data from human bone marrow-and adipose tissue-derived mesenchymal stem cells[38,39].Moreover, the decrease in Ca 2+ oscillation during AFMSC osteoblastic differentiation (Figs.1C, D) is in agreement with previous reports[34,40]. On the other hand, the observed increment in basal [Ca 2+ ]i is in line with the adipocyte differentiation program[41], as an increment in [Ca 2+ ]i upregulates the expression of PPARγ, a nuclear hormone receptor that acts as a critical transcriptional factor in adipocyte differentiation[42].…”

supporting

confidence: 93%

Molecular and Phenotypic Characterization of Human Amniotic Fluid-Derived Cells: A Morphological and Proteomic Approach

Pipino

PierdomenicoLaura

TomoPamela

et al. 2015

Stem Cells and Development

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Mesenchymal Stem Cells derived from Amniotic Fluid (AFMSCs) are multipotent cells of great interest for regenerative medicine. Two predominant cell types, that is, Epithelial-like (E-like) and Fibroblast-like (F-like), have been previously detected in the amniotic fluid (AF). In this study, we examined the AF from 12 donors and observed the prevalence of the E-like phenotype in 5, whereas the F-like morphology was predominant in 7 samples. These phenotypes showed slight differences in membrane markers, with higher CD90 and lower Sox2 and SSEA-4 expression in F-like than in E-like cells; whereas CD326 was expressed only in the E-like phenotype. They did not show any significant differences in osteogenic, adipogenic or chondrogenic differentiation. Proteomic analysis revealed that samples with a predominant E-like phenotype (HC1) showed a different profile than those with a predominant F-like phenotype (HC2). Twenty-five and eighteen protein spots were differentially expressed in HC1 and HC2 classes, respectively. Of these, 17 from HC1 and 4 from HC2 were identified by mass spectrometry. Protein-interaction networks for both phenotypes showed strong interactions between specific AFMSC proteins and molecular chaperones, such as preproteasomes and mature proteasomes, both of which are important for cell cycle regulation and apoptosis. Collectively, our results provide evidence that, regardless of differences in protein profiling, the prevalence of E-like or F-like cells in AF does not affect the differentiation capacity of AFMSC preparations. This may be valuable information with a view to the therapeutic use of AFMSCs.

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supporting

confidence: 93%

Molecular and Phenotypic Characterization of Human Amniotic Fluid-Derived Cells: A Morphological and Proteomic Approach

Pipino

PierdomenicoLaura

TomoPamela

et al. 2015

Stem Cells and Development

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“…Though in vitro studies subjecting osteocytes to fluid shear strongly support a role for Ca 2þ signalling to encode information important for mechanotransduction in osteocytes, limited systems were available to show that Ca 2þ signals in osteocytes embedded within the bone could be induced by load. In explanted fragments of chicken embryonic calvariae, bone cells were demonstrated to exhibit autonomous Ca 2þ responses [76], and embedded osteocytes were found to respond with elevated Ca 2þ to bone matrix deformation [77] and shear stress applied over the explant surfaces [78]. We recently modified the ex vivo system described earlier to observe Ca 2þ responses in live osteocytes in a mouse long bone subjected to dynamic, deformational loading (figure 2a).…”

Section: Mechanosensation and Early Mechanotransduction In Osteocytesmentioning

confidence: 99%

Experimental studies of bone mechanoadaptation: bridging in vitro and in vivo studies with multiscale systems

Brown¹,

Sattler²

2016

Interface Focus.

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Despite advancements in technology and science over the last century, the mechanisms underlying Wolff's law—bone structure adaptation in response to physical stimuli—remain poorly understood, limiting the ability to effectively treat and prevent skeletal diseases. A challenge to overcome in the study of the underlying mechanisms of this principle is the multiscale nature of mechanoadaptation. While there exist in silico systems that are capable of studying across these scales, experimental studies are typically limited to interpretation at a single dimension or time point. For instance, studies of single-cell responses to defined physical stimuli offer only a limited prediction of the whole bone response, while overlapping pathways or compensatory mechanisms complicate the ability to isolate critical targets in a whole animal model. Thus, there exists a need to develop experimental systems capable of bridging traditional experimental approaches and informing existing multiscale theoretical models. The purpose of this article is to review the process of mechanoadaptation and inherent challenges in studying its underlying mechanisms, discuss the limitations of traditional experimental systems in capturing the many facets of this process and highlight three multiscale experimental systems which bridge traditional approaches and cover relatively understudied time and length scales in bone adaptation.

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“…Mechanotransduction of osteocytes in calvarial bone fragments via a micro-needle displacement or surface fluid shear have been investigated [32, 33]. Recent research has also shown the real-time Ca 2+ oscillations in response to a direct mechanical stimulation on an intact ex vivo mouse tibia [31].…”

Section: Discussionmentioning

confidence: 99%

“…Furthermore, in response to in vitro and in situ mechanical stimulations, osteocytes seem to be more sensitive than osteoblasts, in terms of Ca 2+ oscillations. Recent studies have observed the real-time Ca 2+ oscillations in response to fluid shear on ex vivo bone segments, as well as a direct mechanical stimulation on an intact ex vivo mouse tibia [31–33]. However, these studies either investigated the effect of fluid shear on osteocytic Ca 2+ oscillations only on ex vivo bone segment surfaces that were not yet adapted for dynamic mechanical events within bone, or studied the effect of osteocytic Ca 2+ oscillations of intact mouse long bones deformable bone loading that the recording was done only at the resting period with time delay.…”

Section: Introductionmentioning

confidence: 99%

Dynamic fluid flow induced mechanobiological modulation of in situ osteocyte calcium oscillations

Guo-wei

Gibbons

et al. 2015

Archives of Biochemistry and Biophysics

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Distribution of intramedullary pressure (ImP) induced bone fluid flow (BFF) has been suggested to influence the magnitude of mechanotransductory signals within bone. As osteocytes have been suggested as major mechanosensors in bone network, it is still unclear how osteocytes embedded within a mineralized bone matrix respond to the external mechanical stimuli derived from direct coupling of dynamic fluid flow stimulation (DFFS). While in vitro osteocytes show unique Ca2+ oscillations to fluid shear, the objective of this study was to use a confocal imaging technique to visualize and quantify Ca2+ responses in osteocytes in situ under DFFS into the marrow cavity of an intact ex vivo mouse femur. This study provided significant technical development for evaluating mechanotransduction mechanism in bone cell response by separation of mechanical strain and fluid flow factors using intramedullary pressure stimulation, provides the ability for true real-time imaging and monitoring of bone cell activities during stimulation. Loading frequency dependent Ca2+ oscillations in osteocytes indicated the optimized loading at 10Hz, where such induced response was significantly diminished via blockage of the Wnt/β-catenin signaling pathway. The results provided a pilot finding of the potential crosstalk or interaction between Wnt/β-catenin signaling and Ca2+ influx signaling of in situ osteocytes in response to mechanical signals. Findings from the present study make a valuable tool to investigate how in situ osteocytes respond and transduce mechanical signals, e.g. DFFS, as a central mechanosensor.

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Ex vivo real-time observation of Ca2+ signaling in living bone in response to shear stress applied on the bone surface

Cited by 49 publications

References 59 publications

Molecular and Phenotypic Characterization of Human Amniotic Fluid-Derived Cells: A Morphological and Proteomic Approach

Molecular and Phenotypic Characterization of Human Amniotic Fluid-Derived Cells: A Morphological and Proteomic Approach

Experimental studies of bone mechanoadaptation: bridging in vitro and in vivo studies with multiscale systems

Dynamic fluid flow induced mechanobiological modulation of in situ osteocyte calcium oscillations

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