Abstract
MicroRNA´s are regulatory sequences which influence the posttranscriptional synthesis of about 70% of protein encoding genes. In different studies, MicroRNA-146a (miR-146a) was associated with inflammatory and autoimmunological processes. In vitro it was shown, that miR-146a influences the bone metabolism by regulating differentiation of mesenchymal stem cells. The miR-146a deficient mouse starts to develop lymphoproliferative and myeloproliferative disease by 6–8 months of age. In this study, we investigate the influence of miR-146a deficiency on bone structure and stability dependent on age and gender.
Material and Methods
Male and female mice of wild type (WT) and miR-146a deficient (KO) animals at the age of 2–3 and 5–7 month were analyzed Femur, Tibia and lumbar vertebra (LWK4) were dissected and used für structural analyses by microcomputer tomography (µCT). Parameters like bone volume/tissue volume, trabecular bone volume, trabecular thickness, number and separation as well as cortical thickness were determined. Biomechanical stability as load to failure testing was determined using torsional testing for the long bones and axial compression testing for the vertebra body. Statistical analysis was performed using Graph Pad Prism (Mann-Whitney-U-Test, significance: p<0.05).
Results
Structural analyses of the bone structure in the long bones (femur, tibia) revealed a significant higher bone volume/tissue volume (BV/TV) and trabecular bone mass in the elder (5–7 month) miR-146a deficient female mice compared to the male group or wild type animals of either age. In the diaphysis of the femur a BV/TV of 21% was determined for the elder miR-146a deficient females compared to 9% BV/TV in the age matching WT group. These changes were due to an increase in trabecular thickness and trabecular number in this area. In contrast to that, the cortical thickness of all bones analyzed was lowered in the miR-146a deficient animals (male and female) compared to wild type. Biomechanical stability of long bones as well as the vertebra body of the older, female KO group was significantly lower compared to wild type bones. Femurs showed a maximal torque of 20Nmm compared to 34Nmm in the wild type group. The vertebra of the KO mice showed a maximal force at failure of 22N compared to 40N in the wild type group. Male groups and younger females revealed values comparable to wild type animals.
Conclusion
The deficiency of miR-146a leads to an increase of trabecular bone in the long bones of female 5–7 month old mice, but to lowered biomechanical bone stability. If this is due to alterations in differentiation or proliferation of mesenchymal stem cells remains unclear and will be analyzed further. Additionally, gender relation of our observations points to the influence of female specific regulatory mechanisms like the involvement of estrogen receptor related mechanisms
