Lipid droplets synthesized during luteinization are degraded after pregnancy.

After pregnancy, the corpus luteum (CL) functions as a transient endocrine gland that produces progesterone, which is necessary to maintain pregnancy. To maintain constant progesterone production, CLs are enriched in lipids as its precursors. Lipid droplets (LDs) are organelles that originate from the endoplasmic reticulum and store neutral lipids such as triacylglycerols and cholesteryl esters. The size and number of LDs in a cell are regulated by LD-associated proteins that coat their surface. LD degradation is regulated by either neutral lipid hydrolases (lipolysis), selective autophagic mechanism (lipophagy), or both. Mammalian CLs are long known to be enriched in LDs, but LDs are rapidly depleted after pregnancy and reappear near the time of delivery. In this present study, we hypothesized that LDs synthesized by luteinization are massively degraded after pregnancy. Using mCherry-HPos mice, in which LD synthesis can be visualized in vivo, we found that LD synthesis, which was activated during luteal development, was suppressed after implantation. In CLs, LD synthesis remained low during pregnancy, but was reactivated before and after delivery. These changes in LDs were confirmed using electron microscopy and immunostaining. Furthermore, LD degradation was mediated by lipolysis rather than lipophagy. In summary, our findings indicate that luteinization-induced LD synthesis is suppressed after pregnancy onset and that CLs are lipid-poor during pregnancy because LDs stored during luteal development are extensively degraded by lipolysis.

Lipid droplet formation is spatiotemporally regulated in oocytes during follicular development in mice.

Communication between oocytes and the surrounding granulosa cells during follicular development is essential for complete oocyte growth. Oocytes contain lipid droplets (LDs), organelles assembled in the endoplasmic reticulum (ER) that store neutral lipids, including triglycerides and cholesterol esters. Although the LD content varies among animals, LDs stored in oocytes have been shown to play an important role in oocyte maturation and preimplantation embryonic development. However, knowledge is lacking regarding how and when LDs are initially produced in developing oocytes within follicles. In the present study, we found that LDs appeared in mouse oocytes in a specific phase during follicular development. The emergence of LDs in intrafollicular oocytes was induced within a similar time window in vitro and in vivo. Fluorescence imaging and electron microscopy revealed that LDs emerging in oocytes during the early stages of follicular growth were in close proximity to the ER. Furthermore, fatty-acid-tracking experiments have revealed that exogenous fatty acids are rapidly incorporated into oocytes, and their uptake is regulated by the interaction between oocytes and granulosa cells, likely in part through transzonal projections. In summary, our results suggest that LD synthesis observed in growing oocytes is spatiotemporally regulated and that oocyte-granulosa cell contact may be involved in LD biosynthesis during follicular development.

Lipid droplet synthesis is associated with angiogenesis in mouse ovarian follicles†.

Lipid droplets (LDs) are endoplasmic reticulum (ER)-derived organelles comprising a core of neutral lipids surrounded by a phospholipid monolayer. Lipid droplets play important roles in lipid metabolism and energy homeostasis. Mammalian ovaries have been hypothesized to use neutral lipids stored in LDs to produce the hormones and nutrients necessary for rapid follicular development; however, our understanding of LD synthesis remains incomplete. In this study, we generated transgenic reporter mice that express mCherry fused to HPos, a minimal peptide that localizes specifically to nascent LDs synthesized at the ER. With this tool for visualizing initial LD synthesis in ovaries, we found that LDs are synthesized continuously in theca cells but rarely in inner granulosa cells (Gc) during early follicular development. Administration of exogenous gonadotropin enhances LD synthesis in the Gc, suggesting that LD synthesis is hormonally regulated. In contrast, we observed copious LD synthesis in the corpus luteum, and excessive LDs accumulation in atretic follicles. Furthermore, we demonstrated that LD synthesis is synchronized with angiogenesis around the follicle and that suppressing angiogenesis caused defective LD biosynthesis in developing follicles. Overall, our study is the first to demonstrate a spatiotemporally regulated interplay between LD synthesis and neovascularization during mammalian follicular development.

11C-Labeled Radiotracer for Noninvasive and Quantitative Assessment of the Thiocyanate Efflux System in the Brain.

Thiocyanate (SCN-) alters the potency of certain agonists for the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor, and dysfunctions in AMPA receptor signaling are considered to underlie a number of neurological diseases. While humans may be exposed to SCN- from the environment, including food sources, a carrier-mediated system transports SCN- from the brain into the blood and is an important regulator of SCN- distribution in the central nervous system. The assessment of this SCN- efflux system in the brain would thus be useful for understanding the mechanisms underlying the neurotoxicity of SCN- and for elucidating the relationship between the efflux system and brain diseases. However, the currently available technique for studying SCN- efflux is severely limited by its invasiveness. Here, we describe the development of a SCN- protracer, 9-pentyl-6-[11C]thiocyanatopurine ([11C]1), to overcome this limitation. [11C]1 was synthesized by the reaction of the iodo-precursor and [11C]SCN- or the reaction of the disulfide precursor with [11C]NH4CN. The protracer [11C]1 entered the brain after intravenous injection into mice and was rapidly metabolized to [11C]SCN-, which was then eliminated from the brain. The efflux of [11C]SCN- was dose-dependently inhibited by perchlorate, a monovalent anion, and the highest dose caused an 82% reduction in the efflux rate. Our findings demonstrate that [11C]1 can be used for the noninvasive and quantitative assessment of the SCN- efflux system in the brain.

Homeostatic regulation of lipid droplet content in mammalian oocytes and embryos.

Lipid droplets (LDs) consist of a core of neutral lipids such as triacylglycerols and cholesteryl esters covered by a phospholipid monolayer. Recent studies have shown that LDs not only store neutral lipids but are also associated with various physiological functions. LDs are found in most eukaryotic cells and vary in size and quantity. It has long been known that mammalian oocytes contain LDs. Porcine and bovine oocytes contain substantial amounts of LDs, which cause their cytoplasm to darken, whereas mouse and human oocytes are translucent due to their low LD content. A sufficient amount of LDs in mammalian oocytes has been thought to be associated with oocyte maturation and early embryonic development, but the necessity of LDs has been questioned because embryonic development proceeds normally even when LDs are removed. However, recent studies have revealed that LDs play a crucial role during implantation and that maintaining an appropriate amount of LDs is important for early embryonic development, even in mammalian species with low amounts of LDs in their oocytes. This suggests that a fine-tuned balance of LD content is essential for successful mammalian embryonic development. In this review, we discuss the physiological importance of LDs in mammalian oocytes and preimplantation embryos based on recent findings on LD biology.

mRNA decapping factor Dcp1a is essential for embryonic growth in mice.

mRNA decapping is a critical step in posttranscriptional regulation of gene expression in eukaryotes. Although Dcp1a is a well characterized and widely conserved mRNA decapping factor, little is known about its physiological function. To extend our understanding of Dcp1a function in vivo, we employed a transgenic rescue strategy to produce Dcp1a-deficient mice using the CRISPR/Cas9 system. This approach arrowed us to generate heterozygous Dcp1a mice and define the phenotype of Dcp1a-deficient embryos. We found that expression of Dcp1a protein, which is detectable in most mouse tissues, was developmentally regulated through embryonic growth, and that depletion of the Dcp1a gene resulted in embryonic lethality around embryonic day 10.5 (E10.5) concomitant with massive growth retardation and cardiac developmental defects. Moreover, the embryonic lethality was fully rescued by transgenic expression of exogenous human Dcp1a. Together, our results suggest that Dcp1a is required for embryonic growth.

Impact of short-term high-fat feeding on lipid droplet content in mouse oocytes.

Mature mammalian oocytes contain lipid droplets (LDs), which are neutral lipid storage organelles critically important for energy metabolism. In mice, maternal obesity, induced by long-term (> 3 months) high-fat feeding, contributes to the accumulation of LDs in mature oocytes. However, few studies have investigated the influence of short-term high-fat feeding on LD content. In this study, we demonstrated that 3 weeks of high-fat feeding is sufficient to increase LD content and intracellular triacylglycerol levels. Using a two-step centrifugation technique to release LDs into the perivitelline space, we found that short-term high-fat feeding increased the level of LDs in MII oocytes and that 3 days of high-fat feeding were sufficient to increase efficiency of LD release. Collectively, our study suggests that short-term high fat feeding can have a higher impact on lipid metabolism during oocyte maturation.

Synthesis and maintenance of lipid droplets are essential for mouse preimplantation embryonic development.

Lipid droplets (LDs), which are ubiquitous organelles consisting of a neutral lipid core coated with a phospholipid monolayer, play key roles in the regulation of cellular lipid metabolism. Although it is well known that mammalian oocytes and embryos contain LDs and that the amount of LDs varies among animal species, their physiological functions remain unclear. In this study, we have developed a method based on two-step centrifugation for efficient removal of almost all LDs from mouse MII oocytes (delipidation). We found that delipidated MII oocytes could be fertilized in vitro, and developed normally to the blastocyst stage even when the embryos were cultured in the absence of a fatty acid supply. LDs were newly synthesized and accumulated soon after delipidation, but chemical inhibition of long chain acyl-CoA synthetases (ACSLs) blocked this process, resulting in severe impairment of early embryonic development. Furthermore, we found that overabundance of LDs is detrimental to early embryonic development. Our findings demonstrate the importance of synthesis and maintenance of LDs, mediated in part by ACSL activity, during preimplantation embryonic development.