Ultimate Breaking Force Research Articles

Autophagy Is Possibly Involved in Osteoblast Responses to Mechanical Loadings.

Both mechanical loading and autophagy play important roles in regulating bone growth and remodeling, but the relationship between the two remains unclear. In this study, we examined bone structure with micro-CT imaging and measured bone mechanical properties with three-point bending experiments using bones from wild-type (WT) mice and conditional knockout (cKO) mice with Atg7 deletion in their osteoblasts. We found that the knockout mice had significantly less bone volume, bone thickness, bone ultimate breaking force, and bone stiffness compared to wild-type mice. Additionally, bone marrow cells from knockout mice had reduced differentiation and mineralization capacities in terms of alkaline phosphatase and calcium secretion, as well as Runx2 and osteopontin expression. Knockout mice also had significantly less relative bone formation rate due to mechanical loading. Furthermore, we found that the osteoblasts from wild-type mice had stronger responses to mechanical stimulation compared to autophagy-deficient osteoblasts from knockout mice. When inhibiting autophagy with 3 MA in wild-type osteoblasts, we found similar results as we did in autophagy-deficient osteoblasts. We also found that mechanical loading-induced ATP release is able to regulate ERK1/2, Runx2, alkaline phosphatase, and osteopontin activities. These results suggest that the ATP pathway may play an important role in the possible involvement of autophagy in osteoblast mechanobiology.

Effect of Oblique Tendon Laceration on Core Suture Strength: A Biomechanical Evaluation.

The effect of obliquity of tendon laceration on repair strength is not well studied. The overwhelming majority of biomechanical studies assess repair strength following a laceration that is perpendicular to the long axis of the tendon. The aim of this study was to investigate whether the angle of tendon laceration affects the core suture strength. In all, 110 fresh human cadaveric flexor tendons were cut at varying angles of 15°, 30°, 45°, and 60° and the control group at 90°. All tendons were repaired with 6-strand modified Tang technique. The repair strength was tested using a custom-made tensioning machine, and the initial static gap force and the ultimate breaking force were measured. The mean gap force and 95% confidence interval (CI) for the 15°, 30°, 45°, 60°, and 90° groups were 15.2 N (11.4-19.0 N), 15.8 N (13.6-18.1 N), 15.6 N (13.1-18.4 N), 16.6 N (13.7-19.5 N), and 22.3 N (16.6-27.9 N), respectively. In the same respective order, the break force and 95% CI were 25.9 N (21.9-29.8 N), 26.5 N (23.2-29.7 N), 31.1 N (26.1-36.1 N), 35.6 N (28.2-43.1 N), and 51.8 N (62.5-41.0), respectively. The Fisher least significant difference demonstrated significant differences between the control group and all experimental groups for both gap force and break force. Obliquity of tendon laceration affects the core suture strength when compared with a transverse cut. Flexor tendons cut at 90° demonstrated a higher overall gap force and breaking strength that were statistically significant when compared with all obliquely cut groups. These findings should be considered when repairing and starting postoperative therapy for obliquely cut tendons.

Erythropoietin stimulates patellar tendon healing in rats

BackroundErythropoietin (EPO), regulating erythropoiesis, is used to provide protective and regenerative activity in non-haematopoietic tissues. There is insufficient knowledge about the role of EPO activity in tendon healing. Therefore, we investigated the effect of EPO treatment on healing in rat patellar tendons. MethodsOne hundred and twenty-six, four-month-old male Sprague–Dawley rats were randomly assigned to three experimental groups: 1, no treatment; 2, treatment with isotonic saline (NaCl) and 3, treatment with EPO. Each group was randomly subdivided into two groups for sacrifice at three (1a, 2a, 3a) or sixweeks (1b, 2b, 3b). Complete incision of the left patellar tendon from the distal patellar pole was performed. We applied body casts for 20days after the incised edges of the patellar tendon were brought together with a surgical technique. Both legs were harvested and specimens from each group underwent histological, biomechanical, and protein mRNA expression analyses. ResultsThere were statistically significant differences in the ultimate breaking force between the EPO group and others at both weeks three and six (p<0.05); significant differences in fibroblast proliferation, capillary vessel formation, and local inflammation were found between groups 1a and 3a, and 2a and 3a (p<0.05). There were statistical differences between 1a, 3a and 2a, 3a for Col III, TGF-β1, and VEGF and between 1b, 3b and 2b, 3b for Col I, Col III, TGF-β1, and VEGF mRNA expressions. ConclusionEPO had an additive effect with surgery on the injured tendon healing process in rats compared to the control groups biomechanically, histopathologically and with tissue protein mRNA expression. Clinical relevanceThis is the first experimental study to analyze the relationship between EPO treatment and the patellar tendon repair process by biomechanical, histopathological, and tendon tissue mRNA expression methodologies.

The Roles of P2Y2 Purinergic Receptors in Osteoblasts and Mechanotransduction

We previously demonstrated, using osteoblastic MC3T3-E1 cells, that P2Y2 purinergic receptors are involved in osteoblast mechanotransduction. In this study, our objective was to further investigate, using a knockout mouse model, the roles of P2Y2 receptors in bone mechanobiology. We first examined bone structure with micro-CT and measured bone mechanical properties with three point bending experiments in both wild type mice and P2Y2 knockout mice. We found that bones from P2Y2 knockout mice have significantly decreased bone volume, bone thickness, bone stiffness and bone ultimate breaking force at 17 week old age. In order to elucidate the mechanisms by which P2Y2 receptors contribute to bone biology, we examined differentiation and mineralization of bone marrow cells from wild type and P2Y2 knockout mice. We found that P2Y2 receptor deficiency reduces the differentiation and mineralization of bone marrow cells. Next, we compared the response of primary osteoblasts, from both wild type and P2Y2 knockout mice, to ATP and mechanical stimulation (oscillatory fluid flow), and found that osteoblasts from wild type mice have a stronger response, in terms of ERK1/2 phosphorylation, to both ATP and fluid flow, relative to P2Y2 knockout mice. However, we did not detect any difference in ATP release in response to fluid flow between wild type and P2Y2 knock out osteoblasts. Our findings suggest that P2Y2 receptors play important roles in bone marrow cell differentiation and mineralization as well as in bone cell mechanotransduction, leading to an osteopenic phenotype in P2Y2 knockout mice.

Enzyme‐Free Quinone Crosslinking Reaction for Proteins: A Macromolecular Characterization Study Using Gelatin

To mimic the quinone hardening of extracellular proteins in invertebrates, we investigated an enzyme-free crosslinking of gelatin by HQ in a neutral aqueous phase. The mixture was rapidly transformed to a yellowish brown, thermally and mechanically stable hydrogel in the presence of a simple copper(II) salt. A dehydrated thin film made of the mixture was flexible, tough, and showed a large ultimate breaking force. Physicochemical examination of the gel suggested that the basic amino acid residues (lysine, hydroxylysine, and histidine) of the protein were modified by the quinone ring to form 2-6 crosslinks per protein. The enzyme-free crosslinking reaction is discussed with consideration of a copper(II) ion-catalyzed oxidation of HQ and the hydroquinone/protein adducts.

Abnormal Response to Physical Activity in Femurs after Heterozygous Inactivation of One Allele of the Col2a1 Gene for Type II Collagen in Mice

The objective of this study was to evaluate the influence of heterozygous inactivation of one allele of the type II collagen gene (Col2a1) on biomechanical properties and mineral density of bone under physical loading conditions. C57BL/6-TGN mice with heterozygous knockout (HZK) inactivation of Col2a1 gene and their nontransgenic littermate controls were housed in individual cages with running wheels for 9 and 15 months. The running activity of each mouse was monitored continuously throughout the experiment. Bone mineral density (BMD) of mice femora was measured using dual-energy X-ray absorptiometry (DXA) and peripheral quantitative computerized tomography (pQCT). Biomechanical properties were determined using three-point bending tests. Vertebral bone samples were prepared for quantitative polarized light microscopy and digital densitometry of proteoglycans. The concentration of total collagen and collagen cross-links were analyzed using high-performance liquid chromatograpy (HPLC). The average daily running distance was shorter for the HZK mice between the age of 4 and 15 months as compared with normal runners (P < 0.05). The ultimate breaking force was 14.8% and 23.6% (9 vs. 15 months) lower in HZK-runners than in wild-type runners. BMD of the femur was 6.1% lower in HZK-runners at the age of 9 months (P < 0.05). Physical activity increased cortical BMD in wild-type runners but not in the HZK runners at the age of 9 months. The collagen network of the HZK mice was less organized. There were only minor changes in BMD and mechanical and structural properties between sedentary HZK mice and their wild-type controls. Increased physical activity induced significantly lower bone density, mechanical properties, and organization of collagen fibers in male HZK mice. However, there were no major differences in biomechanical parameters between sedentary HZK and wild-type male mice. This suggests an important guiding role of collagen type II in bone remodelling and maturation.

Adaptations to free-fall impact are different in the shafts and bone ends of rat forelimbs.

Impact exercise can have beneficial effects on the growing skeleton. To understand what changes it promotes in the shafts and ends of weight-bearing bones, we measured the effects of impact from repetitive free falls in growing rats. Fischer 344 female rats, 6.5 wk old, were assigned to one of three groups (n = 10 each). Controls were not dropped, whereas those subjected to impact were dropped from 30 or 60 cm. Rats in both free-fall groups were dropped 10 times per day for 8 wk. Leg bones were mechanically tested, and their cross-sectional area (CSA), cross-sectional moments of inertia, and volumetric bone mineral density (BMD) were measured by peripheral quantitative computed tomography. In the shafts of the forelimbs, but not the hindlimbs, free-fall impact resulted in greater ultimate breaking force, minimum and maximum second moments of area, and CSA but not BMD. In the bone ends of the forelimb and tibial bones, trabecular BMD increased but CSA did not. Landing from 30 and 60 cm produced peak impact forces of 12.0 and 16.7 times the standing forefoot weight for each front leg and of 4.5 and 7.7 times the standing hind foot weight for each hind foot. Overall, free-fall impact affected the forelimbs by increasing trabecular bone density in the bone ends and improving the strength at the shaft as a result of geometric improvements. These results indicate that adaptation to impact may occur by different mechanisms in bone end and shaft regions.

The effects of replacing dicalcium phosphate with Busumbu rock phosphate on performance and the mechanical properties of bone in growing chicks.

Three hundred, day-old broiler chicks, with an average initial weight of 41.8 +/- 1.79 g, were used in a 15-day study (10 birds per battery cage) to characterize their performance and fluorine status when dicalcium phosphate (DCP) was replaced by Busumbu rock phosphate (BRP) as the source of phosphorus in the chicks' ration. The treatments comprised a standard ration with BRP replacing 0, 25%, 50%, 75% or 100% of DCP. Replacing DCP with BRP significantly reduced the final weight of the chicks (p < 0.01), feed intake (p < 0.01), weight gains (p < 0.01) and dry matter digestibility (p < 0.05) but increased (p < 0.05) the feed-to-gain ratio. True phosphorus absorption and the percentage of phosphorus in the tibia were not affected by increasing amounts of BRP in the diet. Increasing levels of BRP in the diet linearly reduced (p < 0.01) the percentage bone ash, calcium, Ca:P ratio, ultimate breaking force, bending moment, stress, and modulus of elasticity. Leg stiffness, lameness, reduced feed intake, and a decline in general health were recorded in 10-40% of the chicks on 75% and 100% BRP, respectively. These results suggest that excessive ingestion of fluorine from the BRP caused the reduction in the chicks' performance.

Effect of high-impact and low-repetition training on bones in ovariectomized rats.

This study was designed to investigate the effect of high-impact and low-repetition jump training on bones in ovariectomized (OVX) rats. Forty female Wistar rats were sham-operated (sham) or OVX at the age of 11 weeks. The rats were divided randomly into the following four groups: sham-sedentary (SS; n = 10), sham-exercised (SE; n = 10), OVX-sedentary (OS; n = 10), and OVX-exercised (OE; n = 10). The rats started the jump training at the age of 12 weeks. The jump-training protocol was 10 times/day, 5 days/week and the jumping-height was 40 cm. After 8 weeks of training, the mass and breaking force in the tibia and ulna, cross-sectional areas of diaphysis in the tibia, and serum bone turnover markers were measured. The jump training significantly increased the fat-free dry weight, ash weight, and ultimate breaking force in the tibia. The rate of increase in these parameters was similar in both the sham and the OVX groups. On the other hand, in the ulna, there were no significant changes in the ultimate breaking force. The jump training significantly increased the periosteal perimeter and cortical area, although the increase in these parameters in OE compared with OS was lower than that in SE compared with SS. The jump training significantly increased serum osteocalcin in the OVX groups, as well as in the sham groups. These results suggest that high-impact and low-repetition training had beneficial effects on bone formation and bone biomechanical properties in OVX rats, as well as in sham rats.