Effects of processing and gamma radiation on mechanical properties and organic composition of frozen, freeze-dried and demineralised human cortical bone allograft.

Bone processing and radiation were reported to influence mechanical properties of cortical bones due in part to structural changes and denaturation of collagen composition. This comparative study was to determine effects of bone processing on mechanical properties and organic composition, and to what extent the radiation damaging after each processing. Human femur cortical bones were processed by freezing, freeze-drying and demineralisation and then gamma irradiated at 5, 15, 20, 25 and 50 kGy. In the compression test, freeze drying significantly decreased the Young's Modulus by 15%, while demineralisation reduced further by 90% (P < 0.05) when compared to the freezing. Only demineralisation significantly reduced ultimate strength of bone by 93% (P < 0.05). In the bending test, both freeze drying and demineralisation significantly reduced the ultimate strength and the work to failure. Radiation at 25 kGy showed no effect on compression for ultimate strength in each processing group. However, high dose of 50 kGy significantly reduced bending ultimate strength by 47% in demineralisation group. Alterations in collagen in bones irradiated at 25 and 50 kGy showed by the highest peak of the amide I collagen in the Fourier Transfer Infra-Red spectra indicating more collagen was exposed after calcium was removed in the demineralised bone, however radiation showed no effect on the collagen crosslink. The study confirmed that demineralisation further reduced the ability to resist deformation in response to an applied force in freeze-dried bones due to calcium reduction and collagen composition. Sterilisation dose of 25 kGy has no effect on mechanical properties and collagen composition of the processed human cortical bone.

Effects of long-term plate fixation with different fixation modes on the radial cortical bone in dogs.

The aim of this study was to examine the effect of long-term locking plate fixation on the cortical bone of the canine radius. Locking compression plates were fixed to the left and right radius in dogs (n = 3). The left radius was fixed with a locking head screw (Locking Plate group, LP). The locking compression plate was compressed periosteally in the right radius using a cortex screw (Compression Plate group, CP). Radial bones from dogs that were euthanized for other purposes were collected as an untreated control group (Control group). After euthanasia at 36 weeks following plate fixation, radial bones were evaluated for bone mineral density and underwent histological analysis. Bone metabolic markers were analyzed by quantitative polymerase chain reaction (qPCR). Statistical analyses were performed for comparisons between groups. The LP group showed no significant difference in bone mineral density after plate fixation, whereas the CP group showed significantly lower bone mineral density. Histological analysis indicated that the number of osteoclasts and rate of empty lacunae increased significantly in the CP group relative to the Control and LP groups. qPCR analysis indicated increased expression of inflammatory cytokines, such as tumor necrosis factor-alpha, interleukin-6, and tumor necrosis factor ligand superfamily member 11 in the CP group, whereas Runt-related transcription factor 2, an osteoblast marker, was similar in all groups. The expression of hypoxia-inducible factor-1α in the CP group was also increased relative to that in the Control and LP groups. Thus, locking plate fixation is a biologically superior fixation method that does not cause implant-induced osteoporosis in the bone in the long term.

Is the 0.2%-Strain-Offset Approach Appropriate for Calculating the Yield Stress of Cortical Bone?

The 0.2% strain offset approach is mostly used to calculate the yield stress and serves as an efficient method for cross-lab comparisons of measured material properties. However, it is difficult to accurately determine the yield of the bone. Especially when computational models require accurate material parameters, clarification of the yield point is needed. We tested 24 cortical specimens harvested from six bovine femora in three-point bending mode, and 11 bovine femoral cortical specimens in the tensile mode. The Young's modulus and yield stress for each specimen derived from the specimen-specific finite element (FE) optimization method was regarded as the most ideal constitutive parameter. Then, the strain offset optimization method was used to find the strain offset closest to the ideal yield stress for the 24 specimens. The results showed that the 0 strain offsets underestimated (- 25%) the yield stress in bending and tensile tests, while the 0.2% strain offsets overestimated the yield stress (+ 65%) in three-point bending tests. Instead, the yield stress determined by 0.007 and 0.05% strain offset for bending and tensile loading respectively, can effectively characterize the biomechanical responses of the bone, thereby helping to build an accurate FE model.

Quantitative Magnetic Resonance Imaging of Cortical and Trabecular Bone.

Bone is a composite material consisting of mineral, organic matrix, and water. Water in bone can be categorized as bound water (BW), which is bound to bone mineral and organic matrix, or as pore water (PW), which resides in Haversian canals as well as in lacunae and canaliculi. Bone is generally classified into two types: cortical bone and trabecular bone. Cortical bone is much denser than trabecular bone that is surrounded by marrow and fat. Magnetic resonance (MR) imaging has been increasingly used for noninvasive assessment of both cortical bone and trabecular bone. Bone typically appears as a signal void with conventional MR sequences because of its short T2*. Ultrashort echo time (UTE) sequences with echo times 100 to 1,000 times shorter than those of conventional sequences allow direct imaging of BW and PW in bone. This article summarizes several quantitative MR techniques recently developed for bone evaluation. Specifically, we discuss the use of UTE and adiabatic inversion recovery prepared UTE sequences to quantify BW and PW, UTE magnetization transfer sequences to quantify collagen backbone protons, UTE quantitative susceptibility mapping sequences to assess bone mineral, and conventional sequences for high-resolution imaging of PW as well as the evaluation of trabecular bone architecture.

Comparison of small-angle neutron and X-ray scattering for studying cortical bone nanostructure.

In this study, we present a combined small-angle neutron and X-ray scattering (SANS and SAXS) study of the nanoscale structure of cortical bone specimens from three different species. The variation of the scattering cross section of elements across the periodic table is very different for neutrons and X-rays. For X-rays, it is proportional to the electron density while for neutrons it varies irregularly with the atomic number. Hence, combining the two techniques on the same specimens allows for a more detailed interpretation of the scattering patterns as compared to a single-contrast experiment. The current study was performed on bovine, porcine and ovine specimens, obtained in two perpendicular directions with respect to the main axis of the bone (longitudinal and radial) in order to maximise the understanding of the nanostructural organisation. The specimens were also imaged with high resolution micro-computed tomography (micro-CT), yielding tissue mineral density and microstructural orientation as reference. We show that the SANS and SAXS patterns from the same specimen are effectively identical, suggesting that these bone specimens can be approximated as a two-component composite material. Hence, the observed small-angle scattering results mainly from the mineral-collagen contrast, apart from minor features associated with the internal collagen structure.

Detection and imaging of gadolinium accumulation in human bone tissue by micro- and submicro-XRF.

Gadolinium-based contrast agents (GBCAs) are frequently used in patients undergoing magnetic resonance imaging. In GBCAs gadolinium (Gd) is present in a bound chelated form. Gadolinium is a rare-earth element, which is normally not present in human body. Though the blood elimination half-life of contrast agents is about 90 minutes, recent studies demonstrated that some tissues retain gadolinium, which might further pose a health threat due to toxic effects of free gadolinium. It is known that the bone tissue can serve as a gadolinium depot, but so far only bulk measurements were performed. Here we present a summary of experiments in which for the first time we mapped gadolinium in bone biopsy from a male patient with idiopathic osteoporosis (without indication of renal impairment), who received MRI 8 months prior to biopsy. In our studies performed by means of synchrotron radiation induced micro- and submicro-X-ray fluorescence spectroscopy (SR-XRF), gadolinium was detected in human cortical bone tissue. The distribution of gadolinium displays a specific accumulation pattern. Correlation of elemental maps obtained at ANKA synchrotron with qBEI images (quantitative backscattered electron imaging) allowed assignment of Gd structures to the histological bone structures. Follow-up beamtimes at ESRF and Diamond Light Source using submicro-SR-XRF allowed resolving thin Gd structures in cortical bone, as well as correlating them with calcium and zinc.

Characterization of a prevascularized biomimetic tissue engineered scaffold for bone regeneration.

Significant bone loss due to disease or severe injury can result in the need for a bone graft, with over 500,000 procedures occurring each year in the United States. However, the current standards for grafting, autografts and allografts, can result in increased patient morbidity or a high rate of failure respectively. An ideal alternative would be a biodegradable tissue engineered graft that fulfills the function of bone while promoting the growth of new bone tissue. We developed a prevascularized tissue engineered scaffold of electrospun biodegradable polymers PLLA and PDLA reinforced with hydroxyapatite, a mineral similar to that found in bone. A composite design was utilized to mimic the structure and function of human trabecular and cortical bone. These scaffolds were characterized mechanically and in vitro to determine osteoinductive and angioinductive properties. It was observed that further reinforcement is necessary for the scaffolds to mechanically match bone, but the scaffolds are successful at inducing the differentiation of mesenchymal stem cells into mature bone cells and vascular endothelial cells. Prevascularization was seen to have a positive effect on angiogenesis and cellular metabolic activity, critical factors for the integration of a graft.

Which type of bone releases the most vancomycin? Comparison of spongious bone, cortical powder and cortico-spongious bone.

Bone infections can be challenging to treat and can lead to several surgeries and relapses. When a graft is needed, cavitary bone loss can be grafted with cancellous or cortical bone. Both can be used for grafting. However, the antibiotic releasing capacity of these grafts has not been compared. Which type of bone is best at releasing the most antibiotic has not been well established. The aim of this study was to determine which type of bone is best for antibiotic release when the bone is suffused with antibiotics by the surgeon. The hypothesis is that there would be a difference between the type of bone tested due to different release capacities of cortical and cancellous bone. This was an experimental study. Cortical spongy bone in chips, Spongy bone in chips and demineralized cortical bone powder were compared. For each type of bone, 5 samples were tested. Processed and decontaminated grafts were freeze-dried to be kept at room temperature. The primary endpoint was the amount of vancomycin released by the graft as it affects the concentration of antibiotic around the graft in clinical practice. The procedure for the study consisted of full graft immersion in a vancomycin solution. Then, the liquid was removed with aspiration. In order to measure the quantity of antibiotic released, the bone was put into distilled water in agitation in a heated rocker at 37 °C. After 30 min of soaking, 1 mL of the liquid was removed. The same extraction process was also carried out after 60 min soaking, 2 h, 3 h, 24 h, and 48 h. No differences were found between each type of bone relative to the concentration of vancomycin released at each time of the assessment. There was a significant difference in the weight of the bone with a higher weight for the cortical powder (1.793 g) versus cortical spongy bone and spongy bone (1.154 g and 1.013 g) with a p value < 0.0001. A significant difference was seen in the weight of the bone with vancomycin after the aspiration of the liquid with 3.026 g for cortical powder, 2.140 g and 2.049 g for the cortical spongy bone and the spongy bone with a p value < 0.0001. In daily clinical practice, one can use cancellous bone, cortico-cancellous bone or cortical powder in order to add vancomycin to a bone graft. Our results show the release kinetics of the soaked allografts. With a maximum of 14 mg/mL in the first minutes and a rapid decrease it shows a pattern comparable to antibiotic loaded bone cement. The method used appears favourable for prophylactic use, protecting the graft against contamination at implantation, but is not sufficient for treating chronic bone infection. LEVEL OF EVIDENCE: V.

Sequential Treatment of Estrogen Deficient, Osteopenic Rats with Alendronate, Parathyroid Hormone (1-34), or Raloxifene Alters Cortical Bone Mineral and Matrix Composition.

Anti-resorptive and anabolic treatments can be used sequentially to treat osteoporosis, but their effects on bone composition are incompletely understood. Osteocytes may influence bone tissue composition with sequential therapies because bisphosphonates diffuse into the canalicular network and anabolic treatments increase osteocyte lacunar size. Cortical bone composition of osteopenic, ovariectomized (OVX) rats was compared to that of Sham-operated rats and OVX rats given monotherapy or sequential regimens of single approved anti-osteoporosis medications. Adult female Sprague-Dawley rats were OVX (N = 37) or Sham-OVXd (N = 6). After 2 months, seven groups of OVX rats were given three consecutive 3-month periods of treatment with vehicle (V), h-PTH (1-34) (P), alendronate (A), or raloxifene (R), using the following orders: VVV, PVV, RRR, RPR, AAA, AVA, and APA. Compositional properties around osteocyte lacunae of the left tibial cortex were assessed from Raman spectra in perilacunar and non-perilacunar bone matrix regions. Sequential treatments involving parathyroid hormone (PTH) caused lower mean collagen maturity relative to monotherapies. Mean mineral:matrix ratio was 2.2% greater, mean collagen maturity was 1.4% greater, and mean carbonate:phosphate ratio was 2.2% lower in the perilacunar than in the non-perilacunar bone matrix region (all P < 0.05). These data demonstrate cortical bone tissue composition differences around osteocytes caused by sequential treatment with anti-osteoporosis medications. We speculate that the region-specific differences demonstrate the ability of osteocytes to alter bone tissue composition adjacent to lacunae.

A Review of Bomb Pulse Dating and its Use in the Investigation of Unidentified Human Remains.

In cases where there is limited antemortem information, the examination of unidentified human remains as part of the investigation of long-term missing person's cases is a complex endeavor and consequently requires a multidisciplinary approach. Bomb pulse dating, which involves the analysis and interpretation of 14C concentration, is one technique that may assist in these investigations by providing an estimate of year of birth and year of death. This review examines the technique of bomb pulse dating and its use in the identification of differentially preserved unknown human remains. Research and case studies implementing bomb pulse dating have predominantly been undertaken in the Northern Hemisphere and have demonstrated reliable and accurate results. Limitations were, however, identified throughout the literature. These included the small sample sizes used in previous research/case studies which impacted on the statistical significance of the findings, as well as technique-specific issues. Such limitations highlight the need for future research.

Compositional and mechanical properties of growing cortical bone tissue: a study of the human fibula.

Human cortical bone contains two types of tissue: osteonal and interstitial tissue. Growing bone is not well-known in terms of its intrinsic material properties. To date, distinctions between the mechanical properties of osteonal and interstitial regions have not been investigated in juvenile bone and compared to adult bone in a combined dataset. In this work, cortical bone samples obtained from fibulae of 13 juveniles patients (4 to 18 years old) during corrective surgery and from 17 adult donors (50 to 95 years old) were analyzed. Microindentation was used to assess the mechanical properties of the extracellular matrix, quantitative microradiography was used to measure the degree of bone mineralization (DMB), and Fourier transform infrared microspectroscopy was used to evaluate the physicochemical modifications of bone composition (organic versus mineral matrix). Juvenile and adult osteonal and interstitial regions were analyzed for DMB, crystallinity, mineral to organic matrix ratio, mineral maturity, collagen maturity, carbonation, indentation modulus, indicators of yield strain and tissue ductility using a mixed model. We found that the intrinsic properties of the juvenile bone were not all inferior to those of the adult bone. Mechanical properties were also differently explained in juvenile and adult groups. The study shows that different intrinsic properties should be used in case of juvenile bone investigation.

Verifying measurements of residual calcium content in demineralised cortical bone.

Calcium contents of demineralised human cortical bone determined by titrimetric assay and atomic absorption spectrophotometry technique were verified by comparing to neutron activation analysis which has high recovery of more than 90%. Conversion factors determined from the comparison is necessary to correct the calcium content for each technique. Femurs from cadaveric donors were cut into cortical rings and demineralised in 0.5 M hydrochloric acid for varying immersion times. Initial calcium content in the cortical bone measured by titration was 4.57%, only 21% of the measurement by neutron activation analysis; while measured by atomic absorption spectrophotometer was 13.4%, only 61% of neutron activation analysis. By comparing more readings with the measurements by neutron activation analysis with 93% recovery, a conversion factor of 4.83 was verified and applied for the readings by titration and 1.45 for atomic absorption spectrophotometer in calculating the correct calcium contents. The residual calcium content started to reduce after the cortical bone was demineralised in hydrochloric acid for 8 h and reduced to 13% after 24 h. Using the linear relationship, the residual calcium content could be reduced to less than 8% after immersion in hydrochloric acid for 40 h. Atomic absorption spectrophotometry technique is the method of choice for calcium content determination as it is more reliable compared to titrimetric assay.

Diet and adult age-at-death among mobile foragers: A synthesis of bioarcheological methods.

OBJECTIVES: The research explores whether the combined study of cortical bone histology, bone morphology, and dietary stable isotopes can expand insights into past human health and adaptations, particularly dietary sufficiency and life span. MATERIALS AND METHODS: Midthoracic rib cortices from 54 South African Late Holocene adult skeletons (28 M, 24 F, two sex undetermined) are assessed by transmitted-light microscopy for cross-sectional area measurements, osteon area (On.Ar), osteon population density, and presence/absence of secondary osteon variants. Values for δ13 Cbone collagen , δ15 Nbone collagen , 14 C dates, Southwestern and Southern Cape geographic regions, body size measures, estimated ages-at-death from both morphological and histological methods are integrated into analyses, which include Spearman correlations, χ2 tests and Kruskal-Wallis ANOVAs. RESULTS: There is reduced On.Ar variability with higher δ15 N (r = -.41, p = .005); rib %cortical area and δ15 N are negatively correlated in the Southern Cape group (r = -.60, p = .03). Osteon variants are more common in older adults; histological ages at death are significantly older than those determined from gross morphology. DISCUSSION: We found bone tissue relationships with measures of diet composition, but indicators of dietary adequacy remain elusive. Relationships of tissue quality and isotopes suggest that some Southern Cape adults lived long lives. Osteon variants are associated with age-at-death; some association with diet remains possible. Gross morphological methods appear to underestimate adult ages-at-death, at least among small-bodied adults.

Identification of collagen fibrils in cross sections of bone by electron energy loss spectroscopy (EELS).

Transmission electron microscopic (TEM) images of ion-milled bovid cortical bone cut approximately normal to the axes of fibrils show that mineral occurs in the form of plates surrounding and laying between circular or elliptical features about 50 nm in diameter. The classification of these features as either pores or collagen fibrils is highly debated. Electron energy loss spectroscopy (EELS) mapping of these features in ion milled sections shows that they are lacking significant amounts of mineral or collagen, although their appearance suggests that they are cross sections of collagen fibrils. However, analogous sections prepared using an ultramicrotome show that, while these circular features show reduced concentrations of calcium and phosphorus, some of them contain quantities of carbon and nitrogen in bonding states comparable to the composition of collagen. This work demonstrates that the observed circular features are sections of collagen fibrils, but that bombardment by argon ions during broad beam ion milling destroys the collagen and associated gap-zone mineral.

Analysis of Mechanical Properties and Mechanical Anisotropy in Canine Bone Tissues of Various Ages.

The effect of age on mechanical behavior and microstructure anisotropy of bone is often ignored by researchers engaged in the study of biomechanics. The objective of our study was to determine the variations in mechanical properties of canine femoral cortical bone with age and the mechanical anisotropy between the longitudinal and transverse directions. Twelve beagles divided into three age groups (6, 12, and 36 months) were sacrificed and all femurs were extracted. The longitudinal and transverse samples of cortical bone were harvested from three regions of diaphysis (proximal, central, and distal). A nanoindentation technique was used for simultaneously measuring force and displacement of a diamond tip pressed 2000nm into the hydrated bone tissue. An elastic modulus was calculated from the unloading curve with an assumed Poisson ratio of 0.3, while hardness was defined as the maximal force divided by the corresponding contact area. The mechanical properties of cortical bone were determined from 852 indents on two orthogonal cross-sectional surfaces. Mean elastic modulus ranged from 7.56±0.32 GPa up to 21.56±2.35 GPa, while mean hardness ranged from 0.28±0.057 GPa up to 0.84±0.072 GPa. Mechanical properties of canine femoral cortical bone tended to increase with age, but the magnitudes of these increase for each region might be different. The longitudinal mechanical properties were significantly higher than that of transverse direction (P<0.01). A significant anisotropy was found in the mechanical properties while there was no significant correlation between the two orthogonal directions in each age group (r 2<0.3). Beyond that, the longitudinal mechanical properties of the distal region in each age group were lower than the proximal and central regions. Hence, mechanical properties in nanostructure of bone tissue must differ mainly among age, sample direction, anatomical sites, and individuals. These results may help a number of researchers develop more accurate constitutive micromechanics models of bone tissue in future studies.

Structural description of surfaces and interfaces in biominerals by DNP SENS.

Biological mineralized tissues are hybrid materials with complex hierarchical architecture composed of biominerals often embedded in an organic matrix. The atomic-scale comprehension of surfaces and organo-mineral interfaces of these biominerals is of paramount importance to understand the ultrastructure, the formation mechanisms as well as the biological functions of the related biomineralized tissue. In this communication we demonstrate the capability of DNP SENS to reveal the fine atomic structure of biominerals, and more specifically their surfaces and interfaces. For this purpose, we studied two key examples belonging to the most significant biominerals family in nature: apatite in bone and aragonite in nacreous shell. As a result, we demonstrate that DNP SENS is a powerful approach for the study of intact biomineralized tissues. Signal enhancement factors are found to be up to 40 and 100, for the organic and the inorganic fractions, respectively, as soon as impregnation time with the radical solution is long enough (between 12 and 24 h) to allow an efficient radical penetration into the calcified tissues. Moreover, ions located at the biomineral surface are readily detected and identified through 31P or 13C HETCOR DNP SENS experiments. Noticeably, we show that protonated anions are preponderant at the biomineral surfaces in the form of HPO42- for bone apatite and HCO32- for nacreous aragonite. Finally, we demonstrate that organo-mineral interactions can be probed at the atomic level with high sensitivity. In particular, reliable 13C-{31P} REDOR experiments are achieved in a few hours, leading to the determination of distances, molar proportion and binding mode of citrate bonded to bone mineral in native compact bone. According to our results, only 80% of the total amount of citrate in bone is directly interacting with bone apatite through two out of three carboxylic groups.

Anisotropic aspects of solubility behavior in the demineralization of cortical bone revealed by XRD analysis.

Dissolution of cortical bone mineral under demineralization in 0.1 M HCl and 0.1 M EDTA solutions is studied by X-ray diffraction (XRD). The bone specimens (in the form of planar oriented pieces) were cut from a diaphysial fragment of a mature mammal bone so that a cross-section surface and a longitudinal section surface could be analyzed individually. This permitted to compare the dissolution behavior of bone apatite of different morphologies: crystals having the c-axis of the hexagonal unit-cell generally parallel to the long axis of the bone (major morphology) and those having the c-axis almost perpendicular to the bone axis (minor morphology). For these two types of morphology, the crystallite sizes in two mutually perpendicular directions (namely, [002] and [310]) were estimated by Scherrer formula in the initial and the stepwise-demineralized specimens. The data obtained reveal that the crystals belonging to the minor morphology dissolve faster than the crystals of the major morphological type, despite the fact that the crystallites of the minor morphology seem to be only a little smaller than those of the major morphology; the apatite crystallites irrespective of the morphology type are elongated in the c-axis direction. We hypothesize that the revealed difference in solubility may be caused by diverse chemical modifications of apatite of these two morphological types, since the solubility of apatite is strictly regulated by anionic and cationic substitutions in the lattice. The anisotropy effect in solubility of bone mineral seems to be functionally predetermined and this should be a crucial factor in the resorption and remodeling behavior of a bone. Some challenges arising at XRD examination of partially decalcified cortical bone blocks are discussed, as well as the limitations of estimation of bone crystallite size by XRD line-broadening analysis.

The Effect of Formalin Preservation Time and Temperature on the Material Properties of Bovine Femoral Cortical Bone Tissue.

Literature has reported controversial findings on whether formalin affected bone properties, or not, especially when different preservation time durations and temperatures were involved. Hence, accurately and systematically quantifying the effect of formalin on the mechanical properties of bone using a large dataset is crucial for assessing biomechanical responses based on fixed specimens. A total of 154 longitudinal and 149 transverse cuboid-shaped (12 mm × 2 mm × 0.5 mm) specimens from the midsection of 12 bovine femora from six bovines were prepared and assigned to ten groups, including fresh-frozen, formalin-preserved at 25 °C for 4 weeks and 8 weeks, and formalin-preserved at 4 °C for 4 weeks and 8 weeks. All specimens underwent quasi-static three-point bending tests with a loading rate of 0.02 mm/s. The Young's modulus, yield stress, yield strain, tangent modulus, effective plastic strain, ultimate stress, and toughness were calculated by optimizing the material parameters to make the force-displacement curve of the finite element prediction consistent with the experimental curve, combined with specimen-specific finite element models. Preservation time and temperature both had significant effects on the Young's modulus, yield stress, effective plastic strain, yield strain and ultimate stress of cortical bone (p < 0.05). The Young's modulus, yield stress, and ultimate stress of longitudinal specimens decreased significantly with the increase of preservation time, and the yield strain increased significantly. As the preservation temperature increases, the Young's modulus of the transverse sample decreased significantly, and the yield strain increased significantly. The preservation time mainly affects the longitudinal specimens, while the preservation temperature mainly affects the transverse specimens. Formalin preservation of bovine femoral cortical bones at a lower temperature and less than 4 weeks is recommended for biomechanical testing.

Supportive Role of Probiotic Strains in Protecting Rats from Ovariectomy-Induced Cortical Bone Loss.

Osteoporosis is a major health problem that occurs as a result of an imbalance between bone formation and bone resorption. Different approaches have been established for treating osteoporosis. Recently, because of their health benefits and also low adverse reaction, probiotics have been receiving considerable attention. In this study, we compared the effectiveness of five probiotic strains, Lactobacillus acidophilus, Lactobacillus reuteri, Lactobacillus casei, Bifidobacterium longum, and Bacillus coagulans, in protecting rats from ovariectomized (OVX)-induced bone loss. Forty-nine adult female Sprague-Dawley rats were allocated into seven groups as follows: group 1, control; group 2, OVX; group 3, OVX + Lactobacillus acidophilus; group 4, OVX + Lactobacillus casei; group 5, OVX + Bacillus coagulans; group 6, OVX + Bifidobacterium longum; and group 7, OVX + Lactobacillus reuteri. Probiotics were fed to OVX groups at the concentration of (1 × 109 CFU/ml/day) for 4 weeks. Then, biochemical parameters, including vitamin D, calcium (Ca), phosphorus (P), and alkaline phosphatase (ALP), were assessed. Dual-energy X-ray absorptiometry (DEXA) scans were used that assess bone mineral density (BMD), bone marrow concentration (BMC), and area of global, femur, spine, and tibia. Lactobacillus acidophilus and Lactobacillus casei significantly increased Ca and ALP and decreased P in treated groups. Lactobacillus casei, Lactobacillus reuteri, and Bifidobacterium longum increased vitamin D significantly. Lactobacillus acidophilus and Lactobacillus casei indicated the most effects on BMD. In terms of BMC, and bone area, Lactobacillus acidophilus, Lactobacillus reuteri, and Lactobacillus casei demonstrated the significant enhancement in OVX groups treated with. Among the probiotics used in this study, Lactobacillus acidophilus and Lactobacillus casei showed the most effects in terms of BMD, BMC, bone area, and biochemical parameters. It seems that probiotics effects on bone health are strain dependent, but further studies should be done to prove these findings.

Acid bone lysate activates TGFß signalling in human oral fibroblasts.

Demineralized bone matrix is a widely used allograft from which not only the inorganic mineral but also embedded growth factors are removed by hydrochloric acid (HCl). The cellular response to the growth factors released during the preparation of demineralized bone matrix, however, has not been studied. Here we investigated the in vitro impact of acid bone lysate (ABL) prepared from porcine cortical bone chips on oral fibroblasts. Proteomic analysis of ABL revealed a large spectrum of bone-derived proteins including TGF-ß1. Whole genome microarrays and RT-PCR together with the pharmacologic blocking of TGF-ß receptor type I kinase with SB431542 showed that ABL activates the TGF-ß target genes interleukin 11, proteoglycan 4, and NADPH oxidase 4. Interleukin 11 expression was confirmed at the protein level by ELISA. Immunofluorescence and Western blot showed the nuclear localization of Smad2/3 and increased phosphorylation of Smad3 with ABL, respectively. This effect was independent of whether ABL was prepared from mandible, calvaria or tibia. These results demonstrate that TGF-ß is a major growth factor that is removed upon the preparation of demineralized bone matrix.