Mineralization Of New Bone Research Articles

Abstract 2956: Chemotherapy-induced adipocyte senescence triggers bone loss through osteoclast activation

Abstract Despite breakthroughs in cancer treatment, chemotherapy-induced osteo-toxicity is a major problem that compromises the quality of life and overall survival of cancer patients regardless of cancer type. Recently we demonstrated that chemotherapy-induced senescence drives bone loss by both limiting mineralization of new bone and increasing bone resorption. However, the underlying mechanism of action remains elusive. Therefore, it is critical that we understand the mechanisms that drive these toxicities and develop approaches to mitigate their severity. To establish whether senescent bone resident cells or systemic responses to chemotherapy drove therapy-induced bone loss, we used a vertebral body transplant (vossicle) model. Using this approach, we found that the specific elimination of senescent cells in L4 and L5 donor vossicles (INK-ATTAC) implanted into wildtype mice, protects from chemotherapy-induced bone loss within the vossicles but not the femur of the recipient mice. This demonstrated that chemotherapy-induced senescence in resident bone cells is responsible for bone loss. To determine which bone resident cell(s) underwent senescence in response to chemotherapy and how their gene expression was impacted, we used the p16-CreERT2-tdTomato mouse model and found that chemotherapy triggers senescence in bone marrow adipocytes. Subsequently, we observed that postnatal fat ablation in adipoqCre-inducible DTR transgenic mice (iDTRADQ) prevented chemotherapy-induced bone loss, indicating senescent adipocytes trigger bone loss. Furthermore, we observed chemotherapy-induced bone loss is attributed to RANKL-mediated high osteoclasts activity. Collectively, our data demonstrate that chemotherapy causes senescence in marrow adipocytes which in turn triggers bone loss via RANKL-mediated osteoclasts activation and bone loss can be protected by eliminating senescent cells. Citation Format: Ganesh Kumar Raut, Taylor Malachowski, Taylor Holt, Renata Oliveira, Xianmin Luo, Douglas Faget, Qihao Ren, Sheila Stewart. Chemotherapy-induced adipocyte senescence triggers bone loss through osteoclast activation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2956.

Abstract 137: Abrogating therapy induced senescence driven bone loss to improve quality of life

Abstract Despite the central role chemotherapy plays in prolonging survival, the toxicities associated with its use can negatively impact quality of life and in some cases, be so severe that patients forego further life preserving treatments. Therefore, it is critical that we understand the mechanisms that drive these toxicities and develop approaches to mitigate their severity. Recently we demonstrated that chemotherapy-induced senescence drives bone loss by both limiting mineralization of new bone and increasing bone resorption. To establish whether senescent bone resident cells or systemic responses to chemotherapy drove therapy-induced bone loss, we used a vossicle model in which neonatal vertebral bones (L4 and L5) were transplanted from 4-day old wildtype or INKATTAC pups (allow to selectively kill senescent cells) into wildtype or INKATTAC adult mice. Using this approach, we found that the elimination of senescent cells in donor vossicles protects from chemotherapy-induced bone loss, indicating that senescent bone resident cells are responsible for therapy-induced bone loss. To understand the mechanism(s) that contributed to therapy-induced bone loss, we used scRNA-seq to determine which bone resident cells underwent senescence in response to chemotherapy and how their gene expression was impacted. Using this approach, we found that adipocytes, fibroblast, chondrocytes and osteoblasts underwent senescence as evidenced by expression of p16, p21 and loss of Mki67 and displayed evidence of a senescence associated secretory phenotype (SASP), which we postulate contributes to therapy-induced bone loss. Collectively, our data indicated that chemotherapy causes bone loss via senescence and can be protected by eliminating senescent cells. Citation Format: Ganesh Kumar Raut, Zhangting Yao, Tom Cole, Xainmin Luo, Qihao Ren, Sheila A. Stewart. Abrogating therapy induced senescence driven bone loss to improve quality of life [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 137.

Flow-through anterolateral thigh flap transfer combined with bone transport technique for repairing the segamental mutilating injury in lower extremity

Objective To explore the clinical application and effect of Flow-through anterolateral thigh flap transfer combined with bone transport technique for repairing the segamental mutilating injury in lower extremity. Methods From June, 2010 to June, 2016, Flow-through anterolateral thigh flap transfer combined with Ilizarov technique were applied to repair the segamental mutilating injury in lower extremity in 10 cases. Of the 10 cases, there were 8 males and 2 females (their ages ranged from 26 to 55, 36 on average). Four caused by road accident, 4 caused by crush injury, 2 cases caused by squeeze injury. First stage, Flow-through anterolateral thigh flap transfer were applied to revascularize the lower limb and repair the soft tissue defect in emergency. Second stage, bone transfort with external fixator were applied to repair bone defect in second stage. The area of the flap ranged from 12.0 cm×15.5 cm to 20.0 cm×25.0 cm. The repair of bone defect ranged from 4.0 cm to 10.0 cm. Time interval between two stages ranged from 2 to 4 months, 3 months on average. Followed-up method was regular outpatient-care after discharge. The main contents include: the presence of red swollen with pus, the condition of infection in the pin, if the fixed pin was loosening, and adjust the speed of bone transport according to the mineralization of new bone. Results All cases were successfully repaired in 10 cases. Ten cases were followed-up with a mean followed-up time of 42 months. All flaps survived, in with 6 cases were healed in first stage, 4 case were healed in second stage (healing time ranged from 14 to 30 days), and bone transfort time ranged from 6 to 16 months. The docking site union occurred in 6 cases and were healed by use of bone debridement, bone graft and compression. Deformity in foot and ankle occurred during the bone transport in 2 cases and were cured by draft therapy with adding ring fixator. The functions of donor sites were not found malfunctional. Conclusion Flow-through anterolateral thigh flap transfer combined with bone transport have high success rate of limb salvage, good quality of osteogenesis, and satisfactory function recovery. It is an optimal method to repair the segamental mutilating injury of the lower extremity. Key words: Anterolateral thigh flap; Flow-through flap; Bone transport technique; Segamental mutilating injury; Lower extremity

Subchondral bone response to injected adipose-derived stromal cells for treating osteoarthritis using an experimental rabbit model

Although articular cartilage is the target of osteoarthritis (OA), its deterioration is not always clearly associated with patient symptoms. Because a functional interaction between cartilage and bone is crucial, the pathophysiology of OA and its treatment strategy must focus also on subchondral bone. We investigated whether adipose-derived stromal cells (ASCs) injected into a joint at two different concentrations could prevent subchondral bone damage after the onset of mild OA in a rabbit model. We measured both volumetric and densitometric aspects of bone remodeling. Although OA can stimulate bone remodeling either catabolically or anabolically over time, the accelerated turnover does not allow complete mineralization of new bone and therefore gradually reduces its density. We measured changes in morphometric and densitometric bone parameters using micro-CT analysis and correlated them with the corresponding parameters in cartilage and meniscus. We found that ASCs promoted cartilage repair and helped counteract the accelerated bone turnover that occurs with OA.

Increased osteoinductivity and mineralization by minimal concentration of bone morphogenetic protein-2 loaded onto biphasic calcium phosphate in a rabbit sinus.

PurposeThe purpose of the present study was to evaluate the effectiveness of a minimal concentration of bone morphogenetic protein-2 (BMP-2) in terms of quantitative and qualitative analyses of newly formed bone in a rabbit maxillary sinus model.MethodsIn 7 rabbits, sinus windows were prepared bilaterally. Biphasic calcium phosphate (BCP) loaded with 0.05 mg/mL BMP-2 was grafted into one sinus (the BMP group) and saline-soaked BCP was placed into the other (the control group) in each animal. The animals were allowed an 8-week healing period before being sacrificed. Specimens including the augmented area and surrounding tissues were then removed and evaluated both radiographically and histologically.ResultsThere was a difference in the mineralization of new bone between the groups. In the BMP group, the greater part of the new bone consisted of mature lamellar bone with an evident trabecular pattern, whereas the control group showed mostly woven bone, consisting only partially of lamellar bone. Histometrically, the area of new bone was significantly greater (4.55±1.35 mm2 vs. 2.99±0.86 mm2) in the BMP group than in the control group (P<0.05); however, the total augmentation volumes were not significantly different between the groups.ConclusionsWithin the limitations of this study, it can be suggested that a minimal concentration of BMP-2 (0.05 mg/mL) had an osteoinductive effect with accelerated mineralization in a rabbit sinus model using a BCP carrier.

Overexpression of PLAP-1 in bone marrow stromal cells inhibits the rat critical-size skull defect repair.

Periodontal ligament-associated protein-1 (PLAP-1) is an osteogenisis negative regulatory gene that inhibits the differentiation of rat bone marrow stromal cells (rBMSCs) into osteoblasts in vitro. The aim of this study was to verify whether PLAP-1 acted as an inhibitory factor of rBMSCs in rat critical-size skull defect repair in vivo. Six-week old male Wistar rats treated with a drill-hole injury were randomly assigned into five groups PLAP-1-transfected rBMSC group, empty vector-transfected rBMSC group, normal rBMSC group, collagen group and blank control group according to the treatment factors. Skull specimens were obtained at 8 weeks after surgery. X-ray examination, histological observation of hard tissue slices (HE, Masson staining and von Kossa staining), immunohistochemical staining were applied to evaluate the repair effects. X-ray examination showed that repair effect of the PLAP-1-transfected rBMSC group was significantly worse than that of the empty vector-transfected rBMSC group and normal rBMSC group. In HE staining, regenerated bone could only be observed in the partial defect area of the PLAP-1-transfected rBMSC group. However, new bone filled nearly the entire defects in the empty vector-transfected rBMSC group and normal rBMSC group. Mineralization of new bone in the two groups was significantly higher than that of the PLAP-1-transfected rBMSC group, according to Masson and von Kossa staining. Meanwhile, the transfected PLAP-1 gene worked well in vivo. Positive expression of PLAP-1 protein was only distributed in the newly formed bone of the PLAP-1-transfected rBMSC group, no positive staining was observed in the other four groups. Overexpression of PLAP-1 in rBMSCs inhibits new bone formation and mineralization in rat critical-size skull defects in vivo.

Changes in Bone Quality Associated with the Mineralization of New Bone Formed Around Implants - Using XPS, Polarized Microscopy, and FTIR imaging -

The purpose of this study was to investigate changes in bone quality associated with the mineralization of new bone forming around implants. Eight 18-week-old male New Zealand white rabbits and implants fabricated from Ti-15%Zr-4%Nb-4%Ta alloy were used in the study. Characteristics of new bone formed around implants at 4 and 8 weeks and cortical bone were compared using X-ray photoelectron spectroscopy (XPS) for elemental qualitative and quantitative analysis, polarized microscopy (PM) for histological examination, and Fourier transform infrared Imaging (FTIR Imaging) to characterize molecular distributions in new bone and cortical bone. Results from XPS analysis showed changes in the chemical bonding in new bone over time (4 weeks vs. 8 weeks). PM showed changes in collagen birefringence, orientation and intensity in new bone over time, becoming more similar to those of cortical bone after 8 weeks. FTIR Imaging showed that in new bone close to cortical bone, amounts of PO 4 3- and CO 3 2- ions (representing mineral or inorganic components), and amounts of amide I (representing the organic component) increased over time as mineralization progressed. In new bone more distant from cortical bone, PO 4 3- , CO 3 2- , and amide I distributions also increased over time, but to a lesser extent to that observed near cortical bone. Our findings clearly showed changes in the quality of new bone forming around implants, and these changes were related to mineralization.

Effects of intermittent intravenous ibandronate injections on bone quality and micro-architecture in women with postmenopausal osteoporosis: The DIVA study

In the Dosing IntraVenous Administration (DIVA) study, IV ibandronate injections (15–30 s duration) provided significantly greater gains in bone mineral density than daily oral ibandronate (P<0.001). Single transiliac bone biopsy was performed in a subgroup of women (n=109/1395) from DIVA to assess the impact of ibandronate on newly formed bone and bone remodeling. Patients received ibandronate IV injections 2 mg every 2 months, 3 mg every 3 months or oral ibandronate 2.5 mg daily, plus oral or IV placebo, as appropriate to maintain blinding. Of the 1395 participants from the DIVA study, 122 were enrolled in the substudy. Qualitative histological analysis was performed on all biopsy cores and 89 cores were considered to be evaluable for quantitative histomorphometry. Following 2 years of ibandronate treatment, trabecular bone maintained its normal lamellar structure with no evidence of woven bone, marrow fibrosis, cellular toxicity, or other qualitative abnormalities. Primary mineralization of new bone remained normal, as indicated by the slightly lower osteoid thickness and osteoid volume, with normal mineral apposition rate compared to healthy, postmenopausal women. Mineralizing surface, osteoid surface, activation frequency and bone formation rate were decreased in all ibandronate-treated groups compared with values from healthy, postmenopausal women. Specifically, the bone formation rate (BFR/BV and BFR/BS) was approximately 5 times lower in the ibandronate-treated (3 mg) group than in healthy, postmenopausal women. Histomorphometric analysis of transiliac bone biopsies demonstrated normal micro-structure of newly formed bone with normal mineralization and reduced remodeling after oral or IV ibandronate.

Analysis of callus pattern of tibia lengthening in achondroplasia and a novel method of regeneration assessment using pixel values

To relate morphology of new bone formation to outcome after tibial lengthening performed in patients with achondroplasia. A retrospective analysis of 60 tibial segments in 30 achondroplasia patients was performed. There were 22 female patients and eight male patients, with a mean age of 9.8 years. New bone formation was classified by shape, homogeneity and density. Pixel values in relation to original bone were measured using a picture-archiving communication system (PACS). Clinical outcome was described by the external fixator and maturation indices. Mean lengthening was 9.2 cm (range 3-12.7 cm). The mean external fixator index was 23.4 (range 15.1-50). The mean maturation index was 12.3 days/cm (range 6-40 days/cm). Homogeneous pathways were associated with the best clinical results (fixator index 20.4, maturation index 10.8), followed by heterogeneous pathway (external fixator index 26.5, maturation index 16.8) and radiolucent pathway (fixator index 31.2, maturation index 21.4). Both cylindrical (external fixator index 25.2, maturation index 14.5) and concave (external fixator index 26.6, maturation index 16.3) callus shapes were favourable. Mineralization of new bone became equal to that of normal bone within 16 weeks (mean) for homogeneous pathway, 12 weeks for heterogeneous pathway and 32 weeks for lucent pathway. The type of new bone formation seen on radiographs is related to clinical outcome, with homogeneous pathways being the most favourable ones.

Fluoride effects on bone formation and mineralization are influenced by genetics

Introduction A variation in bone response to fluoride (F −) exposure has been attributed to genetic factors. Increasing fluoride doses (0 ppm, 25 ppm, 50 ppm, 100 ppm) for three inbred mouse strains with different susceptibilities to developing dental enamel fluorosis (A/J, a “susceptible” strain; SWR/J, an “intermediate” strain; 129P3/J, a “resistant” strain) had different effects on their cortical and trabecular bone mechanical properties. In this paper, the structural and material properties of the bone were evaluated to explain the previously observed changes in mechanical properties. Materials and methods This study assessed the effect of increasing fluoride doses on the bone formation, microarchitecture, mineralization and microhardness of the A/J, SWR/J and 129P3/J mouse strains. Bone microarchitecture was quantified with microcomputed tomography and strut analysis. Bone formation was evaluated by static histomorphometry. Bone mineralization was quantified with backscattered electron (BSE) imaging and powder X-ray diffraction. Microhardness measurements were taken from the vertebral bodies (cortical and trabecular bones) and the cortex of the distal femur. Results Fluoride treatment had no significant effect on bone microarchitecture for any of the strains. All three strains demonstrated a significant increase in osteoid formation at the largest fluoride dose. Vertebral body trabecular bone BSE imaging revealed significantly decreased mineralization heterogeneity in the SWR/J strain at 50 ppm and 100 ppm F −. The trabecular and cortical bone mineralization profiles showed a non-significant shift towards higher mineralization with increasing F − dose in the three strains. Powder X-ray diffraction showed significantly smaller crystals for the 129P3/J strain, and increased crystal width with increasing F − dose for all strains. There was no effect of F − on trabecular and cortical bone microhardness. Conclusion Fluoride treatment had no significant effect on bone microarchitecture in these three strains. The increased osteoid formation and decreased mineralization heterogeneity support the theory that F − delays mineralization of new bone. The increasing crystal width with increasing F − dose confirms earlier results and correlates with most of the decreased mechanical properties. An increase in bone F − may affect the mineral–organic interfacial bonding and/or bone matrix proteins, interfering with bone crystal growth inhibition on the crystallite faces as well as bonding between the mineral and organic interface. The smaller bone crystallites of the 129P3/J (resistant) strain may indicate a stronger organic/inorganic interface, reducing crystallite growth rate and increasing interfacial mechanical strength.

Establishment of a model of cortical bone repair in mice.

A model of cortical bone repair has been established for use in mice. The cortical defect consisted of a hole drilled through the entire diameter of the tibial diaphysis. The hematoma that initially filled the drill site was invaded by cells of mesenchymal appearance within 5 days of injury. Trabeculae of mineralized woven bone were present throughout the drill site by day 9. A reaction in the periosteum adjacent to the drill site, consisting of both new bone and cartilage formation, preceded deposition of bone tissue in the drill site. New woven bone was modeled to restore the marrow cavity to normal by 4 weeks after injury, and almost normal cortical structure was achieved by 6 weeks after injury. Immunohistochemical studies indicated that type III collagen was expressed within the drill site by day 5, reached a peak at day 7, and was diminished by day 9. In contrast, type I collagen was first detectable in the drill site at day 7, and staining was more intense by day 9. Osteopontin expression in the drill site coincided with the process of mineralization of new bone in this location. The model of bone repair described here provides a method for inducing reproducible bone lesions in a readily identifiable location in mice. It will be useful in the investigation of bone cell function in mouse strains that have been subjected to genetic manipulation.

Bone turnover model based on a continuously expanding exchangeable calcium pool.

The rate of mineralization of new bone has been worked out on the basis of a continually expanding exchangeable calcium pool. A model is proposed in which the serum specific activity at any time is determined by the combined effects of pool expansion on the one hand, and excretion and mineralization on the other.