Determination of the minimal necessary resolution for valid evaluation of trabecular architecture in small animal models

Introduction MicroCT analysis of bone has become a standard methodology in bone research. One main factor for the quality of the microCT-data is its spatial resolution. Since high-resolution scanning requires long acquisitions times, as well as huge data processing, reaching a compromise for the dichotomy between ideally small voxel size and analysis time is necessary. Here we aimed to analyze the minimum necessary resolution that still generates reliable data for the different architectural features of trabecular bone in a rat animal model.

Methods The 3 rd lumbar vertebral body of 25-week-old female Sprague-Dawley rats (Harlan Winkelmann, Borchen, Germany; 12 weeks; n = 3) were scanned using a high-definition micro-CT system (µCT 50, SCANCO Medical AG, Switzerland; source voltage = 90 kVp, intensity=88 µA). As the trabecular thickness of rats is typically reported at 60 µm, we chose five different voxel sizes: 4.4, 7.4, 10.0, 14.8, and 34.4 µm resulting in object size to voxel size ratios between 13 and 2.43. The volume of interest was 7.5 mm3 in the mid-section of the vertebral body, excluding the cortical bone. Bone morphometric parameters such as bone volume fraction (BV/TV), trabecular thickness (Tb.Th.), trabecular number (Tb.N.), bone mineral density (BMD), and connectivity density (Conn.D.) were then measured directly in 3 D models. The mean values at 4.4 µm were deemed as reference points. The necessary resolution was defined as the lowest scan resolution (i. e. highest voxel size) where the differences of the morphometric parameters were not statistically different compared to the reference data at maximum resolution. Statistical evaluation was carried out using SPSS package (IBM SPSS, version 25.0). The level of significance was designated as follows: no significant difference p > 0.05 (NS), p < 0.05 (*), p < 0.01 (**), and p < 0.001 (***).

Results By increasing the voxel size, we found obvious trends for all morphometric parameters; Namely, BV/TV, Tb.N., and Conn.D. reduced, Tb.Th. increased, while BMD showed erratic fluctuations. Up to 14.8 µm, the differences compared to the reference point (4.4 µm) were insignificant for all parameters, except for Conn.D., which already showed a significant decline at 14.8 µm (*). At 34.4 µm, however, BV/TV, Tb.N., Conn.D., and BMD were significantly 74.0 % (**), 64.7 % (***), 95.3 % (***), and 5 % (*) reduced, respectively. On the other hand, the value of Tb.Th. increased by 39.2 % (*). At 14.8 and 34.4 µm, the ratio of “Tb.Th./voxel size” were 4.0 and 2.4, respectively.

Discussion Our results indicate that massive changes of the data occur, if the resolution is too low. The minimal necessary resolution to examine the trabecular structure of rat bones is at least 10.0 µm for the Conn.D. and 14.8 µm for other main morphometric parameters. To translate the results to other models, we propose a minimum “Tb.Th./voxel size” of 4.

Keywords Micro-computed Tomography, Spatial Resolution, Trabecular Bone, Bone Morphometry

Korrespondenzadresse Shahed Taheri, Universitätsmedizin Göttingen, Clinic for Trauma Surgery, Orthopaedic Surgery and Plastic Surgery, Robert-Koch-Str, 40, 37075 Göttingen, Germany,

E-Mail [e-mail]