The role of PLIN protein in healthy lipid storage and lipid droplet expansion

In skeletal muscle, fatty acids are stored in vesicle‐like organelles known as lipid droplets (LDs). The importance of how lipids are stored in skeletal muscle has emerged from studies demonstrating a strong association between muscle lipids and insulin sensitivity. In human obesity, excess skeletal muscle lipid is associated with insulin resistance; however, in endurance trained athletes excess lipid storage is associated with an enhanced insulin sensitivity. Recent research has tried to elucidate the different mechanisms of LD synthesis and lipid storage in healthy versus unhealthy individuals. It has been shown that exercise‐trained individuals preferentially store fatty acids as intramuscular triacylglycerides (IMTGs) rather than diacylglycerides (DAGs) and ceramides but the exact mechanisms remain unknown (Chow et al. 2014). High concentrations of DAGs in skeletal muscle have been previously linked to impaired insulin signalling leading to insulin resistance, thus increasing the risk of developing type 2 diabetes (Szendroedi et al. 2014). Although there could be many factors affecting skeletal muscle insulin sensitivity, researchers have focused on the role of LD associated proteins known as perilipins (PLIN). The PLIN family has five members (PLIN 1–5), which, with the exception of PLIN1, are all expressed in skeletal muscle fibres. Our understanding of the specific roles of each PLIN protein in skeletal muscle is in its infancy, with evidence indicating roles in lipid storage, breakdown and transport (for review see MacPherson & Peters, 2015). Thus far, the majority of research investigating skeletal muscle PLIN proteins has explored their role in the breakdown of IMTGs to free fatty acids (FFAs) and glycerol, and the delivery of fatty acids to mitochondria. A role for these proteins in regulating how fatty acids are stored in skeletal muscle from trained individuals and if they play a role in LD growth and expansion remains to be elucidated.

A recent study published in The Journal of Physiology by Shepherd et al. (2017) aimed to determine if the increase in IMTG storage observed in trained individuals was accompanied by a redistribution of the cellular pool of PLIN proteins in response to a simultaneous infusion of lipids and insulin. This unique area of research explores exercise training and the mechanisms behind its effect on healthy lipid storage and LD synthesis. In this study, Shepherd and colleagues (2017) sought to investigate potential differences in the redistribution of skeletal muscle PLIN proteins with increased IMTG storage between trained and sedentary individuals. Specifically, the aim of the study was to determine the effects of lipid infusion with insulin and training status on LD growth and PLIN distribution in type I and II muscle fibre types. The researchers performed muscle biopsies at baseline and two time points (120 min and 360 min) following a 6 h lipid infusion or glycerol infusion (control), as well as a concurrent 6 h hyperinsulinaemic–euglycaemic clamp, which was used to ensure optimal skeletal muscle FFA uptake. Muscle fibre types were then differentiated using staining techniques and immunofluorescence images were captured to analyse the co‐localization of individual PLIN proteins with LDs, as well as LD density and size (Shepherd et al. 2017).

The increased skeletal muscle lipid uptake and storage with a lipid infusion is expected; however, the novelty of this study arises with the analysis of the changes in skeletal muscle LD number/size and associated PLIN proteins in each fibre type. Shepherd et al. (2017) demonstrated a higher association of PLIN2, PLIN3 and PLIN5 with LDs in type I fibres of trained individuals while the number of LDs devoid of PLIN2, 3 and 5 increased in the sedentary group. In addition, the rise in type I fibre IMTG content was accompanied by increases in both LD size and density in the trained individuals, while sedentary individuals only had an increase in LD density following lipid infusion. This finding provides evidence for a role of these skeletal muscle PLIN proteins aiding in the storage and expansion of the LD pool in trained individuals. As an explanation, the authors speculate that the increased accumulation and size of LDs in trained individuals may be due to an increased ability to store a more flexible or dynamic LD pool. The authors also commented on the possibility that lower PLIN content in sedentary participants could lead to less PLIN available for redistribution following lipid infusion. This introduces the hypothesis that the availability of PLIN proteins prior to a lipid load is important for the safe storage of fatty acids in healthy individuals.

An alternative hypothesis to the findings by Shepherd and colleagues (2017) is that both trained and sedentary individuals have the ability to take up and store fatty acids in nascent LDs following infusion; however, due to more PLIN + LDs and increases in LD size in trained individuals, it could be that trained individuals have an increased capacity to expand the size of the LDs. This explanation would introduce a role for PLIN proteins in expanding and maturing nascent LDs. Healthy and trained individuals not only have an increased number, but also have a better ability to expand LDs, which could be due to a faster rate of nascent droplets maturing or the fusion of already matured LDs. Following this hypothesis, the increase in storage of FFAs to IMTGs seen in healthy individuals could be linked to the larger, more mature LDs, and the increase in PLIN + LDs. Future studies could focus on individual PLIN functions and their role with this proposed mechanism of LD expansion.

In the current study the authors did not measure changes in DAG or ceramide content between individuals. Our team believes this would be an important measure for future studies, as Chow et al. (2014) showed an increase of DAGs in sedentary individuals after an acute lipid elevation compared to trained participants. Due to the association with insulin resistance, this information could have tied in further health implications such as the risk of insulin resistance and type 2 diabetes. It may further provide insight into the roles that PLIN proteins may play in regulating how the fatty acids are stored. We believe the authors’ choice to measure PLIN4, despite its role remaining very much unknown, was a clear strength of the paper. Unlike the other PLINs, Shepherd et al. (2017) discovered that lipid infusion increased the density of LDs not associated with PLIN4 equally in both groups and no changes were seen in LDs associated with PLIN4. This result further exemplifies a functional difference of PLIN4 and its unique role beyond that of lipid storage in skeletal muscle. In line with this, Pourteymour et al. (2015) investigated the intracellular location of PLIN4 and discovered its location was predominantly on the subsarcolemmal LDs versus being uniformly expressed throughout the muscle fibre. Using this novel co‐localization technique adopted by Shepherd's group, future work could focus on PLIN redistribution with respect to cellular location to determine where PLIN proteins are being redistributed within the muscle fibre. The location of redistribution in response to lipid infusion could further uncover the function of these PLIN proteins. For example, a redistribution to the subsarcolemmal region may indicate a role in assisting in sarcolemmal transport of fatty acids into the cell and the initial growth of LDs in this region. Future studies could also include analysis at time points exceeding 360 min after infusion to determine if and when PLIN association with LD is recovered to the baseline level and how this varies between training groups.

In conclusion, this novel work by Shepherd et al. (2017) has brought us one step closer to understanding the mechanisms underlying healthy lipid storage and LD synthesis through comparison of exercise‐trained and sedentary individuals. The finding that the trained group had increased association of LDs with PLIN2, 3 and 5 post‐lipid infusion provides novel insight into the notion that PLIN proteins assist in healthy skeletal muscle lipid storage and LD growth. This discovery could have implications for PLIN proteins having a role in FFA storage and not solely IMTG breakdown. This suggests that PLIN proteins facilitate favourable IMTG synthesis from FFAs in trained individuals rather than what is seen in sedentary individuals who accumulate more FFAs post infusion and LDs not associated with PLIN proteins. The main findings support the authors’ hypothesis that trained individuals have a more dynamic or flexible PLIN pool, as well as our proposal of nascent droplets maturing faster due to PLIN redistribution in trained individuals. This study provides evidence for the mechanisms behind healthy, exercise‐trained individuals having an advantage in lipid metabolism. Furthermore, it has laid the groundwork for future studies to examine PLIN redistribution and LD synthesis/growth mechanisms in other models, including a larger spectrum of individuals from obese to lean individuals and across various age groups and sexes.

Additional information

Competing interests

None declared.

Author contributions

All authors contributed to the conception or design of the work and drafting the work or revising it critically for important intellectual content. All authors have approved the final version of the manuscript and agree to be accountable for all aspects of the work. All persons designated as authors qualify for authorship, and all those who qualify for authorship are listed.

Funding

The lab is currently funded by NSERC through G.C.H.’s supervisor Dr Rebecca MacPherson.