μCT Analysis Research Articles

Live and heat-killed Lactobacillus reuteri reduce alveolar bone loss on induced periodontitis in rats

ObjectiveLactobacillus reuteri is reported as a beneficial adjunct to conventional periodontitis treatment or as monotherapy for gingivitis in several clinical trials. However, probiotics are not free from the risk of adverse effects. We investigated the effects of live and heat-killed (paraprobiotic) L. reuteri during the development of ligature-induced periodontitis in rats. MethodsThirty-two male rats were divided into four groups—control group: healthy rats, LIP group: rats with ligature induced periodontitis, LIP/PRO group: rats with periodontitis that received live L. reuteri, LIP/PARA group: rats with periodontitis that received heat-killed L. reuteri. Treatment was administered for 30 days prior to ligation and continued for 14 days until euthanasia. The hemimandibles were removed and submitted to radiographic analysis, microcomputerized microtomography (μCT) and histomorphometric analysis. ResultsRadiographic analysis revealed that pro- and paraprobiotics enhanced the percentage of mesial bone support. The μCT analysis showed that paraprobiotic treatment increased bone volume and trabecular number while decreasing total bone porosity and trabecular separation. Live probiotic led to higher trabecular thickness. In the histomorphometric analysis, paraprobiotics significantly prevented furcation bone loss and collagen breakdown, although both pro- and paraprobiotics similarly reduced mesial bone loss. There were no significant differences in attachment loss or the number of tartrate-resistant acid phosphatase -positive cells across treatments. ConclusionsAlthough viable and heat-killed bacteria reduced mesial alveolar bone loss and ameliorated furcation microarchitecture parameters, paraprobiotics presented the most pronounced results preventing periodontitis development. This can contribute to the future use of paraprobiotics in the prevention and treatment of periodontitis.

X-ray μCT analysis to characterize cork spot disorder in Chinese pear ‘Chili’ (Pyrus bretschneideri)

Abstract Healthy and disordered fruit of Chinese ‘Chili’ (Pyrus bretschneideri Rehd.), a well-known pear cultivar in China, are used to investigate the cork spot disorder by non-destructive X-ray computed microtomography (μCT) scanning technology, with pixel resolution of 67-70 μm and scanning time of 60 min on intact fruit. The results suggested that X-ray μCT could clearly and accurately distinguish the internal organization, size and location of cork spot in the pear fruit. CT images showed that the spots mostly locate in the pulp tissue between the calyx end and the equator of disordered fruit, and usually within 14 mm below the pericarp. STL format 3D images had great potential to distinguish cork spotted fruit core from healthy one. In addition, total number of pores is strongly decreased while larger pores bigger than 900 μm are considerably increased in disordered fruit as compared to healthy fruit, which indicates that larger pores maybe formed by the extension and interconnection with adjacent pores of some small pores. Further analysis in pore skeleton-network shows that extremely higher porosity and highly branched, more connected and thicker pores are observed in cork spotted fruit than those in healthy fruit. In addition, the average length of the healthy pore throat is 160.08 μm, which is significantly smaller than that of disordered fruit (277.91 μm). Moreover, the coordination number of pores in cork spotted fruit is 2, while it is 0 in healthy fruit. All the above results indicate that X-ray μCT provides a new perspective to the non-destructive visualization, quantification and knowledge of the 3D image structure of pear fruit with cork spot, and it is highly essential for early diagnosis and further study of the pathogenic mechanism of this disorder.

The neuropeptide calcitonin gene-related peptide alpha is essential for bone healing.

BackgroundImpaired fracture healing represents an ongoing clinical challenge, as treatment options remain limited. Calcitonin gene-related peptide (CGRP), a neuropeptide targeted by emerging anti-migraine drugs, is also expressed in sensory nerve fibres innervating bone tissue.MethodBone healing following a femoral osteotomy stabilized with an external fixator was analysed over 21 days in αCGRP-deficient and WT mice. Bone regeneration was evaluated by serum analysis, µCT analysis, histomorphometry and genome-wide expression analysis. Bone-marrow-derived osteoblasts and osteoclasts, as well as the CGRP antagonist olcegepant were employed for mechanistic studies.FindingsWT mice with a femoral fracture display increased CGRP serum levels. αCGRP mRNA expression after skeletal injury is exclusively induced in callus tissue, but not in other organs. On protein level, CGRP and its receptor, calcitonin receptor-like receptor (CRLR) complexing with RAMP1, are differentially expressed in the callus during bone regeneration. On the other hand, αCGRP-deficient mice display profoundly impaired bone regeneration characterised by a striking reduction in the number of bone-forming osteoblasts and a high rate of incomplete callus bridging and non-union. As assessed by genome-wide expression analysis, CGRP induces the expression of specific genes linked to ossification, bone remodeling and adipogenesis. This suggests that CGRP receptor-dependent PPARγ signaling plays a central role in fracture healing.InterpretationThis study demonstrates an essential role of αCGRP in orchestrating callus formation and identifies CGRP receptor agonism as a potential approach to stimulate bone regeneration. Moreover, as novel agents blocking CGRP or its receptor CRLR are currently introduced clinically for the treatment of migraine disorders, their potential negative impact on bone regeneration warrants clinical investigation.FundingThis work was funded by grants from the Else-Kröner-Fresenius-Stiftung (EKFS), the Deutsche Forschungsgemeinschaft (DFG), and the Berlin Institute of Health (BIH).

Impact of NSAID etoricoxib on side effects of orthodontic tooth movement

ObjectivesThe non-steroidal anti-inflammatory drug etoricoxib is the most highly selective inhibitor of cyclooxygenase-2 available (344:1) and has been approved for postoperative pain therapy following dental interventions in Europe. At clinically relevant doses it has been reported to only have marginal effects on the velocity of orthodontic tooth movement (OTM). Its effects on associated dental root resorptions, osteoclastogenesis, trabecular number in the alveolar bone and periodontal bone loss during OTM, however, have not yet been investigated. Material and methods40 male Fischer344 rats were divided into four groups: 1.5ml tap water/day p.o. (control, 1), additional 7.8mg/kg/day etoricoxib (normal dose) for three (2) or seven (3) days/week and 13.1mg/kg/day (high dose) for seven days/week, respectively (4). After a week of premedication, OTM in anterior direction of the first left upper molar was performed for 28 days by means of a nickel-titanium coil spring (0.25N). We quantified OTM-associated dental root resorptions, osteoclastogenesis, trabecular number and periodontal bone loss by histomorphometrical, histochemical and μCT analyses of the disected tooth-bearing upper jaw sections. ResultsAfter 28 days of OTM, associated reduction of trabecular number seemed to be slightly alleviated by high doses of etoricoxib, whereas no significant other etoricoxib effects in the doses administered could be detected regarding OTM-induced or -associated dental root resorptions, osteoclastogenesis or periodontal bone loss. ConclusionsDental root resorptions, osteoclastogenesis and periodontal bone loss during OTM in rats were not significantly affected by etoricoxib in the clinically relevant dosages investigated with only a slight inhibitory effect on bone remodelling to be expected at high dosages. Etoricoxib is therefore not suitable for the prevention of these detrimental effects, but could be a suitable analgesic during OTM, as it has been reported not to affect tooth movement.

Abstract LB-212: Circadian melatonin signal disruption by exposure to artificial light at night promotes bone lytic breast cancer metastases

Abstract Breast cancer (BC) metastasis to bone is most common in patients with advanced metastatic BC. Erosion of bone by BC metastases increases bone fragility, risk of fracture and mortality in BC patients. Bone metastatic BC cannot be surgically removed and can only be treated with chemotherapy and/or radiation therapy. Circadian rhythms are daily cycles of ~24 h that control most physiologic processes, disruption of these processes by exposure to artificial light at night (LAN) has been shown to be strongly associated with the development of cancer, particularly breast cancer. Disruption (suppression) of melatonin (MLT) production by LAN is considered as a risk factor for BC. The present study addressed the hypothesis that circadian MLT disruption by dim LAN (dLAN) promotes osteolytic bone metastatic BC. Our research shows that the disruption of the anti-cancer circadian hormone MLT signaling by exposure to dLAN can significantly enhance the metastatic potential of BC cells. This supports the report of the International Agency for Research on Cancer that night shift work is a “probable human carcinogen” and highlights the association between exposure to LAN and invasive BC. We have previously reported that human breast tumor xenografts grown in athymic nude female rats housed in a photoperiod of 12h light at day:12h dLAN (suppressed melatonin signal), display resistance to doxorubicin (Dox). More importantly, tumor growth and drug resistance could be blocked by the administration of Dox in circadian alignment with nocturnal MLT. Our present results show that moderately metastatic luminal A estrogen receptor alpha (ERα) positive MCF-7 BC cells, when inoculated into the tibia (to mimic bone metastatic disease) of MLT-producing Foxn1nu athymic nude mice and housed in dLAN (suppressed nocturnal melatonin production), developed robust bone metastatic tumors (P<0.05) that were highly osteolytic (P<0.05) as determined by IVIS bioluminescent imaging, x-ray and µCT analysis. Administration of nighttime MLT at physiological levels was able to partially reduce the bone metastatic tumor burden. The MLT receptor (MT1 and MT2) antagonist luzindole blocked the inhibitory effect of nighttime MLT on bone metastatic tumor growth supporting a receptor-mediated mechanism. These findings demonstrate for the first time the importance of intact nighttime MLT anti-cancer signaling in suppressing bone metastatic breast tumor growth. Our results provide a potentially strong conceptual rationale for the development of a unique clinical strategy to improve the survival rates of bone metastatic BC patients with the use of circadian-optimized MLT therapy. Acknowledgements: Supported in part by U54 GM104940 from the National Institute of General Medical Sciences of the NIH, which funds the Louisiana Clinical and Translational Science Center (LACATS). Citation Format: Muralidharan Anbalagan. Circadian melatonin signal disruption by exposure to artificial light at night promotes bone lytic breast cancer metastases [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr LB-212.

Assessment of bone healing using (Ti,Mg)N thin film coated plates and screws: Rabbit femur model.

Magnesium (Mg) based implants such as plates and screws are often preferred to treat bone defects because of the positive effects of magnesium in bone growth and healing. Their low corrosion resistance, however, leads to fast degradation and consequently failure before healing was completed. Previously, we developed Mg doped titanium nitrate (TiN) thin film coatings to address these limitations and demonstrated that <10 at% Mg doping led to enhanced mineralization in vitro. In the present study, in vivo performance of (Ti,Mg)N coated Ti6Al4V based plates and screws were studied in the rabbit model. Bone fractures were formed on femurs of 16 rabbits and then fixed with either (Ti,Mg)N coated (n = 8) or standard TiN coated (n = 8) plates and screws. X-ray imaging and μCT analyses showed enhanced bone regeneration on fracture sites fixed with (Ti,Mg)N coated plates in comparison with the Mg free ones. Bone mineral density, bone volume, and callus volume were also found to be 11.4, 23.4, and 42.8% higher, respectively, in accordance with μCT results. Furthermore, while TiN coatings promoted only primary bone regeneration, (Ti,Mg)N led to secondary bone regeneration in 6 weeks. These results indicated that Mg presence in the coatings accelerated bone regeneration in the fracture site. (Ti,Mg)N coating can be used as a practical method to increase the efficiency of existing bone fixation devices of varying geometry.

Radix Rehmanniae Praeparata promotes bone fracture healing through activation of TGF-β signaling in mesenchymal progenitors

BackgroundRadix Rehmanniae Praeparata (RR), the steamed roots of Rehmannia glutinosa, is a traditional Chinese medicine with the function of kidney-nourishing, and it has been safety used for centuries to treat bone-related disorders. The aim of this study is to investigate the positive effect and underlying mechanism of RR enhancing bone fracture healing in mouse model. MethodsTen-week-old C57BL/6J mice were subjected to a unilateral open transverse tibial fracture and provided a daily treatment of RR. Bone samples were harvested for tissue analyses including x-ray, μCT, histology, histomorphometry, biomechanical testing, immunohistochemical (IHC) and quantitative gene expression analysis. To determine the role of TGF-β in accelerating fracture healing effect of RR, aforementioned experiments were performed on Gli1-CreER; Tgfbr2 flox/flox (Tgfbr2Gli1ER) conditional knockout mice. ResultsRR promoted bone fracture healing and strengthened bone intensity in wild-type and Cre− mice with the activation of TGF-β/Smad2 signaling, on the contrary, RR failed to accelerating fracture healing in Tgfbr2Gli1ER mice. ConclusionRR promotes bone fracture healing by intensify the contribution of Gli1+ cells on bone and cartilage formation mainly in TGF-β-dependent manner. RR is an alternative option for clinical treatment of fracture.

The loss of STAT3 in mature osteoclasts has detrimental effects on bone structure.

Signal Transducer and Activator of Transcription 3 (STAT3) has recently been shown to be involved in bone development and has been implicated in bone diseases, such as Job’s Syndrome. Bone growth and changes have been known for many years to differ between sexes with male bones tending to have higher bone mass than female bones and older females tending to lose bone mass at faster rates than older males. Previous studies using conditional knock mice with Stat3 specifically deleted from the osteoblasts showed both sexes exhibited decreased bone mineral density (BMD) and strength. Using the Cre-Lox system with Cathepsin K promotor driving Cre to target the deletion of the Stat3 gene in mature osteoclasts (STAT3-cKO mice), we observed that 8-week old STAT3-cKO female femurs exhibited significantly lower BMD and bone mineral content (BMC) compared to littermate control (CN) females. There were no differences in BMD and BMC observed between male knock-out and male CN femurs. However, micro-computed tomography (μCT) analysis showed that both male and female STAT3-cKO mice had significant decreases in bone volume/tissue volume (BV/TV). Bone histomorphometry analysis of the distal femur, further revealed a decrease in bone formation rate and mineralizing surface/bone surface (MS/BS) with a significant decrease in osteoclast surface in female, but not male, STAT3-cKO mice. Profiling gene expression in an osteoclastic cell line with a knockdown of STAT3 showed an upregulation of a number of genes that are directly regulated by estrogen receptors. These data collectively suggest that regulation of STAT3 differs in male and female osteoclasts and that inactivation of STAT3 in osteoclasts affects bone turnover more in females than males, demonstrating the complicated nature of STAT3 signaling pathways in osteoclastogenesis. Drugs targeting the STAT3 pathway may be used for treatment of diseases such as Job’s Syndrome and osteoporosis.

Evaluation of synthetic ceramics as compression resistant matrix to promote osteogenesis of autologous blood coagulum containing recombinant human bone morphogenetic protein 6 in rabbit posterolateral lumbar fusion model.

Posterolateral lumbar fusion (PLF) is a commonly performed surgical procedure for the treatment of pathological conditions of the lumbosacral spine. In the present study, we evaluated an autologous bone graft substitute (ABGS) containing rhBMP6 in autologous blood coagulum (ABC) and synthetic ceramics used as compression resistant matrix (CRM) in the rabbit PLF model. In the pilot PLF rabbit experiment, we tested four different CRMs (BCP 500-1700 μm, BCP 1700-2500 μm and two different TCP in the form of slabs) which were selected based on achieving uniform ABC distribution. Next, ABGS implants composed of 2.5 mL ABC with 0.5 g ceramic particles (TCP or BCP (TCP/HA 80/20) of particle size 500-1700 μm) and 125 μg rhBMP6 (added to blood or lyophilized on ceramics) were placed bilaterally between transverse processes of the lumbar vertebrae (L5-L6) following exposition and decortication in 12 New Zealand White Rabbits observed for 7 weeks following surgery. Spinal fusion outcome was analysed by μCT, palpatory segmental mobility testing and selected specimens were either tested biomechanically (three-point bending test) and/or processed histologically. The total fusion success rate was 90.9% by both μCT analyses and by palpatory segmental mobility testing. The volume of newly formed bone between experimental groups with TCP or BCP ceramics and the different method of rhBMP6 application was comparable. The newly formed bone and ceramic particles integrated with the transverse processes on histological sections resulting in superior biomechanical properties. The results were retrospectively found superior to allograft devitalized mineralized bone as a CRM as reported previously in rabbit PLF. Overall, this novel ABGS containing rhBMP6, ABC and the specific 500-1700 μm synthetic ceramic particles supported new bone formation for the first time and successfully promoted posterolateral lumbar fusion in rabbits.

Quantitative µ-CT Analysis of Scale Topology Formed During Oxidation of High SiMo Cast Iron

High SiMo cast iron components in automotive exhaust systems are exposed to high-temperature oxidation over time. Quantitative analysis of formed oxide scale is therefore important for the assessment of a component durability. The brittle nature of multilayered scale and thermal stress limits capture of a true topology using traditional 2D destructive cut and polish methods. In this study, nondestructive high spatial-resolution 3D µCT analysis was performed on 2.90-mm-diameter oxidized specimens which permitted direct observation with 3.5 µm pixel resolution. The specimens were oxidized in three sequential time steps for a total 100 h at 700 °C and 800 °C in air and combustion gas atmospheres. A MATLAB-coded algorithm was used to quantify the topology, thickness variation in internal and external scale layers, and scale/metal interface unevenness. Scale topology was linked to oxidation temperature and gas atmosphere. A water vapor environment increases scale/metal interface unevenness and scale layer thickness irregularity which were related to an accelerated oxidation rate.

Understanding decay in marine calcifiers: Micro‐CT analysis of skeletal structures provides insight into the impacts of a changing climate in marine ecosystems

AbstractCalcifying organisms and their exoskeletons support some of the most diverse and economically important ecosystems in our oceans. Under a changing climate, we are beginning to see alterations to the structure and properties of these exoskeletons due to ocean acidification, warming and accelerated rates of bioerosion. Our understanding has grown as a result of using micro‐computed tomography (µCT) but its applications in marine biology have not taken full advantage of the technological development in this methodology. We present a significant advancement in the use of this method to studying decalcification in a marine calcifier.We present a detailed workflow on best practice for µCT image processing and analysis of marine calcifiers, designed using coral skeletons subjected to acute, short‐term microbial bioerosion. This includes estimating subresolution microporosity and describing pore space morphological characteristics of macroporosity, in perforate and imperforate exoskeletons. These metrics are compared between control and bieroded samples, and are correlated with skeletal hardness as measured by nanoindentation.Our results suggest that using subresolution microporosity analysis improves the spatiotemporal resolution of µCT data and can detect changes not seen in macroporosity, in both perforate and imperforate skeletons. In imperforate samples, the mean size and relative number of pores in the macroporous portion of the images changed significantly where total macroporosity did not. The increased number of pores and higher microporosity are both directly related to a physical weakening of the calcareous exoskeletons of imperforate corals only. In perforate corals, increased macroporosity was accompanied by an overall widening of pore spaces though this did not correlate with sample hardness.These novel techniques complement traditional approaches and in combination demonstrate the potential for using µCT scanning to sensitively track the process of decalcification from a structural and morphological perspective. Importantly, these approaches do not necessarily rely on ultra‐high resolution (i.e. single micron) scans and so maintain the accessibility of this technology. The continued optimization of these tools for a variety of marine calcifiers will advance our understanding of the effect of climate change on marine biogenic calcified structures.

Macrolactin A protects against LPS-induced bone loss by regulation of bone remodeling

An imbalance between bone resorption and bone formation leads to several kinds of bone diseases such as rheumatoid arthritis, osteoporosis and Paget’s disease. The imbalance between bone formations relative to bone resorption is responsible in bone remodeling. Several studies have suggested that macrolactin A (MA) has potent anti-inflammatory, anti-cancer and anti-angiogenic effects in various cell types. We investigate whether macrolactin A (MA) could inhibit bone loss and enhance bone formation. We used bone marrow monocytes/macrophages (BMMs) cells to study osteoclast activity and MC3T3-E1 cells to study osteoblast activity. MA suppressed tartrate resistant acid phosphatase (TRAP) positive multinucleated cells in a concentration-dependent manner, as well as at a specific time point. MA markedly reduced bone resorption activity and F-actin ring formation. Moreover, MA markedly suppressed receptor activator of nuclear factor k-B ligand (RANKL)-induced osteoclastogenic marker genes and transcription factors in-vitro. MA repressed osteoclast differentiation via activation of the phosphoinositide kinase-3/Akt, extracellular signal-regulated kinase 1/2 (ERK 1/2), c-Jun N-terminal kinase (JNK), nuclear factor of activated T cells, cytoplasmic 1 (NFATc1) and c-Fos signaling pathways. MA enhanced pre-osteoblast cell differentiation on mineralization activity, alkaline phosphatase (ALP) activity, and the expression of osteoblastogenic markers including osterix, RUNX-2, SMAD4, BMP-2, and ALP. Importantly, MA repressed lipopolysaccharide (LPS)-induced inflammatory bone loss in mice as shown by TRAP staining of femurs and μCT analysis. Therefore, MA could be a promising candidate for the inhibition and management of osteoporosis, arthritis, and bone lytic diseases.

Variability in the number of infraorbital foramina in rhesus macaques (Macaca mulatta) and cynomolgus macaques (Macaca fascicularis).

This study aimed to determine the number of infraorbital foramina in monkeys of the Papionini tribe. The authors performed a μCT analysis of the morphology of the infraorbital foramina. A total number of 52 simian skulls belonged to two macaque species: Macaca mulatta and Macaca fascicularis were used in the study. The number of infraorbital foramina was counted macroscopically and with the use of a magnifying glass. Next, the skull representing the most common morphological type was selected and scanned by micro-computed tomography (μCT). The Shapiro-Wilk normality test was used in the study. To compare the differences in the number of infraorbital foramen between species, sex and sides, the Mann-Whitney U test was applied. Three infraorbital foramina were present in most individuals from the test group. The Mann-Whitney test revealed no statistically significant difference between the number of foramina on the right- and left-hand side. Likewise, no statistically significant differences between the numbers of infraorbital foramina across sexes were observed. Volumetric reconstructions revealed the presence of separate infraorbital canals for each infraorbital foramen. Craniofacial innervation in macaques is formed by complex branching patterns of cranial nerves. Variability in the number of infraorbital foramina suggests a variable maxillary innervation pattern in these animals. Based on the analysis of volumetric projections, the presence of two labial branches and a single nasal branch of the infraorbital nerve is suggested. Detailed descriptions are supported by quantitative data and μCT evidence.

Acid bone lysates reduce bone regeneration in rat calvaria defects.

Acid bone lysates (ABLs) represent the growth factors and other molecules released during autologous graft resorption. However, the impact of these bone‐derived growth factors on the healing of bone defects has not yet been investigated. The aim of the present study was, therefore, to examine the impact of ABLs adsorbed to collagen membranes on bone regeneration. To this end, in 16 female Sprague Dawley rats, a standardized 5‐mm‐diameter critical size defect on the calvarial bone was created. The defects were covered with collagen membranes that had been soaked either in serum‐free media or ABLs followed by lyophilization. After a healing period of 4 weeks, micro‐computed tomography (μCT) and histological analyses by means of undecalcified thin ground sections were performed. μCT analysis of the inner 4 mm of the calvaria defect showed a greater bone defect coverage in the control group when compared to ABL group, 29.8% (confidence interval [CI]: 17.7–50.3) versus 5.6% (CI: 1.0–29.8, p = .03), respectively. Moreover, we found significantly more absolute bone volume (BV) in the control group when compared to ABL group, 0.59 mm3 (CI: 0.27–1.25) versus 0.07 mm3 (CI: 0.06–0.59, p = .04), respectively. Histomorphometry confirmed these findings with a relative BV in the central compartment of 14.1% (CI: 8.4–20.6) versus 5.6% (CI: 3.4–7.9, p = .004), respectively. These findings indicate that bone‐derived growth factors contained in ABLs are able to attenuate bone regeneration within collagen membranes.

Pre-natal manifestation of systemic developmental abnormalities in spinal muscular atrophy.

Spinal muscular atrophy (SMA) is a neuromuscular disease caused by mutations in survival motor neuron 1 (SMN1). SMN-restoring therapies have recently emerged; however, preclinical and clinical studies revealed a limited therapeutic time window and systemic aspects of the disease. This raises a fundamental question of whether SMA has presymptomatic, developmental components to disease pathogenesis. We have addressed this by combining micro-computed tomography (μCT) and comparative proteomics to examine systemic pre-symptomatic changes in a prenatal mouse model of SMA. Quantitative μCT analyses revealed that SMA embryos were significantly smaller than littermate controls, indicative of general developmental delay. More specifically, cardiac ventricles were smaller in SMA hearts, whilst liver and brain remained unaffected. In order to explore the molecular consequences of SMN depletion during development, we generated comprehensive, high-resolution, proteomic profiles of neuronal and non-neuronal organs in SMA mouse embryos. Significant molecular perturbations were observed in all organs examined, highlighting tissue-specific prenatal molecular phenotypes in SMA. Together, our data demonstrate considerable systemic changes at an early, presymptomatic stage in SMA mice, revealing a significant developmental component to SMA pathogenesis.

Dauricine Protects from LPS-Induced Bone Loss via the ROS/PP2A/NF-κB Axis in Osteoclasts.

Dauricine (DAC), an isoquinoline alkaloid, exhibits anti-inflammatory activity. We hypothesized that DAC may prevent the inflammatory bone loss induced by lipopolysaccharide (LPS). LPS-induced bone loss was decreased by DAC in female C57BL/6J mice as evaluated by micro-computerized tomography (μCT) analysis. In vivo tartrate-resistant acid phosphatase (TRAP) staining showed that the increased number of osteoclasts (OCs) in LPS-treated mice was attenuated by DAC, indicating that DAC exhibited bone sparing effects through acting on OCs. DAC also decreased the differentiation and activity of OCs after LPS stimulation in vitro. LPS-induced cytosolic reactive oxygen species (cROS) oxidized PP2A, a serine/threonine phosphatase, leading to the activation of IKKα/β, followed by the nuclear localization of p65. DAC decreased LPS-induced ROS, resulting in the recovery of the activity of PP2A by reducing its oxidized form. Consequently, DAC reduced the phosphorylation of IKKα/β to block the nuclear localization of p65, which decreased NF-κB activation. Taken together, DAC reduced the differentiation and activity of OCs by decreasing ROS via the ROS/PP2A/NF-κB axis, resulting in protection from LPS-induced bone loss. We have demonstrated that LPS-induced bone loss was inhibited by DAC via its action on OCs, implying the therapeutic potential of DAC against inflammatory bone loss.

Combination of micro-computed X-ray tomography and electronic microscopy to understand the influence of graphene nanoplatelets on the thermal conductivity of epoxy composites

An innovative combination of micro-computed X-ray tomography (μCT) and electron microscopy (SEM) coupled to an image-processing software was used to study the thermal conductivity of graphene nanoplatelets(GNP)/epoxy composites. This approach afforded a precise and systematic analysis of the dispersion state of the fillers in the epoxy matrix. Here, the influence of GNP aspect ratio on thermal conductivity enhancement (TCE) and the relationship between dispersion state of the fillers and TCE is revealed. The aggregation level of GNP was finely tuned by varying the sonication power. An unexpected trend towards enhanced thermal conductivity upon aggregation of the fillers is demonstrated. Moreover, μCT analysis revealed the encapsulation of micro-voids, undetectable with conventional methods, by large fillers aggregates. This study demonstrates that aggregation of the fillers increases the thermal conductivity of epoxy composites as long as it does not lead to voids encapsulation.

Regeneration of Secondary Dentin Using Recombinant Human Platelet-Derived Growth Factor and MTA for Pulp Capping: A Randomized Controlled Human Clinical Trial.

This in vivo study assessed the effect of mineral trioxide aggregate (MTA) as a matrix carrier for recombinant human platelet-derived growth factor (rhPDGF) and enamel matrix protein (EMP) on pulp tissue healing following pulp capping. Eighteen intact human premolars scheduled for extraction were included. Coronal access and pulpotomy were performed, and each tooth was left exposed to the oral cavity for 1 hour before pulp capping was performed. Teeth were randomly assigned to one of the following pulp-capping groups (n = 6 each): Group 1 (MTA only); Group 2 (MTA+EMP); or Group 3 (MTA+rhPDGF). Coronal access cavities were then sealed. Immediate preoperative, postoperative, and 4-month follow-up radiographs were taken. At 4 months, the teeth were extracted atraumatically, and histomorphometric and micro-computed tomography (μCT) analyses were performed. Group 1 showed a thin, uneven, irregular dentin-like structure. Its average thickness was 0.3 ± 0.084 mm measured histologically and 0.29 ± 0.091 mm measured by μCT. Group 2 showed of a nonporous, even-thickness dentin-like structure with multiple root-canal obliterations. Highly dense, atubular dentin-like structures associated with presence of odontoblastic lacunae were seen. The structure's average thickness was 0.87 ± 0.09 mm (histologically) and 0.81 ± 0.17 mm (μCT). Group 3 showed a thick and complete 3D continuous seal of newly formed dentin-like structure covering the pulpal space. It resembled secondary dentin in form, porosity, and tubular structural organization, and its average thickness was 0.94 ± 0.02 mm (histologically) and 0.91 ± 0.09 mm (μCT). Groups 2 and 3 showed higher amounts of newly formed dentin-like structure, that was also thicker, than Group 1 (P < .05). No statistically significant differences in structure thickness were found between Groups 2 and 3. The nature of the structure can differ if rhPDGF or EMP is added to MTA for pulp-capping purposes. Combination of rhPDGF and MTA resulted in a newly formed structure resembling secondary dentin, whereas a combination of EMP and MTA produced a nonporous, highly dense dentinal-like structure associated with significant root-canal obliterations.

X-ray Micro-Computed Tomography: An Emerging Technology to Analyze Vascular Calcification in Animal Models.

Vascular calcification describes the formation of mineralized tissue within the blood vessel wall, and it is highly associated with increased cardiovascular morbidity and mortality in patients with chronic kidney disease, diabetes, and atherosclerosis. In this article, we briefly review different rodent models used to study vascular calcification in vivo, and critically assess the strengths and weaknesses of the current techniques used to analyze and quantify calcification in these models, namely 2-D histology and the o-cresolphthalein assay. In light of this, we examine X-ray micro-computed tomography (µCT) as an emerging complementary tool for the analysis of vascular calcification in animal models. We demonstrate that this non-destructive technique allows us to simultaneously quantify and localize calcification in an intact vessel in 3-D, and we consider recent advances in µCT sample preparation techniques. This review also discusses the potential to combine 3-D µCT analyses with subsequent 2-D histological, immunohistochemical, and proteomic approaches in correlative microscopy workflows to obtain rich, multifaceted information on calcification volume, calcification load, and signaling mechanisms from within the same arterial segment. In conclusion we briefly discuss the potential use of µCT to visualize and measure vascular calcification in vivo in real-time.

Pore characterization of PM Mg–0.6Ca alloy and its degradation behavior under physiological conditions

Several material parameters affect degradation characteristics of Mg and its alloys under physiological conditions. Porous Mg materials are interesting for their simultaneous degradation and drug delivery capabilities. However, an increase in pore surface area is detrimental to both degradation resistance and subsequent mechanical properties. The present work aims at determining the threshold porosity value in Mg–0.6Ca specimens produced by powder metallurgy (PM) below which low degradation rates persist with acceptable mechanical properties. Seven different porous Mg–0.6Ca specimens containing both closed and open pore structures were fabricated with porosities ranging from 3% to 21%. Degradation profiles were obtained via a semi static immersion test over 16 days under physiological conditions using Dulbecco's modified Eagle's medium with Glutamax and 10% fetal bovine serum as supplements. The results are related to morphological pore parameters like pore size distribution, pore interconnectivity and pore curvatures that were quantified using an ex situ µCT analysis. In general, with decreasing porosity a decrease in pore interconnectivity is seen followed by rounding of the pores. Low degradation rates (MDR < 0.3 mm/year) are observed in specimens until 10% porosity, however, the upper bound for reproducible degradation is observed to be in specimens until 12% porosity. This porosity level also marks the transition from closed to open pore nature with a simultaneous change in pore interconnectivity from less than 10% to greater than 95%, below and above this porosity level, respectively. The tensile strength and elongation to failure recorded for specimens with 10% porosity were 70 MPa and 2%, respectively displaying positive traits of both homogenous degradation and mechanical properties. The results suggest that high pore interconnectivity is the dominant factor controlling degradation and mechanical properties in porous Mg-0.6Ca specimens. The results also indicate a good sintering response of Mg-0.6Ca specimens providing further material development towards biomaterial applications.