MicroRNAs are fine regulators of the whole-body adaptive response but their use as biomarkers is limited by the lack of standardized pre- and post-analytical procedures. This work aimed to compare different normalization approaches for RT-qPCR data analyses, in order to identify the most reliable and reproducible method to analyze circulating miRNA expression profiles in sedentary and highly-trained subjects. As the physically active status is known to affect miRNA expression, they could be effective biomarkers of the homeostatic response. Following RNA extraction from plasma, a panel of 179 miRNAs was assayed by RT-qPCR and quantified by applying different normalization strategies based on endogenous miRNAs and exogenous oligonucleotides. hsa-miR-320d was found as the most appropriate reference miRNA in reducing the technical variability among the experimental replicates and, hence, in highlighting the inter-cohorts differences. Our data showed an association between the physically active status and specific skeletal muscle- and bone-associated circulating miRNAs profiles, revealing that established epigenetic modifications affect the baseline physiological status of these tissues. Since different normalization strategies led to different outputs, in order to avoid misleading interpretation of data, we remark the importance of the accurate choice of the most reliable normalization method in every experimental setting.
Physical inactivity has been recognized, by the World Health Organization as the fourth cause of death (5.5 % worldwide). On the contrary, physical activity (PA) has been associated with improved quality of life and decreased risk of several diseases (i.e., stroke, hypertension, myocardial infarction, obesity, malignancies). Bone turnover is profoundly affected from PA both directly (load degree is the key determinant for BMD) and indirectly through the activation of several endocrine axes. Several molecules, secreted by muscle (myokines) and adipose tissues (adipokines) in response to exercise, are involved in the fine regulation of bone metabolism in response to the energy availability. Furthermore, bone regulates energy metabolism by communicating its energetic needs thanks to osteocalcin which acts on pancreatic β-cells and adipocytes. The beneficial effects of exercise on bone metabolism depends on the intermittent exposure to myokines (i.e., irisin, IL-6, LIF, IGF-I) which, instead, act as inflammatory/pro-resorptive mediators when chronically elevated; on the other hand, the reduction in the circulating levels of adipokines (i.e., leptin, visfatin, adiponectin, resistin) sustains these effects as well as improves the whole-body metabolic status. The aim of this review is to highlight the newest findings about the exercise-dependent regulation of these molecules and their role in the fine regulation of bone metabolism.
Highlights d Millimeter-scale model of a vascularized 3D human muscle environment d Endothelial cells acquire organ-specific signature in co-culture with muscle cells d Muscle fibroblasts spontaneously self-assemble an endomysium-like structure d Physiologically relevant 3D model for the study of muscle fibrosis
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