Selected mechanical properties of human cancellous bone subjected to different treatments: short-term immersion in physiological saline and acetone treatment with subsequent immersion in physiological saline

Abstract

Background: Physiological saline (0.9% NaCl) and acetone are extensively used for storage (as well as hydration) and removal of bone marrow, respectively, of cancellous bone during preparation and mechanical testing. Our study aimed to investigate the mechanical properties of cancellous bone subjected to short-term immersion in saline and acetone treatment with subsequent immersion in saline.

Methods: Cylindrical samples (Ø6 × 12 mm) were harvested from three positions (left, middle, and right) of 1 thoracic vertebral body, 19 lumbar vertebral bodies, and 5 sacral bones, as well as from 9 femoral heads. All samples were divided into two groups according to the different treatments, (i) samples from the left and middle sides were immersed in saline at 4℃ for 43 h (saline-immersed group, n = 48); (ii) samples from the respective right side were treated with a combination of acetone and ultrasonic bath (4 h), air-dried at room temperature (21℃, 15 h), and then immersed in saline at room temperature (21℃, 24 h) (acetone and saline-treated group, n = 38). All samples were subjected, both before and after treatment, to a non-destructive compression test with a strain of 0.45%, and finally destructive tests with a strain of 50%. Actual density (ρ_act), initial modulus (E₀), maximum stress (σ_max), energy absorption (W), and plateau stress (σ_p) were calculated as evaluation indicators.

Results: Based on visual observation, a combination of acetone and ultrasonic bath for 4 h failed to completely remove bone marrow from cancellous bone samples. The mean values of ρ_act, σ_max, W, and σ_p were significantly higher in the femoral head than in the spine. There was no significant difference in E₀ between non-treated and saline-immersed samples (non-treated 63.98 ± 20.23 vs. saline-immersed 66.29 ± 20.61, p = 0.132). The average E₀ of acetone and saline-treated samples was significantly higher than that of non-treated ones (non-treated 62.17 ± 21.08 vs. acetone and saline-treated 74.97 ± 23.98, p = 0.043).

Conclusion: Short-term storage in physiological saline is an appropriate choice and has no effect on the E₀ of cancellous bone. Treatment of cancellous bone with acetone resulted in changes in mechanical properties that could not be reversed by subsequent immersion in physiological saline.

Keywords: Acetone; Bone marrow; Cancellous bone; Mechanical properties; Physiological saline.

Fig. 1

The flow chart of the main steps in our experiment. ¹Human donors (n = 34) included 1 thoracic vertebral body (T₁₂), 19 lumbar vertebral bodies (L_1–5), 5 sacral bones (S_1–5), and 9 femoral heads. ²Cylindrical samples (Ø6 × 12 mm, n = 86) were obtained using a tenon cutter and a stationary drilling machine, including 63 samples obtained from spinal vertebral bodies and 23 samples collected from femoral heads. ³Saline-immersed group: the samples from the left and middle sides of spinal vertebral bodies and femoral heads. ²Acetone and saline-treated group: the samples from the respective right side of spinal vertebral bodies and femoral heads

Fig. 2

Sample preparation: A1 shows the location of the femoral head samples within the available region. B1 The main trabecular orientation was visible after cutting. C1 The femoral head was fixed in the clamping jaws and then drilled in the superior-inferior direction along with the main trabecular orientation. A2–C2 Cylindrical cores were harvested from three positions (left, middle, and right) of 1 thoracic vertebral body (T₁₂), 19 lumbar vertebral bodies (L_1–5), and 5 sacral bones (S_1–5) along the long and midline axis. D, E The cores were cut into samples (Ø6 × 12 mm). F The samples were tested after being glued to the platens with cyanoacrylate

Fig. 3

Typical non-destructive and destructive stress–strain curves of a cancellous sample plotted with the evaluation of deformed shape. E₀: initial modulus (0–0.2% strain, MPa), E_max: maximum modulus (0–0.2% strain, MPa), σ_max: maximum stress (MPa), σ_p: plateau stress (20%-40% strain, MPa)

Fig. 4

The appearance of non-treated, as well as acetone and saline-treated samples. A1–A2 the low-density sacral sample before and after treatment by a combination of acetone and ultrasonic bath (4 h); B1–B2/C1–C2: the relatively dense femoral head samples before and after treatment by a combination of acetone and ultrasonic bath (4 h); C3–C4: one of the samples (n = 3) was split longitudinally and vertically through the center. The samples treated with a combination of ultrasonic bath and acetone (4 h) still had a certain amount of bone marrow left

Fig. 5

The distribution of actual density (ρ_act, g/cm³) and initial modulus (E₀, MPa) in the spine (thoracic, lumbar vertebral bodies, and sacral bones) and femoral heads (*p < 0.05)

Fig. 6

The initial modulus (E₀, MPa) in different groups before and after treatment: A saline-immersed group, B acetone and saline-treated group (*p < 0.05)

Fig. 7

From destructive testing, femoral head has higher mechanical properties than spine in several parameters, including maximum stress (σ_max, MPa), plateau stress (σ_p, MPa), and energy absorption (W, mJ/mm³)