Cytoplasmic Droplets Research Articles

Abstract 206: Intestinal LXR Directs Absorbed Lipids to Storage

Post-prandial hypertriglyceridemia has emerged as a cardiovascular risk factor with limited therapeutic options. We have taken genetic and physiological approaches to identify and characertize novel factors that regulate intestinal lipid handling in the hope of revealing cellular pathways that dampen delivery of dietary lipids, opening the therapeutic window to blunting post-prandial lipid excursions. Liver X receptors (Lxrs) are master regulators of cholesterol catabolism, driving the elimination of cholesterol from the periphery to the lumen of the intestine. These nuclear hormone receptors are activated by oxysterol, ligands that accumulate when excess cholesterol is present. Loss of nr1h3 function causes anticipated gene regulatory changes and cholesterol intolerance, collectively reflecting high evolutionary conservation of zebrafish Lxra function. Intestinal nr1h3 activation delays transport of absorbed neutral lipids, with accumulation of neutral lipids in enterocyte cytoplasmic droplets, as visualized with histological stains, live fluorescent lipid probes, and ultrastructural analyses. This delay in transport of ingested neutral lipids protects animals over-expressing nr1h3 in the intestine from hypercholesterolemia and hepatic steatosis induced by a high-fat diet. On a gene regulatory level, Lxra induces expression of acsl3a , which encodes acyl-CoA synthetase long-chain family member 3a, a lipid droplet-anchored protein that directs fatty acyl chains into lipids. We have identified a gene regulatory network in enterocytes whereby the pace of delivery of dietary lipids is gated by Lxrs. Development of pharmacological agents to activate Lxrs has been hindered by synthetic Lxr agonists’ induction of hepatic lipogenesis and hypertriglyceridemia. The development of intestine-limited Lxr agonists could be of therapeutic value in blunting post-prandial hypertriglyceridemia.

Electrical signalling in Nitellopsis obtusa: potential biomarkers of biologically active compounds.

The Nitellopsis obtusa (N.A.Desvaux) J.Groves cell provides a model system for complex investigation of instantaneous effects of various biologically active compounds (BC) on the generation of plant bioelectrical signals in vivo. Experimental evidence using multiple electrical signals as biomarkers of the effects of BC (acetylcholine, asparagine, glutamate, nicotine, aluminium, nickel and cadmium ions) is provided. The effect of BC on membrane transport systems involved in the cell excitability were tested by current clamp, voltage clamp and patch clamp methods. Membrane potential (MP) alterations and action potential (AP) patterns in response to BC were shown to represent the cell state. High discretisation frequency allows precise, high time resolution analysis of real-time processes measuring changes in excitation threshold, AP amplitude and velocity of repolarisation values after application of BC indicating the effect on ion channels involved in AP generation. Application of voltage clamp revealed that changes in AP peak value were caused not only by increment in averaged maximum amplitude of the Cl- current, but in prolonged Cl- channels' opening time also. The cytoplasmic droplet can serve as a model system in which the effects of BC on single tonoplast ion channel can be studied by patch clamping. Investigation of electrical cell-to-cell communication revealed evidence on the electrical signal transduction through plasmodesmata.

Glassy droplet inclusions within the cytoplasm of Kupffer cells: A novel ultrastructural feature for the diagnosis of pediatric autoimmune hepatitis

Since Kupffer cells/macrophages (KCs/MPs) may be involved in the pathogenesis of autoimmune hepatitis (AIH), this pioneer study was undertaken to evaluate KCs/MPs in pediatric AIH in transmission-electron microscope. MethodsUltrastructural analyses were performed using liver biopsies from 14 children with clinicopathologically diagnosed AIH. ResultsIn all AIH children, ultrastructural findings revealed changes in the cells lining sinusoidal vessels, especially KCs/MPs and endothelial cells. KCs/MPs showed increased phagocytic activity and damaged mitochondria, frequently with accompanying intense fibrosis. In 10/14 AIH patients, the cytoplasm of sinusoidal KCs/MPs contained characteristic glassy droplet inclusions. They were round, sharply circumscribed, and contained homogeneous material and distinct translucent fields. Their ultrastructure was identical with the Russel bodies of plasma cells, which were also found in liver biopsies in the same patients. ConclusionUltrastructural identification of characteristic cytoplasmic droplets with glassy appearance in KCs/MPs, never before described in AIH, provides a useful novel morphological feature in the diagnosis of this disease.

Ageing is associated with brown adipose tissue remodelling and loss of white fat browning in female C57BL/6 mice.

Fat storage changes throughout life and affects body metabolism. Ageing impact on brown (BAT) and white adipose tissue (WAT) deserves attention, especially in females, because they are less prone to age-induced weight gain. While in male mice the impact of ageing on adipose tissue remodelling is well characterized, the effects in female mice remain largely unclear. Thus, we investigated BAT and WAT remodelling during ageing in female C57BL/6 mice. At 3months, body weight was 24±0.3g (mean±SD), and it increased from 6 to 9months of age (+20%, P<0.0001). Oral glucose tolerance test showed no disturbance of glucose metabolism. All WAT depots became heavier, and white adipocytes hypertrophied. The subcutaneous and visceral WAT had clusters of beige cells in younger mice, but they were progressively lost by ageing, indicating loss of WAT browning. Older mice had hypertrophied classic brown adipocytes that had larger cytoplasmic lipid droplets than younger mice. Pearson's correlation showed that WAT mass has a weak correlate with BAT mass, although white adipocyte diameter has a strong correlation with classic brown adipocyte size. In conclusion, our results indicate that female C57BL/6 mice have a progressive age-dependent loss of subcutaneous and visceral WAT browning, and this process runs in parallel with BAT morphological changes towards a fat storer phenotype, independent of cycling or disturbances in glucose metabolism.

Differences in the Enterocyte Cytoplasmic Lipid Droplet Proteome in Mice Resistant (Acyl CoA: Diacylglycerol Acyltransferase 1 Deficient) versus Susceptible to Diet‐Induced Obesity

The highly efficient absorption of dietary fat is essential for survival but can contribute to obesity and related metabolic diseases when dietary fat is consumed in excess. The digestive products of dietary fat are taken up by absorptive cells of the small intestine (enterocytes), re‐esterified into triacylglycerol (TAG), and either packaged on chylomicrons (CMs) for secretion into circulation or stored within cytoplasmic lipid droplets (CLDs). Altered enterocyte TAG storage and secretion impacts susceptibility to high fat diet‐induced diseases, such as obesity, diabetes, and hepatic steatosis, but molecular mechanisms directing TAG toward these fates are unclear. Acyl CoA: diacylglycerol acyltransferases (DGATs) are the enzymes that catalyze the final and committed step of TAG synthesis, and DGAT1 is the major DGAT enzyme present in the small intestine. Whole body Dgat1 deficient (Dgat1−/−) mice are resistant to diet‐induced obesity and exhibit improved glucose tolerance and blunted postprandial triglyceridemia compared to high fat fed wild‐type mice. In addition, these mice exhibit an accumulation of TAG within large CLDs in enterocytes in response to an acute dietary fat challenge. We hypothesized that the lean, insulin sensitive phenotype of these mice when fed a chronic high fat diet may be due to alterations in intestinal TAG storage that are regulated by the proteins that associate with enterocyte CLDs. To investigate this we isolated enterocyte CLDs from wild‐type and Dgat1−/− mice after 12 weeks of high fat feeding and determined their proteomes using LC‐MS/MS. Label‐free quantification resulted in the identification of 125 proteins spanning diverse cellular processes, including lipid metabolism. Several of the identified proteins were differentially expressed in Dgat1−/− compared to wild‐type mice, and over a third of these proteins were only identified in the Dgat1−/− model. Further investigation of these proteins will contribute to molecular knowledge of the regulation of enterocyte CLD stores, as well as the role of CLDs in the process of dietary fat absorption. This knowledge can be used to identify potential therapeutic targets for the treatment of obesity, type II diabetes, and cardiovascular disease.Support or Funding InformationThis project was supported by the Purdue University Ingestive Behavior Research Center NIH Training Grant (5T32DK076540‐06, 5T32DK076540‐07) and the American Diabetes Association Innovation Award (7‐13‐IN‐05).

Differential proteomics profiling identifies LDPs and biological functions in high-fat diet-induced fatty livers

Eukaryotic cells store neutral lipids in cytoplasmic lipid droplets (LDs) enclosed in a monolayer of phospholipids and associated proteins [LD proteins (LDPs)]. Growing evidence has demonstrated that LDPs play important roles in the pathogenesis of liver diseases. However, the composition of liver LDPs and the role of their alterations in hepatosteatosis are not well-understood. In this study, we performed liver proteome and LD sub-proteome profiling to identify enriched proteins in LDs as LDPs, and quantified their changes in a high-fat diet (HFD)-induced fatty liver model. Among 5,000 quantified liver proteins, 101 were enriched by greater than 10-fold in the LD sub-proteome and were classified as LDPs. Differential profiling of LDPs in HFD-induced fatty liver provided a list of candidate LDPs for functional investigation. We tested the function of an upregulated LDP, S100a10, in vivo with adenovirus-mediated gene silencing and found, unexpectedly, that knockdown of S100a10 accelerated progression of HFD-induced liver steatosis. The S100A10 interactome revealed a connection between S100A10 and lipid transporting proteins, suggesting that S100A10 regulates the development and formation of LDs by transporting and trafficking. This study identified LD-enriched sub-proteome in homeostatic as well as HFD-induced fatty livers, providing a rich resource for the LDP research field.

Cytoplasmic droplet acting as a mitochondrial modulator during sperm maturation in dogs

Motility acquisition during sperm maturation and passage through the epididymis is closely related to mitochondrial function and appears to occur in parallel with cytoplasmic droplet (CD) migration. However, such mechanism remains unclear in dogs. Thus, the aim of this study was to characterize the influence of sperm CD in the mitochondrial functionality during epididymal sperm maturation in dogs. Twenty-one adult dogs were submitted to elective bilateral orchiectomy. Testicles were stored for 18–24h at 5°C and epididymal sperm samples were then collected from different segments of the epididymis (caput, corpus and cauda). Samples were evaluated for computer-assisted motility analysis (CASA), presence of CD (eosin/nigrosin stain), ultrastructural CD analysis and sperm mitochondrial activity (3,3' diaminobenzidine technique) and membrane potential (JC-1 probe). Samples collected from the corpus epididymis showed higher motility and mitochondrial activity in comparison to the caput sperm. Moreover, corpus sperm had lower percentage of proximal droplets compared to caput samples, while mitochondrial membrane potential remained unchanged. Cauda samples showed higher motility, mitochondrial activity and potential, however, lower presence of sperm droplets (proximal and distal). In conclusion, the CD is essential for epididymal sperm maturation in dogs, showing important functions along the transit in the epididymis. In the corpus segment, the migration of the CD along the sperm midpiece provides a high mitochondrial activity and the onset of sperm motility. On the other hand, sperm from cauda epididymis lack CD but suffered lipid membrane changes which allow a maximum mitochondrial membrane potential and motility.

Human Subtilisin Kexin Isozyme-1 (SKI-1)/Site-1 Protease (S1P) regulates cytoplasmic lipid droplet abundance: A potential target for indirect-acting anti-dengue virus agents.

Viral hijacking and manipulation of host-cell biosynthetic pathways by human enveloped viruses are shared molecular events essential for the viral lifecycle. For Flaviviridae members such as hepatitis C virus and dengue virus (DENV), one of the key subsets of cellular pathways that undergo manipulation is the lipid metabolic pathways, underlining the importance of cellular lipids and, in particular, lipid droplets (LDs) in viral infection. Here, we hypothesize that targeting cellular enzymes that act as key regulators of lipid homeostasis and LD formation could represent a powerful approach to developing a novel class of broad-spectrum antivirals against infection associated with all DENV serotypes (1–4) circulating around the world. Using PF-429242, an active-site-directed inhibitor of SKI-1/S1P, we demonstrate that inhibition of SKI-1/S1P enzymatic activity in human hepatoma Huh-7.5.1 cells results in a robust reduction of the LD numbers and LD-positive areas and provides a means of effectively inhibiting infection by DENV (1–4). Pre-treatment of Huh-7.5.1 cells with PF-429242 results in a dose-dependent inhibition of DENV infection [median inhibitory dose (EC50) = 1.2 microM; median cytotoxic dose (CC50) = 81 microM; selectivity index (SI) = 68)] and a ~3-log decrease in DENV-2 titer with 20 microM of PF-429242. Post-treatment of DENV-2 infected Huh-7.5.1 cells with PF-429242 does not affect viral RNA abundance, but it does compromise the assembly and/or release of infectious virus particles. PF-429242 antiviral activity is reversed by exogenous oleic acid, which acts as an inducer of LD formation in PF-429242-treated and non-treated control cells. Collectively, our results demonstrate that human SKI-1/S1P is a potential target for indirect-acting pan-serotypic anti-DENV agents and reveal new therapeutic opportunities associated with the use of lipid-modulating drugs for controlling DENV infection.

Differentiation of bovine mammary epithelial cells in the presence of linolenic acid in combination with thiazolidenediones

Cytoplasmic lipid droplets (CLD's) formation is critical for lactation and health and so it could be detrimental in upholding the formation of CLD's in bovine mammary epithelial cells (MAC-T). We therefore, treated MAC-T cells with differentiation medium containing alpha-linolenic acid (ALA) (100 μM) and thiazolidenediones (TZD's)(10 μM) and observed CLD's formation in cellular cytoplasm with Oil-red-O staining and elution index percentage. We also observed significant up-regulation of adipogenic and down regulation of epithelial markers. In conclusion, 100 μM ALA plus 10 μM TZD's resulted in formation of CLD's and subsequent differentiation of MAC-T cells.

Dgat1 and Dgat2 regulate enterocyte triacylglycerol distribution and alter proteins associated with cytoplasmic lipid droplets in response to dietary fat

Enterocytes, the absorptive cells of the small intestine, mediate efficient absorption of dietary fat (triacylglycerol, TAG). The digestive products of dietary fat are taken up by enterocytes, re-esterified into TAG, and packaged on chylomicrons (CMs) for secretion into blood or temporarily stored within cytoplasmic lipid droplets (CLDs). Altered enterocyte TAG distribution impacts susceptibility to high fat diet associated diseases, but molecular mechanisms directing TAG toward these fates are unclear. Two enzymes, acyl CoA: diacylglycerol acyltransferase 1 (Dgat1) and Dgat2, catalyze the final, committed step of TAG synthesis within enterocytes. Mice with intestine-specific overexpression of Dgat1 (Dgat1Int) or Dgat2 (Dgat2Int), or lack of Dgat1 (Dgat1−/−), were previously found to have altered intestinal TAG secretion and storage. We hypothesized that varying intestinal Dgat1 and Dgat2 levels alters TAG distribution in subcellular pools for CM synthesis as well as the morphology and proteome of CLDs. To test this we used ultrastructural and proteomic methods to investigate intracellular TAG distribution and CLD-associated proteins in enterocytes from Dgat1Int, Dgat2Int, and Dgat1−/− mice 2h after a 200μl oral olive oil gavage. We found that varying levels of intestinal Dgat1 and Dgat2 altered TAG pools involved in CM assembly and secretion, the number or size of CLDs present in enterocytes, and the enterocyte CLD proteome. Overall, these results support a model where Dgat1 and Dgat2 function coordinately to regulate the process of dietary fat absorption by preferentially synthesizing TAG for incorporation into distinct subcellular TAG pools in enterocytes.

Downsizing cumulus cell layers to improve cryotolerance of germinal vesicle-stage bovine oocytes

This study was undertaken to investigate whether complete removal or downsizing of the cumulus cell layers in germinal vesicle (GV)-stage bovine cumulus-oocyte complexes (COCs) can improve blastocyst development rate following Cryotop vitrification. Downsized COCs (196 μm in mean diameter) and denuded oocytes (141 μm in mean diameter) were prepared by vortex-mixing of full-sized COCs (330 μm in mean diameter) retrieved from abattoir-derived ovaries. Nuclear maturation rates, assessed by the first polar body extrusion, after vitrification and the subsequent 22-h IVM were comparable (61.9–62.9%). Approximately one-third (30.5–31.2%) of the matured oocytes derived from the downsized COCs could develop into high quality blastocysts after 6-h IVF and 8-d IVC, while 13.4 and 23.7% of the matured oocytes derived from denuded oocytes and full-size COCs reached to the blastocysts, respectively. Cytoplasmic lipid droplets of matured oocytes in vitrification group were more clustered with decreased number and increased size of the droplets, when compared to those in fresh control group. However, individual oocyte culture in well-of-the well system suggested that change of lipid droplet distribution in the matured oocytes had no adverse effect on their subsequent developmental competence up to the blastocyst stage. In conclusion, Cryotop vitrification of downsized GV-stage bovine COCs allowed blastocyst yields as high as >30%.

The lipid droplet-associated protein perilipin 3 facilitates hepatitis C virus-driven hepatic steatosis

Hepatitis C virus (HCV) is an enveloped RNA virus responsible for 170 million cases of viral hepatitis worldwide. Over 50% of chronically infected HCV patients develop hepatic steatosis, and steatosis can be induced by expression of HCV core protein (core) alone. Additionally, core must associate with cytoplasmic lipid droplets (LDs) for steatosis development and viral particle assembly. Due to the importance of the LD as a key component of hepatic lipid storage and as a platform for HCV particle assembly, it seems this dynamic subcellular organelle is a gatekeeper in the pathogenesis of viral hepatitis. Here, we hypothesized that core requires the host LD scaffold protein, perilipin (PLIN)3, to induce hepatic steatosis. To test our hypothesis in vivo, we have studied core-induced hepatic steatosis in the absence or presence of antisense oligonucleotide-mediated knockdown of PLIN3. PLIN3 knockdown blunted HCV core-induced steatosis in transgenic mice fed either chow or a moderate fat diet. Collectively, our studies demonstrate that the LD scaffold protein, PLIN3, is essential for HCV core-induced hepatic steatosis and provide new insights into the pathogenesis of HCV.

The CatSper channel modulates boar sperm motility during capacitation

The cation channel of sperm (CatSper) comprises four transmembrane subunits specifically expressed in human, equine, murine and ovine spermatozoa, apparently implicated in capacitation, hyperactivation and acrosome exocytosis. Western blotting and immunocytochemistry showed hereby that CatSper subunits are also present in boar spermatozoa, primarily over the sperm neck, tail and cytoplasmic droplets; albeit CatSper −1 presented in addition some distribution over the membrane of the acrosome and CatSper −2 and −4 over the membrane of the post-acrosome. The role of the Catsper channel in boar spermatozoa was investigated by extending the spermatozoa in media containing different calcium (Ca2+) availability and exposure to the capacitation-trigger bicarbonate, to progesterone or CatSper inhibitors (Mibefradil and NNC 55-0396), separately or sequentially, at physiological and toxicological doses. Extracellular Ca2+ availability, combined with bicarbonate exposure (capacitation-inducing conditions) decreased sperm motility, similarly to when spermatozoa incubated in capacitation-inducing conditions was exposed to Mibefradil and NNC 55-0396. Exposure of these spermatozoa to progesterone did not cause significant changes in sperm motility and nor did it revert its decrease induced by CatSper antagonists. In conclusion, the CatSper channel regulates sperm motility during porcine capacitation-related events in vitro.

Head morphometry and chromatin instability in normal boar spermatozoa and in spermatozoa with cytoplasmic droplets

Head morphometry and chromatin instability in normal boar spermatozoa and in spermatozoa with cytoplasmic droplets

Green tea polyphenols ameliorate non-alcoholic fatty liver disease through upregulating AMPK activation in high fat fed Zucker fatty rats.

AIMTo investigate protective effects and molecular mechanisms of green tea polyphenols (GTP) on non-alcoholic fatty liver disease (NAFLD) in Zucker fatty (ZF) rats.METHODSMale ZF rats were fed a high-fat diet (HFD) for 2 wk then treated with GTP (200 mg/kg) or saline (5 mL/kg) for 8 wk, with Zucker lean rat as their control. At the end of experiment, serum and liver tissue were collected for measurement of metabolic parameters, alanine aminotransferase (ALT) and aspartate aminotransferase (AST), inflammatory cytokines and hepatic triglyceride and liver histology. Immunoblotting was used to detect phosphorylation of AMP-activated protein kinase (AMPK) acetyl-CoA carboxylase (ACC), and sterol regulatory element-binding protein 1c (SREBP1c).RESULTSGenetically obese ZF rats on a HFD presented with metabolic features of hepatic pathological changes comparable to human with NAFLD. GTP intervention decreased weight gain (10.1%, P = 0.052) and significantly lowered visceral fat (31.0%, P < 0.01). Compared with ZF-controls, GTP treatment significantly reduced fasting serum insulin, glucose and lipids levels. Reduction in serum ALT and AST levels (both P < 0.01) were observed in GTP-treated ZF rats. GTP treatment also attenuated the elevated TNFα and IL-6 in the circulation. The increased hepatic TG accumulation and cytoplasmic lipid droplet were attenuated by GTP treatment, associated with significantly increased expression of AMPK-Thr172 (P < 0.05) and phosphorylated ACC and SREBP1c (both P < 0.05), indicating diminished hepatic lipogenesis and triglycerides out flux from liver in GTP treated rats.CONCLUSIONThe protective effects of GTP against HFD-induced NAFLD in genetically obese ZF rats are positively correlated to reduction in hepatic lipogenesis through upregulating the AMPK pathway.

Immunohistochemical staining of lipid droplets with adipophilin in paraffin-embedded glioma tissue identifies an association between lipid droplets and tumour grade

Abstract Background: Cytoplasmic lipid droplets are important in cancer metabolism and a clear relationship has been established between their accumulation and increased tumour grade in glioma. The development of the

Chronic exposure to ethanol causes steatosis and inflammation in zebrafish liver

AIMTo evaluate the effects of chronic exposure to ethanol in the liver and the expression of inflammatory genes in zebrafish.METHODSZebrafish (n = 104), wild type, adult, male and female, were divided into two groups: Control and ethanol (0.05 v/v). The ethanol was directly added into water; tanks water were changed every two days and the ethanol replaced. The animals were fed twice a day with fish food until satiety. After two and four weeks of trial, livers were dissected, histological analysis (hematoxilin-eosin and Oil Red staining) and gene expression assessment of adiponectin, adiponectin receptor 2 (adipor2), sirtuin-1 (sirt-1), tumor necrosis factor-alpha (tnf-a), interleukin-1b (il-1b) and interleukin-10 (il-10) were performed. Ultrastructural evaluations were conducted at fourth week.RESULTSExposing zebrafish to 0.5% ethanol developed intense liver steatosis after four weeks, as demonstrated by oil red staining. In ethanol-treated animals, the main ultrastructural changes were related to cytoplasmic lipid particles and droplets, increased number of rough endoplasmic reticulum cisterns and glycogen particles. Between two and four weeks, hepatic mRNA expression of il-1b, sirt-1 and adipor2 were upregulated, indicating that ethanol triggered signaling molecules which are key elements in both hepatic inflammatory and protective responses. Adiponectin was not detected in the liver of animals exposed and not exposed to ethanol, and il-10 did not show significant difference.CONCLUSIONData suggest that inflammatory signaling and ultrastructural alterations play a significant role during hepatic steatosis in zebrafish chronically exposed to ethanol.

Relationships between milks differentiated on native milk fat globule characteristics and fat, protein and calcium compositions

Many studies have shown that milk fat globule (MFG) diameter varies in dairy cows in relation to diet and/or breed. However, the mechanisms governing the size of the fat globules remain hypothetical. Our objective was to determine the variable biochemical characteristics (fat, protein, fatty acids (FA), casein and calcium (Ca) contents) between individual milk which differed in both MFG diameter and membrane content, in order to speculate about the links between milk synthesis and MFG secretion. With this aim, we built five databases of individual milk samples from 21 experiments performed between 2003 and 2011. Three of them grouped data from trials dealing with breed/diet effects and included information about: (i) MFG size/membrane, fat and protein contents (n=982), (ii) previous parameters plus FA profile (n=529) and (iii) previous parameters plus true protein composition and calcium contents (n=101). A hierarchical clustering analysis performed on these three databases yielded four groups differing in the MFG characteristics. We observed significant differences among groups for the following parameters: (i) fat content and fat : protein ratio; (ii) de novo and polyunsaturated FA contents; (iii) Ca contents. These relationships could result from potential process regulating the synthesis and secretion of MFG: (i) the apical membrane turnover for MFG secretion and (ii) cytoplasmic lipid droplet formation in the lactocyte during its migration from the basal to the apical pole. The two other databases grouped data from trials dealing with milking frequency (n=211), milking kinetics and milk type (residual v. cisternal) (n=224). They were used to study the relationships between the size of the MFG and milk composition for high native fat contents (from 60 up to 100 g/kg in residual milks). We observed curvilinear relationships between the size of the MFG and fat content, as well as with the fat : protein ratio. This result suggests that MFG diameter reaches a threshold but mechanisms are still unknown.

Fat Cell and Fatty Acid Turnover in Obesity.

The ratio of free fatty acid (FFA) turnover decreases significantly with the expansion of white adipose tissue. Adipose tissue and dietary saturated fatty acid levels significantly correlate with an increase in fat cell size and number. Inhibition of adipose triglyceride lipase leads to an accumulation of triglyceride, whereas inhibition of hormone-sensitive lipase leads to the accumulation of diacylglycerol. The G0/G1 switch gene 2 increases lipid content in adipocytes and promotes adipocyte hypertrophy through the restriction of triglyceride turnover. Excess triacylglycerols (TAGs), sterols and sterol esters are surrounded by the phospholipid monolayer surface and form lipid droplets. Following the release of lipid droplets from endoplasmic reticulum, cytoplasmic lipid droplets increase their volume either by local TAG synthesis or by homotypic fusion. The number and the size of lipid droplet distribution is correlated with obesity. Obesity-associated adipocyte death exhibits feature of necrosis-like programmed cell death. NOD-like receptors family pyrin domain containing 3 (NLRP3) inflammasome-dependent caspase-1 activation in hypertrophic adipocytes induces obese adipocyte death by pyroptosis. Actually adipocyte death may be a prerequisite for the transition from hypertrophic to hyperplastic obesity. Major transcriptional factors, CCAAT/enhancer-binding proteins beta and delta, play a central role in the subsequent induction of critical regulators, peroxisome-proliferator-activated receptor gamma, CCAAT/enhancer-binding protein alpha and sterol regulatory element-binding protein 1, in the transcriptional control of adipogenesis in obesity.Collectively, in this chapter the concept of adipose tissue remodeling in response to adipocyte death or adipogenesis, and the complexity of lipid droplet interactions with the other cellular organelles are reviewed. Furthermore, in addition to lipid droplet growth, the functional link between the adipocyte-specific lipid droplet-associated protein and fatty acid turn-over is also debated.

Efeito da imunocastração sobre o perímetro escrotal e a produção espermática em touros da raça nelore

Bopriva® exerts anti-gonadotropin-releasing hormone (GnRH) activity and is used for immunocastration of cattle. The objective of this study was to evaluate the effect of Bopriva® on scrotal circumference and sperm production and quality of Nellore bulls. Five Nellore bulls (Bos taurus indicus), aged 12 months, received two doses of Bopriva® at an interval of 30 days, with each dose of 1 mL/animal containing 400 μg of the product. At 2-weeks intervals, scrotal circumference was measured and semen was collected by electroejaculation (seven samplings) and submitted to physical and morphological analysis. Scrotal circumference significantly decreased after the 4th sampling (P=0.0056) and sperm motility ranged from 20% to 66% (mean of 39%), sperm vigor from 1.0 to 2.8 (mean of 1.74), and sperm concentration from 87.8 to 296.2 million spermatozoa per mL (mean of 242.9 million). The percentage of total sperm defects ranged from 36% to 75.6% (mean of 49.51%), with a high incidence of cytoplasmic droplets, bent tails and detached heads. Bopriva® reduces scrotal circumference and interferes with semen quality in Nellore animals, reducing sperm motility and increasing the percentage of sperm abnormalities in the ejaculate.