Type 2 diabetes (T2D) mellitus is associated with an increased fracture risk, independently of bone mineral density (BMD). A reduction in bone quality might explain this discrepancy in T2D. Appropriate exercise results in bone mass maintenance, bone formation and improved bone strength. miRNAs play a role in the pathogenesis of T2DM and diabetic bone disease. Changes in miRNA levels may precede changes in bone turnover markers (BTMs), making them more sensitive markers of bone disease.
This study aims to identify miRNA signatures of exercise training that modulate the expression of specific genes regulating osteoclast and osteoblast function in T2D, thus impacting bone quality.
In step 1, 30 trained and 30 untrained T2D patients, will be selected from patients participating in the SWEET BONE Trial. Patients, previously randomized to either the Exercise group or the Control group, will be evaluated at the end of the 24-month follow-up of the trial. Patients¿ sera will be used in a screening analysis of the entire miRNome performed by Next Generation Sequencing (NGS). Relevant up/down-regulated miRNAs will then be tested and validated by quantitative Real Time-PCR (RT-PCR). The identified circulating c-miRNA will be correlated with measures of bone quality, BTMs, vertebral fractures, cardiorespiratory and muscular fitness, and body composition.
Should the identified c-miRNAs be associated with clinical outcomes, i.e. measures of bone quality and strength (and number of fractures), we would generate the hypothesis that c-miRNAs are mediators of the effects of exercise training on bone quality and fracture risk in T2D. Further, in vitro studies will be necessary to test this hypothesis in controlled, experimental conditions.
The present study will allow the identification of any c-miRNAs that are modulated by exercise training and that may impact bone remodeling in T2D. Considering the lack of studies assessing effects of exercise training in general, and exercise-associated miRNAs in particular, on bone quality in T2D, we cannot generate a specific hypothesis for the anticipated role of these miRNAs on bone strength and quality.
The exploratory phase of the study might lead to different scenarios. We may identify new miRNAs changing in response to exercise training or we might confirm the up/down regulation of previously reported miRNAs. Even more, having a T2D control group that is not participating in the exercise training might allow the evaluation of the impact of exercise training on bone-relevant miRNAs. More miRNAs, such as miR-378, miR-223, miR-20a, miR-103a and miR-214 and miR-17-92 cluster, previously reported to play a role in diabetes, exercise adaptations or osteogenesis, will also be assessed specifically. This will allow to either confirm or contrast previous findings, and determine their role in bone quality.
Once relevant signature miRNAs are identified, we plan a future, in vitro study, in which differentiating osteoblasts and osteoclasts will be grown in the presence or absence of a glycated matrix. Then, the differentiation and function of cultured osteoblasts and osteoclasts will be assessed after treatment with patients¿ sera and the selected miRNAs. Testing osteoblasts and osteoclasts differentiation and function in glycated conditions too, will help determine whether distinct miRNAs may specifically promote or inhibit the detrimental effect of T2D on bone quality by modulating bone cells function in conditions mimicking the diabetic bone microenvironment.
Should the in vitro study support the clinical outcomes by demonstrating consistent results for some of the differently expressed miRNAs, this would be the first study to identify a mediator of the effect of exercise training on bone quality and fracture risk in T2D. It is possible that some of the identified miRNAs that regulate bone metabolism play additional roles in glucose and lipid metabolism, as shown in previous studies (Wan S. et al. Current Drug Targets 2017), thus affecting bone quality indirectly. Having early biomarkers that are specific to exercise training and diabetes and impact bone quality hold the potential to become established and robust diagnostic tools for diabetic bone disease in clinical research, clinical routine and in personalized medicine.