This study has been approved by the Institutional Review Board (IRB) of the Third Hospital of Hebei Medical University.
Specimen Preparation: Fourteen fresh-frozen cadaveric lower limbs with intact soft tissue were autopsied (all cadavers were provided by the Department of Human Anatomy, Hebei Medical University), average height of the donors was 171 cm (range, 163 to 181 cm), average age was 55 years (range, 42 to 65 years). Each had complete femur, tibia and knee joint, there were no gross deformities of the knee, i.e. hyperflexion, hyperextension, varus and valgus. Passive joint motion are freely. Furthermore, inner knee structures were examined by X ray, pathological (osteoporosis, rheumatism, tuberculosis, or tumors) or anatomical deformities (unsymmetrical joint surface, bone hyperplasia, or other imaging abnormalities) were excluded(Fig. 1).
After all muscular tissues were removed, anatomical axes along the length of the intramedullary canals of the femur and tibial were marked based on the method proposed by Moreland. In order to carry out this biomechanical experiment, we reserved approximately 25 cm of the distal femur, proximal tibia and fibula, wrapped dissected cadaveric knees with polyethylene films to prevent dehydration and cryopreserved at − 20℃.
Establishment of rotatory fixation model: The cadaveric knees were thawed at room temperature for 12 hours before experiment. Cut a horizontal incision about 3–4 cm long at the level of joint space, both sides of the patellar ligament. Separate the subcutaneous fat, cut the sac, and expose the joint space, attention must be paid to reserve anterior and posterior cruciate ligaments, as meniscus is a weight-bearing structure that can buffer pressure and affect the expansion. Then saw the femoral shaft transversely at distal 1/3, guarante that each cut is basically at the same level to eliminate heterogeneity. Take the previously drawn anatomical axes as the measurement benchmark, and predetermined angle was measured with a bone protractor, finally fixed two stumps with plates and screws. In our study, we chose neutral position (0°, anatomically reduced), the internal rotation 5°, 10°, 15° and the external rotation 5°, 10°, 15° as experimental factors.
Pressure-sensitive film inserted
An ultra-low-pressure sensitive film (LLW type, Fujifilm Investment Co. Ltd. Japan) (0.5–2.5 MPa) is used to reflect the contact pressure on tibial plateau, in order to ensure the quality of the pressure-sensitive film, we set the room humidity to 35%RH and the temperature to 20°C. Trim the pressure-sensitive film into somehow match shape according to our preliminary experiment, then seal it with a polyethylene film bag, a total thickness must be less than 250µm, thereafter carefully insert it under the meniscus and fully accessed into the joint cavity, suture the capsule tightly, leakage, bending, breakage of the sealed bag means failure (Fig. 2). In order to distinguish the anterior and posterior side, the corresponding anterior side of the pressure sensitive sheet is clamped with a hemostatic forceps in advance to make an impression.
Clamp the femur and tibial end perpendicularly and reinforce with the denture base resin and solution (type II self-setting dental powder and tray water) (Fig. 3–4). Then transfer and assemble the combination to the biomechanical testing machine (Electroforce 3520-AT, Bose company, USA). As the measurement work will be done dozens of times, so we are intended to ensure conformity between each step.
To simulates a normal male adult in naturally standing state ,we chose 400N load. Start the biomechanical machine, load the test bench, pressurize to 200N at a speed of 10N/s to eliminate creep. After stabilizing, apply a vertical load to the specimen to 400N at a speed of 10N/s and uphold for 2 minutes, unload and get the pressure-sensitive film out.
Pressure value read
FPD-305E density meter and FPD-306E pressure converter were used to read relative pressure value. We divided the contact pressure area (the red area) of each pressure-sensitive film into 4 quadrants (anterolateral, anterior medial, posterior medial and posterior lateral), each quadrant randomly and equally read 5 values, total 20 values in one film, take the average as final values.
The experimental data were organized and computed by SPSS 21.0 software (SPSS, Chicago, IL, USA). The normality is verified using the Shapiro–Wilk test and expressed as x̄±s, we used T-test of two independent samples to access difference between medial and lateral groups, the Student–Newman–Keuls test for pairwise comparisons between the multiple sample measurements. Using the Levene test for variance consistency, and analysis of variance (ANOVA) for random block groups. Data doesn’t fit normality expressed as the median (quartile) and using Mann-Whitney U test to access difference between medial and lateral groups. Kruskal-Wallis H test for random block groups, significance was P < 0.05.