Control Group Selection in Preclinical Rat Bone Defect Models: A Systematic Review

Large bone defects and loss present major orthopedic challenges. In preclinical research, femoral bone defects in rats are commonly used as in vivo models to evaluate new osteoregenerative biomaterials. These test items are typically compared to negative and positive controls. This review aims to summarize the different control groups used to evaluate new osteoregenerative test items in preclinical rat femoral defect models and to identify potential pitfalls related to these controls, ultimately to enhance the future translational success. The protocol for this review was registered in PROSPERO, and no specific funding was received for this work. The systematic search comprised publications between January 2001 and January 2023. 436 studies were included for analysis. The choice of control groups was inconsistent across studies. A negative (e.g., empty defects or inert carriers) and positive (e.g., bone grafts or commercially available bone substitutes) control group was included in 56% (n = 245/436) and 34% (n = 149/436) of the included studies, respectively. Notably, 25% (n = 109/436) of the studies did not include any control group. Bone grafts were used as positive controls in 50% of the studies that included positive controls (n = 74/149), mainly of allogeneic origin (45%, n = 33/74). The control groups used to evaluate the test item impacted the healing comparison, with 81% of studies showing better healing of their test items compared to negative control (n = 198/245) versus 54% compared to positive control (n = 80/149). A qualitative risk-of-bias and reporting assessment was performed using an integrated ARRIVE-SYRCLE framework. Most studies demonstrated moderate concern in several domains, with frequent absence of randomization (67%, high concern) and blinding (84%, high concern), incomplete reporting of inclusion/exclusion criteria (74%, moderate concern), and variable clarity regarding animal characteristics and statistical methodology. The variability in the choice of control groups appears to influence study outcomes. Inadequate control group selection can lead to misleading conclusions regarding the efficacy of new biomaterials. Therefore, standardizing control group selection is crucial to enhance the reliability and comparability of preclinical research findings.