The impact of low-magnitude high-frequency vibration on fracture healing is profoundly influenced by the oestrogen status in mice

Fracture healing is impaired in aged and osteoporotic individuals. Because adequate mechanical stimuli are able to increase bone formation, one therapeutical approach to treat poorly healing fractures could be the application of whole-body vibration, including low-magnitude high-frequency vibration (LMHFV). We investigated the effects of LMHFV on fracture healing in aged osteoporotic mice. Female C57BL/6NCrl mice (n=96) were either ovariectomised (OVX) or sham operated (non-OVX) at age 41 weeks. When aged to 49 weeks, all mice received a femur osteotomy that was stabilised using an external fixator. The mice received whole-body vibrations (20 minutes/day) with 0.3 g peak-to-peak acceleration and a frequency of 45 Hz. After 10 and 21 days, the osteotomised femurs and intact bones (contra-lateral femurs, lumbar spine) were evaluated using bending-testing, micro-computed tomography (μCT), histology and gene expression analyses. LMHFV disturbed fracture healing in aged non-OVX mice, with significantly reduced flexural rigidity (−81%) and bone formation (−80%) in the callus. Gene expression analyses demonstrated increased oestrogen receptor β (ERβ, encoded by Esr2) and Sost expression in the callus of the vibrated animals, but decreased β-catenin, suggesting that ERβ might mediate these negative effects through inhibition of osteoanabolic Wnt/β-catenin signalling. In contrast, in OVX mice, LMHFV significantly improved callus properties, with increased flexural rigidity (+1398%) and bone formation (+637%), which could be abolished by subcutaneous oestrogen application (0.025 mg oestrogen administered in a 90-day-release pellet). On a molecular level, we found an upregulation of ERα in the callus of the vibrated OVX mice, whereas ERβ was unaffected, indicating that ERα might mediate the osteoanabolic response. Our results indicate a major role for oestrogen in the mechanostimulation of fracture healing and imply that LMHFV might only be safe and effective in confined target populations.KEY WORDS: Whole-body vibration, LMHFV, Fracture healing, Oestrogen receptor signalling, Wnt signalling     

Conformational Changes in a Hyperthermostable Glycoside Hydrolase: Enzymatic Activity Is a Consequence of the Loop Dynamics and Protonation Balance

Endo-β-1, 4-mannanase from Thermotoga petrophila (TpMan) is a modular hyperthermostable enzyme involved in the degradation of mannan-containing polysaccharides. The degradation of these polysaccharides represents a key step for several industrial applications. Here, as part of a continuing investigation of TpMan, the region corresponding to the GH5 domain (TpManGH5) was characterized as a function of pH and temperature. The results indicated that the enzymatic activity of the TpManGH5 is pH-dependent, with its optimum activity occurring at pH 6. At pH 8, the studies demonstrated that TpManGH5 is a molecule with a nearly spherical tightly packed core displaying negligible flexibility in solution, and with size and shape very similar to crystal structure. However, TpManGH5 experiences an increase in radius of gyration in acidic conditions suggesting expansion of the molecule. Furthermore, at acidic pH values, TpManGH5 showed a less globular shape, probably due to a loop region slightly more expanded and flexible in solution (residues Y88 to A105). In addition, molecular dynamics simulations indicated that conformational changes caused by pH variation did not change the core of the TpManGH5, which means that only the above mentioned loop region presents high degree of fluctuations. The results also suggested that conformational changes of the loop region may facilitate polysaccharide and enzyme interaction. Finally, at pH 6 the results indicated that TpManGH5 is slightly more flexible at 65°C when compared to the same enzyme at 20°C. The biophysical characterization presented here is well correlated with the enzymatic activity and provide new insight into the structural basis for the temperature and pH-dependent activity of the TpManGH5. Also, the data suggest a loop region that provides a starting point for a rational design of biotechnological desired features.     

Correlation between Body Composition and Walking Capacity in Severe Obesity

Background

Obesity is associated with mobility reduction due to mechanical factors and excessive body fat. The six-minute walk test (6MWT) has been used to assess functional capacity in severe obesity.

Objective

To determine the association of BMI, total and segmental body composition with distance walked (6MWD) during the six-minute walk test (6MWT) according to gender and obesity grade.

Setting

University of São Paulo Medical School, Brazil; Public Practice.

Methods

Functional capacity was assessed by 6MWD and body composition (%) by bioelectrical impedance analysis in 90 patients.

Results

The mean 6MWD was 514.9 ± 50.3 m for both genders. The male group (M: 545.2 ± 46.9 m) showed a 6MWD higher (p = 0.002) than the female group (F: 505.6 ± 47.9 m). The morbid obese group (MO: 524.7 ± 44.0 m) also showed a 6MWD higher (p = 0.014) than the super obese group (SO: 494.2 ± 57.0 m). There was a positive relationship between 6MWD and fat free mass (FFM), FFM of upper limps (FFM_UL), trunk (FFM_TR) and lower limbs (FFM_LL). Female group presented a positive relationship between 6MWD and FFM, FFM_UL and FFM_LL and male group presented a positive relationship between 6MWD and FFM_TR. In morbid obese group there was a positive relationship between 6MWD with FFM, FFM_UL, FFM_TR and FFM_LL. The super obese group presented a positive relationship between 6MWD with FFM, FFM_TR and FFM_LL.

Conclusions

Total and segmental FFM is associated with a better walking capacity than BMI.     

Controlling network topology and mechanical properties of co‐assembling peptide hydrogels

Oligopeptides are well‐known to self‐assemble into a wide array of nanostructures including β‐sheet‐rich fibers that when present above a critical concentration become entangled and form self‐supporting hydrogels. The length, quantity, and interactions between fibers influence the mechanical properties of the hydrogel formed and this is typically achieved by varying the peptide concentration, pH, ionic strength, or the addition of a second species or chemical cross‐linking agent. Here, we outline an alternative, facile route to control the mechanical properties of the self‐assembling octa‐peptide, FEFEFKFK (FEKII); simply doping with controlled quantities of its double length peptide, FEFEFKFK‐GG‐FKFKFEFE (FEKII18). The structure and properties of a series of samples were studied here (0–100 M% of FEKII18) using Fourier transform infrared, small angle X‐ray scattering, transmission electron microscopy, and oscillatory rheology. All samples were found to contain elongated, flexible fibers and all mixed samples contained Y‐shaped branch points and parallel fibers which is attributed to the longer peptide self‐assembling within two fibers, thus creating a cross‐link in the network structure. Such behavior was reflected in an increase in the elasticity of the mixed samples with increasing quantity of double peptide. Interestingly the elastic modulus increased up to 30 times the pure FEKII value simply by adding 28 M% of FEKII18. These observations provide an easy, off‐the‐shelf method for an end‐user to control the cross‐linked network structure of the peptide hydrogel, and consequently strength of the hydrogel simply by physically mixing pre‐determined quantities of two similar peptide molecules. © 2013 Wiley Periodicals, Inc. Biopolymers 101: 669–680, 2014.