Martin Whitham

Small extracellular vesicles (sEVs) are released from all cell types and participate in the intercellular exchange of proteins, lipids, metabolites and nucleic acids. Proteomic, flow cytometry and nanoparticle tracking analyses suggest sEVs are released into circulation with exercise. However, interpretation of these data may be influenced by sources of bias introduced by different analytical approaches. Seven healthy participants carried out a high intensity intermittent training (HIIT) cycle protocol consisting of 4 × 30 s at a work‐rate corresponding to 200% of individual max power (watts) interspersed by 4.5 min of active recovery. EDTA‐treated blood was collected before and immediately after the final effort. Platelet‐poor (PPP) and platelet‐free (PFP) plasma was derived by one or two centrifugal spins at 2500 g, respectively (15 min, room temperature). Platelets were counted on an automated haemocytometer. Plasma samples were assessed with the Exoview R100 platform, which immobilises sEVs expressing common tetraspanin markers CD9, CD63, CD81 and CD41a on microfluidic chips and with the aid of fluorescence imaging, counts their abundance at a single sEV resolution, importantly, without a pre‐isolation step. There was a lower number of platelets in the PFP than PPP, which was associated with a lower number of CD9, CD63 and CD41a positive sEVs. HIIT induced an increase in fluorescence counts in CD9, CD63 and CD81 positive sEVs in both PPP and PFP. These data support the concept that sEVs are released into circulation with exercise. Furthermore, platelet‐free plasma is the preferred, representative analyte to study sEV dynamics and phenotype during exercise. image Key points: Small extracellular vesicles (sEV) are nano‐sized particles containing protein, metabolites, lipid and RNA that can be transferred from cell to cell. Previous findings implicate that sEVs are released into circulation with exhaustive, aerobic exercise, but since there is no gold standard method to isolate sEVs, these findings may be subject to bias introduced by different approaches. Here, we use a novel method to immobilise and image sEVs, at single‐vesicle resolution, to show sEVs are released into circulation with high intensity intermittent exercise. Since platelet depletion of plasma results in a reduction in sEVs, platelet‐free plasma is the preferred analyte to examine sEV dynamics and phenotype in the context of exercise.

Extracellular vesicles (EVs) can be released from most cells in the body and act as intercellular messengers transferring information in their cargo to affect cellular function. A growing body of evidence suggests that a subset of EVs, referred to here as ‘small extracellular vesicles’ (sEVs), can accelerate or slow the processes of ageing and age‐related diseases dependent on their molecular cargo and cellular origin. Continued exploration of the vast complexity of the sEV cargo aims to further characterise these systemic vehicles that may be targeted to ameliorate age‐related pathologies. Marked progress in the development of mass spectrometry‐based technologies means that it is now possible to characterise a significant proportion of the proteome of sEVs (surface and cargo) via unbiased proteomics. This information is vital for identifying biomarkers and the development of sEV‐based therapeutics in the context of ageing. Although exercise and physical activity are prominent features in maintaining health in advancing years, the mechanisms responsible are unclear. A potential mechanism by which plasma sEVs released during exercise could influence ageing and senescence is via the increased delivery of cargo proteins that function as antioxidant enzymes or inhibitors of senescence. These have been observed to increase in sEVs following acute and chronic exercise, as identified via independent interrogation of high coverage, publicly available proteomic datasets. Establishing tropism and exchange of functionally active proteins by these processes represents a promising line of enquiry in implicating sEVs as biologically relevant mediators of the ageing process. image

Physical activity has systemic effects on the body, affecting almost every organ. It is important not only for general health and wellbeing, but also in the prevention of diseases. The mechanisms behind the therapeutic effects of physical activity are not completely understood; however, studies indicate these benefits are not confined to simply managing energy balance and body weight. They also include systemic factors which are released into the circulation during exercise and which appear to underlie the myriad of benefits exercise can elicit. It was shown that along with a number of classical cytokines, active tissues also engage in inter-tissue communication via extracellular vesicles (EVs), specifically exosomes and other small EVs, which are able to deliver biomolecules to cells and alter their metabolism. Thus, EVs may play a role in the acute and systemic adaptations that take place during and after physical activity, and may be therapeutically useful in the treatment of a range of diseases, including metabolic disorders such as type 2 diabetes and obesity; and the focus of this review, neurological disorders such as Alzheimer’s disease.

The systemic response to exercise is dose‐dependent and involves a complex gene expression regulation and cross‐talk between tissues. This context ARISES the need for analyzing the influence of exercise dose on the profile of circulating microRNAs (c‐miRNAs), as emerging posttranscriptional regulators and intercellular communicators. Thus, we hypothesized that different exercise doses will determine specific c‐miRNA signatures that will highlight its potential as exercise dose biomarker. Nine active middle‐aged males completed a 10‐km race (10K), a half‐marathon (HM), and a marathon (M). Blood samples were collected immediately before and after races. Plasma RNA was extracted, and a global screening of 752 microRNAs was analyzed using RT‐qPCR. Three different c‐miRNA profiles were defined according to the three doses. In 10K, 14 c‐miRNAs were found to be differentially expressed between pre‐ and post‐exercise, 13 upregulated and 1 downregulated. Regarding HM, 13 c‐miRNAs were found to be differentially modulated, in all the cases upregulated. A total of 28 c‐miRNAs were found to be differentially expressed in M, 21 overexpressed and 7 repressed after this race. We had also found 3 common c‐miRNAs between 10K and M and 2 common c‐miRNAs between 10K and HM. In silico analysis supported a close association between exercise dose c‐miRNA profiles and cellular pathways linked to energy metabolism and cell cycle. In conclusion, we have observed that different exercise doses induced specific c‐miRNA profiles. So, our results point to c‐miRNAs as emerging exercise dose biomarkers and as one of regulatory mechanisms modulating the response to endurance exercise.

Type 2 diabetes (T2D) manifests from inadequate glucose control due to insulin resistance, hypoinsulinemia, and deteriorating pancreatic β‐cell function. The pro‐inflammatory factor Activin has been implicated as a positive correlate of severity in T2D patients, and as a negative regulator of glucose uptake by skeletal muscle, and of pancreatic β‐cell phenotype in mice. Accordingly, we sought to determine whether intervention with the Activin antagonist Follistatin can ameliorate the diabetic pathology . Here, we report that an intravenous Follistatin gene delivery intervention with tropism for striated muscle reduced the serum concentrations of Activin B and improved glycemic control in the db/db mouse model of T2D. Treatment reversed the hyperglycemic progression with a corresponding reduction in the percentage of glycated‐hemoglobin to levels similar to lean, healthy mice. Follistatin gene delivery promoted insulinemia and abundance of insulin‐positive pancreatic β‐cells, even when treatment was administered to mice with advanced diabetes, supporting a mechanism for improved glycemic control associated with maintenance of functional β‐cells. Our data demonstrate that single‐dose intravascular Follistatin gene delivery can ameliorate the diabetic progression and improve prognostic markers of disease. These findings are consistent with other observations of Activin‐mediated mechanisms exerting deleterious effects in models of obesity and diabetes, and suggest that interventions that attenuate Activin signaling could help further understanding of T2D and the development of novel T2D therapeutics.

It has been suggested that interleukin-6 (IL-6) produced by adipocytes in obesity leads to liver insulin resistance, although this hypothesis has never been definitively tested. Accordingly, we did so by generating adipocyte-specific IL-6-deficient (AdipoIL-6^−/−) mice and studying them in the context of diet-induced and genetic obesity. Mice carrying two floxed alleles of IL-6 (C57Bl/6J) were crossed with Cre recombinase-overexpressing mice driven by the adiponectin promoter to generate AdipoIL-6^−/− mice. AdipoIL-6^−/− and floxed littermate controls were fed a standard chow or high-fat diet (HFD) for 16 wk and comprehensively metabolically phenotyped. In addition to a diet-induced obesity model, we also examined the role of adipocyte-derived IL-6 in a genetic model of obesity and insulin resistance by crossing the AdipoIL-6^−/− mice with leptin-deficient ( ob/ob) mice. As expected, mice on HFD and ob/ob mice displayed marked weight gain and increased fat mass compared with chow-fed and ob/+ (littermate control) animals, respectively. However, deletion of IL-6 from adipocytes in either model had no effect on glucose tolerance or fasting hyperinsulinemia. We concluded that adipocyte-specific IL-6 does not contribute to whole body glucose intolerance in obese mice.

The health-promoting effects of physical activity to prevent and treat metabolic disorders are numerous. However, the underlying molecular mechanisms are not yet completely deciphered. In recent years, studies have referred to the liver as an endocrine organ, since it releases specific proteins called hepatokines. Some of these hepatokines are involved in whole body metabolic homeostasis and are theorized to participate in the development of metabolic disease. In this regard, the present review describes the role of Fibroblast Growth Factor 21, Fetuin-A, Angiopoietin-like protein 4, and Follistatin in metabolic disease and their production in response to acute exercise. Also, we discuss the potential role of hepatokines in mediating the beneficial effects of regular exercise and the future challenges to the discovery of new exercise-induced hepatokines.

Protein signaling between tissues, or tissue cross‐talk is becoming recognized as a fundamental biological process that is incompletely understood. Shotgun proteomic analyses of tissues and plasma to explore this concept are regularly challenged by high dynamic range of protein abundance, which limits the identification of lower abundance proteins. In this viewpoint article, it is highlighted how a focus on proteins contained within extracellular vesicles (EVs) not only partially addresses this issue, but can also reveal an underappreciated complexity of the circulating proteome in various physiological and pathological contexts. Furthermore, how quantitative proteomics can inform EV mediated crosstalk is highlighted and the importance of high coverage, sensitive proteomic analyses of EVs to identify both the optimal methods to isolate EV subtypes of interest and proteins that characterize them is stressed.

Exercise stimulates a wide array of biological processes, but the mechanisms involved are incompletely understood. Many previous studies have adopted transcriptomic analyses of skeletal muscle to address particular research questions, a process that ultimately results in the collection of large amounts of publicly available data that has not been fully integrated or interrogated. To maximize the use of these available transcriptomic exercise data sets, we have downloaded and reanalyzed them and formulated the data into a searchable online tool, geneXX. GeneXX is highly intuitive and free and provides immediate information regarding the response of a transcript of interest to exercise in skeletal muscle. To demonstrate its utility, we carried out a meta-analysis on the included data sets and show transcript changes in skeletal muscle that persist regardless of sex, exercise mode, and duration, some of which have had minimal attention in the context of exercise. We also demonstrate how geneXX can be used to formulate novel hypotheses on the complex effects of exercise, using preliminary data already generated. This resource represents a valuable tool for researchers with interests in human skeletal muscle adaptation to exercise.