scholarly journals Differences in fracture healing between female and male C57BL/6J mice

2020 ◽  
Author(s):  
Melanie Haffner-Luntzer ◽  
Verena Fischer ◽  
Anita Ignatius
Keyword(s):  
Fracture Healing ◽  
Callus Formation ◽  
Fibrous Tissue ◽  
Biomechanical Testing ◽  
Transgenic Animals ◽  
Male Mice ◽  
Mineral Density ◽  
Fracture Callus ◽  
Female Mice ◽  
Tissue Mineral Density

Abstract Background: Mice are increasingly used in fracture healing research because of the opportunity to use transgenic animals. While both, male and female mice are employed, there is no consensus in the literature whether fracture healing differs between both sexes. Therefore, the aim of the present study was to analyse diaphyseal fracture healing in female and male C57BL/6J mice, a commonly used mouse strain in bone research. Methods: For that purpose, 12-week-old female (17–20 g) and male mice (22–26 g) received a standardised femur midshaft osteotomy stabilised by an external fixator. Mice were euthanized 10 and 21 days after fracture and bone healing was analysed by biomechanical testing, µCT, histology, immunohistochemistry and qPCR. Results: Ten days after fracture, male mice displayed significantly more cartilage but less fibrous tissue in the fracture callus compared to female mice, whereas the amount of bone did not differ. At day 21, male mice showed a significantly larger fracture callus compared to female mice. The relative amount of bone in the fracture callus did not significantly differ between both sexes, whereas its tissue mineral density was significantly higher in male mice on day 21, indicating more mature bone and slightly more rapid fracture healing. These results were confirmed by a significantly greater absolute bending stiffness of the fractured femurs of male mice on day 21. On the molecular level, male mice displayed increased active β-catenin expression in the fracture callus, whereas oestrogen receptor α (ERα) expression was lower. Conclusions: These results suggest that male mice display more rapid fracture healing with more prominent cartilaginous callus formation. This might be due to the higher weight of male mice, resulting in increased mechanical loading of the fracture. Furthermore, male mice displayed significantly greater activation of osteoanabolic Wnt/β-catenin signalling, which might also contribute to more rapid bone regeneration.

scholarly journals Differences in Fracture Healing Between Female and Male C57BL/6J Mice

2021 ◽  
Vol 12 ◽  
Author(s):  
Melanie Haffner-Luntzer ◽  
Verena Fischer ◽  
Anita Ignatius
Keyword(s):  
Fracture Healing ◽  
Callus Formation ◽  
Fibrous Tissue ◽  
Biomechanical Testing ◽  
Transgenic Animals ◽  
Estrogen Receptor Α ◽  
Male Mice ◽  
Mineral Density ◽  
Fracture Callus ◽  
Female Mice

BackgroundMice are increasingly used in fracture healing research because of the opportunity to use transgenic animals. While both, male and female mice are employed, there is no consensus in the literature whether fracture healing differs between both sexes. Therefore, the aim of the present study was to analyze diaphyseal fracture healing in female and male C57BL/6J mice, a commonly used mouse strain in bone research.MethodsFor that purpose, 12-week-old Female (17–20 g) and Male mice (22–26 g) received a standardized femur midshaft osteotomy stabilized by an external fixator. Mice were euthanized 10 and 21 days after fracture and bone healing was analyzed by biomechanical testing, μCT, histology, immunohistochemistry and qPCR.ResultsTen days after fracture, Male mice displayed significantly more cartilage but less fibrous tissue in the fracture callus compared to Female mice, whereas the amount of bone did not differ. At day 21, Male mice showed a significantly larger fracture callus compared to Female mice. The relative amount of bone in the fracture callus did not significantly differ between both sexes, whereas its tissue mineral density was significantly higher in Male mice on day 21, indicating more mature bone and slightly more rapid fracture healing. These results were confirmed by a significantly greater absolute bending stiffness of the fractured femurs of Male mice on day 21. On the molecular level, Male mice displayed increased active β-catenin expression in the fracture callus, whereas estrogen receptor α (ERα) expression was lower.ConclusionThese results suggest that Male mice display more rapid fracture healing with more prominent cartilaginous callus formation. This might be due to the higher weight of Male mice, resulting in increased mechanical loading of the fracture. Furthermore, Male mice displayed significantly greater activation of osteoanabolic Wnt/β-catenin signaling, which might also contribute to more rapid bone regeneration.

scholarly journals Heterogeneity of murine periosteum osteochondroprogenitors involved in fracture healing

2020 ◽  
Author(s):  
Brya G Matthews ◽  
Francesca V Sbrana ◽  
Sanja Novak ◽  
Jessica L. Funnell ◽  
Ye Cao ◽  
...  
Keyword(s):  
Fracture Healing ◽  
Progenitor Cells ◽  
Progenitor Cell ◽  
Callus Formation ◽  
Lineage Tracing ◽  
Fracture Callus ◽  
Stem And Progenitor Cells ◽  
Periosteal Cells ◽  
Osteoblast Lineage

AbstractThe periosteum is the major source of cells involved in fracture healing. We sought to characterize differences in progenitor cell populations between periosteum and other bone compartments, and identify periosteal cells involved in fracture healing. The periosteum is highly enriched for progenitor cells, including Sca1+ cells, CFU-F and label-retaining cells. Lineage tracing with αSMACreER identifies periosteal cells that contribute to >80% of osteoblasts and ~40% of chondrocytes following fracture. A subset of αSMA+ cells are quiescent long-term injury-responsive progenitors. Ablation of αSMA+ cells impairs fracture callus formation. In addition, committed osteoblast-lineage cells contributed around 10% of osteoblasts, but no chondrocytes in fracture calluses. Most periosteal progenitors, particularly those that form osteoblasts, can be targeted by αSMACreER. We have demonstrated that the periosteum is highly enriched for skeletal stem and progenitor cells and there is heterogeneity in the populations of cells that contribute to mature lineages during periosteal fracture healing.

scholarly journals Effect of Cervus and Cucumis Peptides on Osteoblast Activity and Fracture Healing in Osteoporotic Bone

2014 ◽  
Vol 2014 ◽  
pp. 1-10 ◽  
Author(s):  
Ai-Yuan Wang ◽  
Yue Tian ◽  
Mei Yuan ◽  
Li Zhang ◽  
Ji-Feng Chen ◽  
...  
Keyword(s):  
Fracture Healing ◽  
Callus Formation ◽  
Biomechanical Testing ◽  
Physiological Saline ◽  
Osteoporotic Bone ◽  
Sprague Dawley ◽  
Matrix Mineralization ◽  
Osteoblast Activity

Osteoporosis is associated with delayed and/or reduced fracture healing. As cervus and cucumis are the traditional Chinese treatments for rheumatoid arthritis, we investigated the effect of supplementation of these peptides (CCP) on bone fracture healing in ovariectomized (OVX) osteoporotic ratsin vitroandin vivo. CCP enhanced osteoblast proliferation and increased alkaline phosphatase activity, matrix mineralization, and expression of runt-related transcription factor 2 (Runx2), bone morphogenetic protein 4 (BMP4), and osteopontin.In vivo, female Sprague-Dawley rats underwent ovariectomy and the right femora were fractured and fixed by intramedullary nailing 3 months later. Rats received intraperitoneal injections of either CCP (1.67 mg/kg) or physiological saline every day for 30 days. Fracture healing and callus formation were evaluated by radiography, micro-CT, biomechanical testing, and histology. At 12 weeks after fracture, calluses in CCP-treated bones showed significantly higher torsional strength and greater stiffness than control-treated bones. Bones in CCP-treated rats reunified and were thoroughly remodeled, while two saline-treated rats showed no bone union and incomplete remodeling. Taken together, these results indicate that use of CCP after fracture in osteoporotic rats accelerates mineralization and osteogenesis and improves fracture healing.

scholarly journals Modulation of fracture healing by the transient accumulation of senescent cells

2021 ◽  
Author(s):  
Sundeep Khosla ◽  
Dominik Saul ◽  
David Monroe ◽  
Jennifer L Rowsey ◽  
Robyn Laura Kosinsky ◽  
...  
Keyword(s):  
Fracture Healing ◽  
Time Course ◽  
Callus Formation ◽  
In Silico Analysis ◽  
Skin Wound ◽  
Luciferase Reporter ◽  
Skin Wound Healing ◽  
Fracture Callus ◽  
Beneficial Effects

Senescent cells have detrimental effects across tissues with aging but may have beneficial effects on tissue repair, specifically on skin wound healing. However, the potential role of senescent cells in fracture healing has not been defined. Here, we performed an in silico analysis of public mRNAseq data and found that senescence and senescence-associated secretory phenotype (SASP) markers increased during fracture healing. We next directly established that the expression of senescence biomarkers increased markedly during murine fracture healing. We also identified a subset of cells in the fracture callus that displayed hallmarks of senescence, including distension of satellite heterochromatin and telomeric DNA damage. Then, using a genetic mouse model (p16LUC) containing a p16Ink4a55-driven luciferase reporter, we demonstrated transient in vivo senescent cell accumulation during callus formation. Finally, we intermittently treated young adult mice following fracture with drugs that selectively eliminate senescent cells ("senolytics", Dasatinib plus Quercetin), and showed that this regimen both decreased senescence and SASP markers in the fracture callus and significantly accelerated the time course of fracture healing. Our findings thus demonstrate that senescent cells accumulate transiently in the murine fracture callus and, in contrast to the skin, their clearance does not impair but rather may improve fracture healing.

A 3D Computational Simulation of Fracture Callus Formation: Influence of the Stiffness of the External Fixator

2005 ◽  
Vol 128 (3) ◽  
pp. 290-299 ◽  
Author(s):  
M. J. Gómez-Benito ◽  
J. M. García-Aznar ◽  
J. H. Kuiper ◽  
M. Doblaré
Keyword(s):  
Fracture Healing ◽  
External Fixator ◽  
Callus Formation ◽  
Computational Simulation ◽  
Element Model ◽  
Regenerative Process ◽  
Fracture Site ◽  
Three Dimensions ◽  
Fracture Callus ◽  
Clinical Observations

The stiffness of the external fixation highly influences the fracture healing pattern. In this work we study this aspect by means of a finite element model of a simple transverse mid-diaphyseal fracture of an ovine metatarsus fixed with a bilateral external fixator. In order to simulate the regenerative process, a previously developed mechanobiological model of bone fracture healing was implemented in three dimensions. This model is able to simulate tissue differentiation, bone regeneration, and callus growth. A physiological load of 500N was applied and three different stiffnesses of the external fixator were simulated (2300, 1725, and 1150N∕mm). The interfragmentary strain and load sharing mechanism between bone and the external fixator were compared to those recorded in previous experimental works. The effects of the stiffness on the callus shape and tissue distributions in the fracture site were also analyzed. We predicted that a lower stiffness of the fixator delays fracture healing and causes a larger callus, in correspondence to well-documented clinical observations.

scholarly journals Ablation of Proliferating Osteoblast Lineage Cells After Fracture Leads to Atrophic Nonunion in a Mouse Model

2020 ◽  
Author(s):  
Katherine R. Hixon ◽  
David A.W. Sykes ◽  
Susumu Yoneda ◽  
Jennifer A. McKenzie ◽  
Austin Hensley ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Ex Vivo ◽  
Fibrous Tissue ◽  
Biomechanical Testing ◽  
Suicide Gene ◽  
Fracture Site ◽  
Fracture Callus ◽  
Atrophic Nonunion ◽  
Osteoblast Lineage

ABSTRACTNonunion is defined as the permanent failure of a fractured bone to heal, often necessitating surgical intervention. Atrophic nonunions are a subtype that are particularly difficult to treat. Animal models of atrophic nonunion are available; however, these require surgical or radiation-induced trauma to disrupt periosteal healing. These methods are highly invasive and not representative of many clinical nonunions where osseous regeneration has been arrested by a “failure of biology”. We hypothesized that arresting osteoblast cell proliferation after fracture would lead to atrophic nonunion in mice. Using mice that express a thymidine kinase (tk) “suicide gene” driven by the 3.6Col1a1 promoter (Col1-tk), proliferating osteoblast lineage cells can be ablated upon exposure to the nucleoside analog ganciclovir (GCV). Wild-type (WT; control) and Col1-tk littermates were subjected to a full femur fracture and intramedullary fixation at 12 weeks age. We confirmed abundant tk+ cells in fracture callus of Col-tk mice dosed with PBS. The remainder of mice were dosed with GCV twice daily for 2 or 4 weeks. Histologically, we observed diminished periosteal cell proliferation in Col1-tk mice 3 weeks post fracture. Moreover, Col1-tk mice had less osteoclast activity, mineralized callus, and vasculature at the fracture site compared to WT mice. Additional mice were monitored for 12 weeks with in vivo radiographs and microCT scans, revealing delayed bone bridging and reduced callus size in Col1-tk mice. Following sacrifice, ex vivo microCT and histology demonstrated failed union with residual bone fragments and fibrous tissue in Col1-tk mice. Biomechanical testing demonstrated a failure to recover torsional strength in Col1-tk mice, in contrast to WT. Our data indicates that suppression of proliferating osteoblast-lineage cells for at least 2 weeks after fracture blunts the formation and remodeling of a mineralized callus leading to a functional nonunion. We propose this as a new murine model of atrophic nonunion.

scholarly journals Review of Fracture Healing and Outcomes Assessment

2020 ◽  
Vol 3 ◽  
Author(s):  
Adam Knoximprs ◽  
Anthony McGuire ◽  
Christopher Collier ◽  
Melissa Kacena ◽  
Roman Natoli
Keyword(s):  
Fracture Healing ◽  
Bone Fractures ◽  
Healing Process ◽  
Biomechanical Testing ◽  
Fracture Repair ◽  
Long Bone ◽  
Micro Computed Tomography ◽  
Fracture Callus ◽  
Medical Interventions ◽  
Healing Assessment

Background/Objective: Long bone fractures are of the most common and costly medical traumas humans experience.  Adequate characterization of the fracture healing process and development of potential medical interventions generally involves fracture induction operations on animal models of varying treatment or genetic groups, then analyzing relative repair success via synthesis of diverse assessment methodologies.  This review discusses the procedures, relevant parameters, special considerations, and key correlations of these major methodologies of fracture repair quantification.  Methods: A literature review was conducted for articles discussing the procedures or identifying correlations between each of the major fracture healing assessment methodologies.    Results: These methodologies include biomechanical testing, which provides the most direct quantification of skeletal functionality; micro-computed tomography, which enables high resolution visualization of fracture callus architecture; histology which helps elucidate the intricate processes underlying fracture repair; and x-ray which offers a non-invasive and clinically relevant view of fracture repair progress.  Each of these methodologies measure parameters directly correlating to restored functionality of fractured bone.  Conclusion: When appropriately integrated, synthesis of relevant parameters from each methodology of fracture repair assessment enables a comprehensive understanding of varying fracture healing outcomes and associated causalities.  Scientific/Clinical Policy Impact and Implications: This review may guide the interpretation and planning of fracture healing studies utilizing murine models. 

Tranexamic Acid Protects Ovary and Testis Functions and Ameliorates Osteoporosis in Mice

Pharmacology ◽  
2020 ◽  
Vol 105 (11-12) ◽  
pp. 652-661 ◽  
Author(s):  
Keiichi Hiramoto ◽  
Hirotaka Oikawa ◽  
Yurika Yamate ◽  
Eisuke F. Sato
Keyword(s):  
Tranexamic Acid ◽  
Transforming Growth Factor ◽  
Transforming Growth Factor Β ◽  
Male Mice ◽  
Mineral Density ◽  
Effective Prevention ◽  
Female Mice ◽  
Ameliorative Effect ◽  
17Β Estradiol ◽  
Old Mice

<b><i>Introduction:</i></b> In a rapidly aging society, the number of people suffering from osteoporosis keeps increasing. However, effective prevention strategies for osteoporosis are not yet currently available. <b><i>Objective:</i></b> In this study, we examined the ameliorative effects of tranexamic acid on osteoporosis in 24-month-old mice. <b><i>Methods:</i></b> During the study period, mice were orally administered tranexamic acid 3 times per week. <b><i>Results:</i></b> Bone mineral density, which is a parameter of osteoporosis, was improved following tranexamic acid administration. In addition, female mice evidenced a stronger phenotypic improvement than male mice. In female mice treated with tranexamic acid, ovary abnormalities were reduced. Furthermore, the levels of transforming growth factor-β, hyaluronic acid, CD44, reactive oxygen species, and apoptosis, as well as the number of infiltrated neutrophils and macrophages in the ovary were lower than those in the control or solvent-administered mice. In addition, 17β-estradiol levels in blood increased when compared with the control or solvent-treated mice. In addition, administration of tranexamic acid to 24-month-old male mice decreased the level of apoptosis in the testis. However, the levels of 17β-estradiol and testosterone in blood increased compared with the control or solvent-administered mice. <b><i>Conclusions:</i></b> The use of tranexamic acid had an ameliorative effect on osteoporosis, possibly by protecting ovaries and testes.

scholarly journals Modulation of fracture healing by the transient accumulation of senescent cells

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Dominik Saul ◽  
David G Monroe ◽  
Jennifer L Rowsey ◽  
Robyn Laura Kosinsky ◽  
Stephanie J Vos ◽  
...  
Keyword(s):  
Fracture Healing ◽  
Time Course ◽  
Callus Formation ◽  
Skin Wound ◽  
Luciferase Reporter ◽  
Skin Wound Healing ◽  
Fracture Callus ◽  
Beneficial Effects ◽  
Genetic Mouse Model

Senescent cells have detrimental effects across tissues with aging but may have beneficial effects on tissue repair, specifically on skin wound healing. However, the potential role of senescent cells in fracture healing has not been defined. Here, we performed an in silico analysis of public mRNAseq data and found that senescence and senescence-associated secretory phenotype (SASP) markers increased during fracture healing. We next directly established that the expression of senescence biomarkers increased markedly during murine fracture healing. We also identified cells in the fracture callus that displayed hallmarks of senescence, including distension of satellite heterochromatin and telomeric DNA damage; the specific identity of these cells, however, requires further characterization. Then, using a genetic mouse model (Cdkn2aLUC) containing a Cdkn2aInk4a-driven luciferase reporter, we demonstrated transient in vivo senescent cell accumulation during callus formation. Finally, we intermittently treated young adult mice following fracture with drugs that selectively eliminate senescent cells ('senolytics', Dasatinib plus Quercetin), and showed that this regimen both decreased senescence and SASP markers in the fracture callus and significantly accelerated the time course of fracture healing. Our findings thus demonstrate that senescent cells accumulate transiently in the murine fracture callus and, in contrast to the skin, their clearance does not impair but rather improves fracture healing.

scholarly journals YAP and TAZ promote periosteal osteoblast precursor expansion and differentiation for fracture repair

2020 ◽  
Author(s):  
Christopher D. Kegelman ◽  
Madhura P. Nijsure ◽  
Yasaman Moharrer ◽  
Hope B. Pearson ◽  
James H. Dawahare ◽  
...  
Keyword(s):  
Bone Formation ◽  
Fracture Healing ◽  
Bone Fracture ◽  
Callus Formation ◽  
Fracture Repair ◽  
Binding Motif ◽  
Adult Onset ◽  
Bone Fracture Healing ◽  
Fracture Callus ◽  
Endochondral Bone

ABSTRACTIn response to bone fracture, periosteal progenitor cells proliferate, expand, and differentiate to form cartilage and bone in the fracture callus. These cellular functions require the coordinated activation of multiple transcriptional programs, and the transcriptional regulators Yes-associated protein (YAP) and transcriptional co-activator with PDZ-binding motif (TAZ) regulate osteochondroprogenitor activation during endochondral bone development. However, recent observations raise important distinctions between the signaling mechanisms used to control bone morphogenesis and repair. Here, we tested the hypothesis that YAP and TAZ regulate osteochondroprogenitor activation during endochondral bone fracture healing. Constitutive YAP and/or TAZ deletion from Osterix-expressing cells impaired both cartilage callus formation and subsequent mineralization. However, this could be explained either by direct defects in osteochondroprogenitor differentiation after fracture, or by developmental deficiencies in the progenitor cell pool prior to fracture. Consistent with the second possibility, we found that developmental YAP/TAZ deletion produced long bones with impaired periosteal thickness and cellularity. Therefore, to remove the contributions of developmental history, we next generated adult onset-inducible knockout mice (using Osx1-CretetOff) in which YAP and TAZ were deleted prior to fracture, but after normal development. Adult onset-induced YAP/TAZ deletion had no effect on cartilaginous callus formation, but impaired bone formation at 14 days post-fracture (dpf). Earlier, at 4 dpf, adult onset-induced YAP/TAZ deletion impaired the proliferation and expansion of osteoblast precursor cells located in the shoulder of the callus. Further, activated periosteal cells isolated from this region at 4 dpf exhibited impaired osteogenic differentiation in vitro upon YAP/TAZ deletion. Finally, confirming the effects on osteoblast function in vivo, adult onset-induced YAP/TAZ deletion impaired bone formation in the callus shoulder at 7 dpf, prior to the initiation of endochondral ossification. Together, these data show that YAP and TAZ promote the expansion and differentiation of periosteal osteoblast precursors to accelerate bone fracture healing.

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