Assessment of Bone Quality Associated with Loosely and Tightly Bound Water, 15-R8061

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Principal Investigators
Qingwen Ni
Armando De Los Santos
Daniel Nicolella

Inclusive Dates:  04/15/09 – 08/15/09

Background - Previous studies have shown that the age-related increase in bone porosity results in a decrease in bone strength, and porosity is related to the volume of mobile water in the pores. In addition, because water is also bound to collagen and mineral, changes in the amount of bound water also will potentially affect bone strength. It is known that the removal of the loosely bound water (via hydrogen bonding) requires less energy than the water molecules trapped inside collagen molecules, which, in turn, requires similar or less energy than water molecules bound to the surface charges of mineral apatite (more ionic in nature). Also, water that is imbedded in the lattice of hydroxyapatite (more covalent in nature) requires the highest energy to dislodge. However, there is no traditional method that can determine mobile and bound water accurately, nondestructively and noninvasively. SwRI researchers propose using NMR Car-Purcell-Meiboom-Gill (CPMG) spin-spin relaxation measurement to determine the mobile water and the NMR inversion T2-FID spectrum derived from NMR free induction decay (FID) measurements for estimating the bound and free water distribution. Furthermore, after comparison of the total water lost (weighing method) within tissue by using drying (free dry) on the air to the total mobile water lost measured by the NMR CPMG method, the total loosely bound water lost can be estimated. Following this, a mechanical test will be used to evaluate the bone quality related to the tightly and loosely bound water within bone. This information can be used to further assess bone quality.

Approach - The project had the following objectives:

  1. Use NMR to determine bone porosity and water (bound and mobile) distribution using proton NMR CPMG and FID spin-spin (T2) relaxation signal measurement techniques;
  2. Determine the changes of loosely and tightly bound water using NMR CPMG and weighing methods;
  3. Monotonically load the sample to failure to determine the mechanical properties under displacement control using a closed-loop servo hydraulic test machine;
  4. Determine the bone strain correlation with the bound (loosely and tightly) water.

Accomplishments - Ten female baboon cortical bone samples, five from a younger group (6 to 8-year-old) and five from an older group (26 to 29-year-old) were collected and used in this project. One year of age in a baboon is approximately equivalent to three years of age in humans. The sample size was about 8 mm by 3mm by 40 mm. An SwRI-built 0.5 to 40 MHz broadline NMR system was set up at a proton frequency of 27 MHz for these measurements. 1H spin-spin (T2) relaxation profiles were obtained by using NMR CPMG {90° [- t - 180° - t(echo)]n - TR} spin echo method with a 9.5 µs wide RF-90° pulse, t of 1000 µs, and TR (sequences repetition rate) of 15 s. Each T2 profile, which included one thousand echoes (one scan with n = 1000), was acquired and sixth-four scans were used for the bone porosity measurements. The FID signal was sampled and recorded at 2 µs intervals using a 9.5 µs wide 90° RF-pulse for the bone mobile water and bound water measurements. An inversion relaxation technique was used to invert both CPMG and FID data to T2 relaxation distribution spectrum.

This is the first time that the porosity, water distributions, mobile and bound water, and specifically the loosely bound water has been measure noninvasively and nondestructively in baboon cortical bone. The porosity distribution pattern differences between baboon and human cortical bone is observed. In addition, the ratio of bound water to mobile water in baboon bone is much higher than in human bone. All these can be related to an overall lower porosity in baboon bone compared to human bone. There was no evidence of age-related differences in either porosity or the ratio of bound to mobile water between young and old baboon bone. However, there was a significant difference in loosely bound water between the young and old bone, and it appears that this loosely bound water may play an important function in bone mechanical properties, particularly in the yield and post-yield behavior of bone tissue. In a previous SwRI study, it was found that bound water plays an important function in human bone mechanical properties. Because the average of bound water to mobile water is much higher than in human bone, it is reasonable to assume that the loosely bound water plays an important function in the mechanical properties. It is concluded that the ratio of bound water to mobile water is not enough to fully assess bone quality and thus bone mechanical integrity, and a more detailed characterization of the bound water (such as the fraction of loosely bound or tightly bound water) is needed to assess the mechanical integrity of bone tissue.

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