Perilipin 5 protects the mitochondrial oxidative functions and improves the alcoholic liver injury in mice.

BACKGROUND AND AIMS: Alcohol consumption is a well-established risk factor for the onset and progression of hepatic steatosis. Perilipin 5 (Plin5), a lipid droplet protein, is an important protective factor against hepatic lipotoxicity induced by excessive lipolysis, but its role and molecular mechanism in alcoholic liver disease (ALD) are not fully elucidated. METHODS: The optimized National Institute on Alcohol Abuse and Alcoholism model was used to construct ALD model mice. Automatic biochemical analyser was used for Biochemical Parameters. The primary hepatocytes and Plin5-overexpressed HepG2 cells (including full-length Plin5 and Plin5 deleting 444-464 aa) were used for in vitro experiment. Haematoxylin and Eosin staining, Oil Red O staining, Bodipy 493/503 staining, Periodic Acid-Schiff staining, immunohistochemistry and JC-1 staining were used to evaluate cell morphology, lipids, glycogen, inflammation and membrane potential. Commercially kits are used to detect glycolipid metabolites, such as triglycerides, glycogen, glucose, reactive oxygen species, lactic acids, ketone bodies. Fluorescently labelled deoxyglucose, NBDG, was used for glucose intake. An XF96 extracellular flux analyser was used to determinate oxygen consumption rate in hepatocytes. The morphological and structural damage of mitochondria was evaluated by electron microscopy. Classical ultracentrifugation is used to separate the subcellular organelles of tissues and cells. Immunoblotting and qPCR were used to detect changes in mRNA and protein levels of related genes. RESULTS: Our results showed that the expression of Plin5 in mouse livers was enhanced by alcohol intake, and Plin5 deficiency aggravated the alcohol-induced liver injury. To clarify the mechanism, we found that Plin5 deficiency significantly elevated the hepatic NADH levels and ketone body production in the alcohol-treated mice. As NADH elevation could promote the reduction of pyruvate into lactate and then inhibit the gluconeogenesis, alcohol-treated Plin5-deficient mice exhibited more lactate production and severer hypoglycemia. These results implied that Plin5 deficiency impaired the mitochondrial oxidative functions in the presence of alcohol. In addition, we demonstrated that Plin5 could be recruited onto mitochondria by alcohol, while Plin5 without mitochondrial targeting sequences lost its mitochondrial protection functions. CONCLUSION: Collectively, this study demonstrated that the mitochondrial Plin5 could protect the alcohol-induced mitochondrial injury, which provides an important new insight on the roles of Plin5 in highly oxidative tissues.

Perilipin 5 deficiency aggravates cardiac hypertrophy by stimulating lactate production in leptin-deficient mice.

BACKGROUND: Perilipin 5 (Plin5) is well known to maintain the stability of intracellular lipid droplets (LDs) and regulate fatty acid metabolism in oxidative tissues. It is highly expressed in the heart, but its roles have yet to be fully elucidated. METHODS: Plin5-deficient mice and Plin5/leptin-double-knockout mice were produced, and their histological structures and myocardial functions were observed. Critical proteins related to fatty acid and glucose metabolism were measured in heart tissues, neonatal mouse cardiomyocytes and Plin5-overexpressing H9C2 cells. 2-NBDG was employed to detect glucose uptake. The mitochondria and lipid contents were observed by MitoTracker and BODIPY 493/503 staining in neonatal mouse cardiomyocytes. RESULTS: Plin5 deficiency impaired glucose utilization and caused insulin resistance in mouse cardiomyocytes, particularly in the presence of fatty acids (FAs). Additionally, Plin5 deficiency increased the NADH content and elevated the expression of lactate dehydrogenase (LDHA) in cardiomyocytes, which resulted in increased lactate production. Moreover, when fatty acid oxidation was blocked by etomoxir or LDHA was inhibited by GSK2837808A in Plin5-deficient cardiomyocytes, glucose utilization was improved. Leptin-deficient mice exhibited myocardial hypertrophy, insulin resistance and altered substrate utilization, and Plin5 deficiency exacerbated myocardial hypertrophy in leptin-deficient mice. CONCLUSION: Our results demonstrated that Plin5 plays a critical role in coordinating fatty acid and glucose oxidation in cardiomyocytes, providing a potential target for the treatment of metabolic disorders in the heart.

Lipid-Independent Regulation of PLIN5 via IL-6 through the JAK/STAT3 Axis in Hep3B Cells.

Perilipin 5 (PLIN5) is a lipid droplet coat protein that is highly expressed in oxidative tissues such as those of muscles, the heart and the liver. PLIN5 expression is regulated by a family of peroxisome proliferator-activated receptors (PPARs) and modulated by the cellular lipid status. So far, research has focused on the role of PLIN5 in the context of non-alcoholic fatty liver disease (NAFLD) and specifically in lipid droplet formation and lipolysis, where PLIN5 serves as a regulator of lipid metabolism. In addition, there are only limited studies connecting PLIN5 to hepatocellular carcinoma (HCC), where PLIN5 expression is proven to be upregulated in hepatic tissue. Considering that HCC development is highly driven by cytokines present throughout NAFLD development and in the tumor microenvironment, we here explore the possible regulation of PLIN5 by cytokines known to be involved in HCC and NAFLD progression. We demonstrate that PLIN5 expression is strongly induced by interleukin-6 (IL-6) in a dose- and time-dependent manner in Hep3B cells. Moreover, IL-6-dependent PLIN5 upregulation is mediated by the JAK/STAT3 signaling pathway, which can be blocked by transforming growth factor-ß (TGF-ß) and tumor necrosis factor-α (TNF-α). Furthermore, IL-6-mediated PLIN5 upregulation changes when IL-6 trans-signaling is stimulated through the addition of soluble IL-6R. In sum, this study sheds light on lipid-independent regulation of PLIN5 expression in the liver, making PLIN5 a crucial target for NAFLD-induced HCC.

Leptin Reduces Plin5 m6A Methylation through FTO to Regulate Lipolysis in Piglets.

Perilipin5 (Plin5) is a scaffold protein that plays an important role in lipid droplets (LD) formation, but the regulatory effect of leptin on it is unclear. Our study aimed to explore the underlying mechanisms by which leptin reduces the N6-methyladenosine (m6A) methylation of Plin5 through fat mass and obesity associated genes (FTO) and regulates the lipolysis. To this end, 24 Landrace male piglets (7.73 ± 0.38 kg) were randomly sorted into two groups, either a control group (Control, n = 12) or a 1 mg/kg leptin recombinant protein treatment group (Leptin, n = 12). After 4 weeks of treatment, the results showed that leptin treatment group had lower body weight, body fat percentage and blood lipid levels, but the levels of Plin5 mRNA and protein increased significantly in adipose tissue (p < 0.05). Leptin promotes the up-regulation of FTO expression level in vitro, which in turn leads to the decrease of Plin5 M6A methylation (p < 0.05). In in vitro porcine adipocytes, overexpression of FTO aggravated the decrease of M6A methylation and increased the expression of Plin5 protein, while the interference fragment of FTO reversed the decrease of m6A methylation (p < 0.05). Finally, the overexpression in vitro of Plin5 significantly reduces the size of LD, promotes the metabolism of triglycerides and the operation of the mitochondrial respiratory chain, and increases thermogenesis. This study clarified that leptin can regulate Plin5 M6A methylation by promoting FTO to affect the lipid metabolism and energy consumption, providing a theoretical basis for treating diseases related to obesity.

Perilipin 5 is a novel target of nuclear receptor LRH-1 to regulate hepatic triglycerides metabolism.

Liver receptor homolog-1 (LRH-1) has emerged as a regulator of hepatic glucose, bile acid, and mitochondrial metabolism. However, the functional mechanism underlying the effect of LRH-1 on lipid mobilization has not been addressed. This study investigated the regulatory function of LRH-1 in lipid metabolism in maintaining a normal liver physiological state during fasting. The Lrh-1f/f and LRH-1 liver-specific knockout (Lrh-1LKO) mice were either fed or fasted for 24 h, and the liver and serum were isolated. The livers were used for qPCR, western blot, and histological analysis. Primary hepatocytes were isolated for immunocytochemistry assessments of lipids. During fasting, the Lrh-1LKO mice showed increased accumulation of triglycerides in the liver compared to that in Lrh-1f/f mice. Interestingly, in the Lrh-1LKO liver, decreases in perilipin 5 (PLIN5) expression and genes involved in ß-oxidation were observed. In addition, the LRH-1 agonist dialauroylphosphatidylcholine also enhanced PLIN5 expression in human cultured HepG2 cells. To identify new target genes of LRH-1, these findings directed us to analyze the Plin5 promoter sequence, which revealed -1620/-1614 to be a putative binding site for LRH-1. This was confirmed by promoter activity and chromatin immunoprecipitation assays. Additionally, fasted Lrh-1f/f primary hepatocytes showed increased co-localization of PLIN5 in lipid droplets (LDs) compared to that in fasted Lrh-1LKO primary hepatocytes. Overall, these findings suggest that PLIN5 might be a novel target of LRH-1 to mobilize LDs, protect the liver from lipid overload, and manage the cellular needs during fasting. [BMB Reports 2021; 54(9): 476-481].

Perilipin 5 links mitochondrial uncoupled respiration in brown fat to healthy white fat remodeling and systemic glucose tolerance.

Exposure of mice or humans to cold promotes significant changes in brown adipose tissue (BAT) with respect to histology, lipid content, gene expression, and mitochondrial mass and function. Herein we report that the lipid droplet coat protein Perilipin 5 (PLIN5) increases markedly in BAT during exposure of mice to cold. To understand the functional significance of cold-induced PLIN5, we created and characterized gain- and loss-of-function mouse models. Enforcing PLIN5 expression in mouse BAT mimics the effects of cold with respect to mitochondrial cristae packing and uncoupled substrate-driven respiration. PLIN5 is necessary for the maintenance of mitochondrial cristae structure and respiratory function during cold stress. We further show that promoting PLIN5 function in BAT is associated with healthy remodeling of subcutaneous white adipose tissue and improvements in systemic glucose tolerance and diet-induced hepatic steatosis. These observations will inform future strategies that seek to exploit thermogenic adipose tissue as a therapeutic target for type 2 diabetes, obesity, and nonalcoholic fatty liver disease.

Isolated Plin5-deficient cardiomyocytes store less lipid droplets than normal, but without increased sensitivity to hypoxia.

Plin5 is abundantly expressed in the heart where it binds to lipid droplets (LDs) and facilitates physical interaction between LDs and mitochondria. We isolated cardiomyocytes from adult Plin5+/+ and Plin5-/- mice to study the role of Plin5 for fatty acid uptake, LD accumulation, fatty acid oxidation, and tolerance to hypoxia. Cardiomyocytes isolated from Plin5-/- mice cultured with oleic acid stored less LDs than Plin5+/+, but comparable levels to Plin5+/+ cardiomyocytes when adipose triglyceride lipase activity was inhibited. The ability to oxidize fatty acids into CO2 was similar between Plin5+/+ and Plin5-/- cardiomyocytes, but Plin5-/- cardiomyocytes had a transient increase in intracellular fatty acid oxidation intermediates. After pre-incubation with oleic acids, Plin5-/- cardiomyocytes retained a higher content of glycogen and showed improved tolerance to hypoxia compared to Plin5+/+. In isolated, perfused hearts, deletion of Plin5 had no important effect on ventricular pressures or infarct size after ischemia. Old Plin5-/- mice had reduced levels of cardiac triacylglycerides, increased heart weight, and apart from modest elevated expression of mRNAs for beta myosin heavy chain Myh7 and the fatty acid transporter Cd36, other genes involved in fatty acid oxidation, glycogen metabolism and glucose utilization were essentially unchanged by removal of Plin5. Plin5 seems to facilitate cardiac LD storage primarily by repressing adipose triglyceride lipase activity without altering cardiac fatty acid oxidation capacity. Expression of Plin5 and cardiac LD content of isolated cardiomyocytes has little importance for tolerance to acute hypoxia and ischemia, which contrasts the protective role for Plin5 in mouse models during myocardial ischemia.

Identification of prognostic and metastasis-related alternative splicing signatures in hepatocellular carcinoma.

As the most common neoplasm in digestive system, hepatocellular carcinoma (HCC) is one of the most important leading cause of cancer deaths worldwide. Its high-frequency metastasis and relapse rate lead to the poor survival of HCC patients. However, the mechanism of HCC metastasis is still unclear. Alternative splicing events (ASEs) have a great effect in cancer development, progression and metastasis. We downloaded RNA sequencing and seven types of ASEs data of HCC samples, in order to explore the mechanism of ASEs underlying tumorigenesis and metastasis of HCC. The data were taken from the The Cancer Genome Atlas (TCGA) and TCGASpliceSeq databases. Univariate Cox regression analysis was used to determine a total of 3197 overall survival-related ASEs (OS-SEs). And based on five OS-SEs screened by Lasso regression, we constructed a prediction model with the Area Under Curve of 0.765. With a good reliability of the model, the risk score was also proved to be an independent predictor. Among identified 390 candidate SFs, Y-box protein 3 (YBX3) was significantly correlated with OS and metastasis. Among 177 ASEs, ATP-binding cassette subfamily A member 6 (ABCA6)-43162-AT and PLIN5-46808-AT were identified both associated with OS, bone metastasis and co-expressed with SFs. Then we identified primary bile acid biosynthesis as survival-related (KEGG) pathway by Gene Set Variation Analysis (GSVA) and univariate regression analysis, which was correlated with ABCA6-43162-AT and PLIN5-46808-AT. Finally, we proposed that ABCA6-43162-AT and PLIN5-46808-AT may contribute to HCC poor prognosis and metastasis under the regulation of aberrant YBX3 through the pathway of primary bile acid biosynthesis.

Skeletal muscle lipid droplets are resynthesized before being coated with perilipin proteins following prolonged exercise in elite male triathletes.

Intramuscular triglycerides (IMTG) are a key substrate during prolonged exercise, but little is known about the rate of IMTG resynthesis in the postexercise period. We investigated the hypothesis that the distribution of the lipid droplet (LD)-associated perilipin (PLIN) proteins is linked to IMTG storage following exercise. Fourteen elite male triathletes (27 ± 1 yr, 66.5 ± 1.3 mL·kg-1·min-1) completed 4 h of moderate-intensity cycling. During the first 4 h of recovery, subjects received either carbohydrate or H2O, after which both groups received carbohydrate. Muscle biopsies collected pre- and postexercise and 4 and 24 h postexercise were analyzed using confocal immunofluorescence microscopy for fiber type-specific IMTG content and PLIN distribution with LDs. Exercise reduced IMTG content in type I fibers (-53%, P = 0.002), with no change in type IIa fibers. During the first 4 h of recovery, IMTG content increased in type I fibers (P = 0.014), but was not increased more after 24 h, where it was similar to baseline levels in both conditions. During recovery the number of LDs labeled with PLIN2 (70%), PLIN3 (63%), and PLIN5 (62%; all P < 0.05) all increased in type I fibers. Importantly, the increase in LDs labeled with PLIN proteins only occurred at 24 h postexercise. In conclusion, IMTG resynthesis occurs rapidly in type I fibers following prolonged exercise in highly trained individuals. Furthermore, increases in IMTG content following exercise preceded an increase in the number of LDs labeled with PLIN proteins. These data, therefore, suggest that the PLIN proteins do not play a key role in postexercise IMTG resynthesis.

Curcumin Recovers Intracellular Lipid Droplet Formation Through Increasing Perilipin 5 Gene Expression in Activated Hepatic Stellate Cells In Vitro.

The activation of hepatic stellate cells (HSCs) is a major event during hepatic fibrogenesis. Restoration of intracellular lipid droplet (LD) formation turns the activated HSC back to a quiescent state. Our previous studies have shown that curcumin suppresses HSC activation through increasing peroxisome proliferator-activated receptor, gamma (PPARγ) and 5' adenosine monophosphate-activated protein kinase (AMPK) activities. This study aims at evaluating the effect of curcumin on lipid accumulation in HSCs and hepatocytes, and further elucidating the underlying mechanisms. Now we showed that curcumin increased LD formation in activated HSCs and stimulated the expression of sterol regulatory element-binding protein and fatty acid synthase, and reduced the expression of adipose triglyceride lipase. Exogenous perilin5 expression in primary HSCs promoted LD formation. Perilipin 5 siRNA eliminated curcumin-induced LD formation in HSCs. These results suggest that curcumin recovers LD formation and lipid accumulation in activated HSCs by increasing perilipin 5 gene expression. Furthermore, inhibition of AMPK or PPARγ activity blocked curcumin's effect on Plin5 gene expression and LD formation. Our results provide a novel evidence in vitro for curcumin as a safe, effective candidate to treat liver fibrosis.

Excision of the expanded GAA repeats corrects cardiomyopathy phenotypes of iPSC-derived Friedreich's ataxia cardiomyocytes.

Friedreich's ataxia is caused by large homozygous, intronic expansions of GAA repeats in the frataxin (FXN) gene, resulting in severe downregulation of its expression. Pathogenic repeats are located in intron one, hence patients express unaffected FXN protein, albeit in low quantities. Although FRDA symptoms typically afflict the nervous system, hypertrophic cardiomyopathy is the predominant cause of death. Our studies were conducted using cardiomyocytes differentiated from induced pluripotent stem cells derived from control individuals, FRDA patients, and isogenic cells corrected by zinc finger nucleases-mediated excision of pathogenic expanded GAA repeats. This correction of the FXN gene removed the primary trigger of the transcription defect, upregulated frataxin expression, reduced pathological lipid accumulation observed in patient cardiomyocytes, and reversed gene expression signatures of FRDA cardiomyocytes. Transcriptome analyses revealed hypertrophy-specific expression signatures unique to FRDA cardiomyocytes, and emphasized similarities between unaffected and ZFN-corrected FRDA cardiomyocytes. Thus, the iPSC-derived FRDA cardiomyocytes exhibit various molecular defects characteristic for cellular models of cardiomyopathy that can be corrected by genome editing of the expanded GAA repeats. These results underscore the utility of genome editing in generating isogenic cellular models of FRDA and the potential of this approach as a future therapy for this disease.

Plin5 deficiency exacerbates pressure overload-induced cardiac hypertrophy and heart failure by enhancing myocardial fatty acid oxidation and oxidative stress.

While cardiac hypertrophy and heart failure are accompanied by significant alterations in energy metabolism, more than 50-70% of energy is obtained from fatty acid ß-oxidation (FAO) in adult hearts under physiological conditions. Plin5 is involved in the metabolism of lipid droplets (LDs) and is highly abundant in oxidative tissues including heart, liver and skeletal muscle. Plin5 protects the storage of triglyceride (TG) in LDs by inhibiting lipolysis, thereby suppressing excess FAO and preventing excessive oxidative stress in the heart. In this study, we investigated the roles of Plin5 in cardiac hypertrophy and heart failure in mice treated with transverse aortic constriction (TAC). The results indicated that Plin5 deficiency aggravated myocardial hypertrophy in the TAC-treated mice and exacerbated the TAC-induced heart failure. We also found that Plin5 deficiency reduced the cardiac lipid accumulation and upregulated the levels of PPARα and PGC-1α, which stimulate mitochondrial proliferation. Moreover, Plin5 deficiency aggravated the TAC-induced oxidative stress. We consistently found that Plin5 knockdown disrupted TG storage and elevated FAO and lipolysis in H9C2 rat cardiomyocytes. In addition, Plin5 knockdown also provoked mitochondrial proliferation and lipotoxic injury in H9C2 cells. In conclusion, Plin5 deficiency increases myocardial lipolysis, elevates FAO and oxidative burden, and thereby exacerbates cardiac hypertrophy and heart failure in TAC-treated mice.

Oleic acid-induced perilipin 5 expression and lipid droplets formation are regulated by the PI3K/PPARα pathway in HepG2 cells.

Perilipin 5 (Plin5), a member of the PAT (Perilipin, ADRP, and Tip47) protein family, has been implicated in the regulation of cellular neutral lipid accumulation in nonalcoholic fatty liver diseases. However, the underlying regulatory mechanisms of Plin5 are not clear. The goal of the present study was to explore the mechanism of oleic acid (OA)-induced Plin5 expression in HepG2 cells. We found that the expression of Plin5 was increased during OA-induced lipid droplets formation in a dose- and time-dependent manner. During this process of OA-stimulated lipid droplets formation, peroxisome proliferator-activated receptor alpha (PPARα) was also upregulated. When PPARα activation was blocked by GW6471, OA-induced Plin5 expression and lipid droplets formation were effectively ablated. We further found that the phosphoinositide 3-kinase (PI3K) inhibitor LY294002 was able to downregulate both PPARα and Plin5 expression and lipid droplets formation. Thus, we concluded that PI3K may, at least in part, act upstream of PPARα to regulate Plin5 expression and lipid droplets formation in HepG2 cells.

Lipid storage droplet protein 5 reduces sodium palmitate­induced lipotoxicity in human normal liver cells by regulating lipid metabolism­related factors.

Lipid storage droplet protein 5 (LSDP5) is specifically expressed in tissues with high oxidative metabolism such as liver and heart. The present study aimed to explored the role of LSDP5 in sodium palmitate­induced lipotoxicity in LO2 normal human liver cells. LO2 cells were treated with various concentrations of sodium palmitate (25, 50, 75, 100, 125 and 150 µmol/l) for 12, 24 and 48 h, and cell viability was determined by Cell Counting Kit­8. Subsequently, LO2 cells were exposed to 100 µmol/l sodium palmitate for 48 h to induce lipotoxicity (Model). Lipotoxicity Model LO2 cells were also transfected with pCMV5­LSDP5 overexpression vector, and reactive oxygen species (ROS) production, mitochondrial membrane potential (MMP) and apoptotic rates were measured. The contents of non­esterified fatty acid (NEFA), malondialdehyde (MDA) and superoxide dismutase (SOD) were also measured. The expression levels of LSDP5, and apoptosis­, mitochondrial­, lipid metabolism­related factors were detected using reverse transcription­quantitative polymerase chain reaction and western blot assays. The results indicated that sodium palmitate exposure inhibited cell viability and induced lipotoxicity in LO2 cells. LSDP5 overexpression decreased ROS and apoptotic rates, and reduced NEFA and MDA content. LSDP5 transfection rescued the loss of MMP and elevated SOD content in lipotoxicity Model LO2 cells. In addition, LSDP5 upregulated the expression levels of B­cell lymphoma­2, acetyl­CoA carboxylase1/2 and fatty acid synthase (Fas), whereas the expression levels of activated­caspase­3, Bcl­2­associated X protein, cytochrome c, cytochrome c oxidase subunits IV, carnitine palmitoyltransferase 1a and peroxisome proliferator­activated receptors α levels were downregulated. LSDP5 may produce a protective effect on sodium palmitate­induced lipotoxicity in LO2 cells by regulating lipid metabolism­related factors.

Perilipin 5 alleviates HCV NS5A-induced lipotoxic injuries in liver.

BACKGROUND: The homeostasis of lipid droplets (LDs) plays a crucial role in maintaining the physical metabolic processes in cells, and is regulated by many LD-associated proteins, including perilipin 5 (Plin5) in liver. As the putative sites of hepatitis C virus (HCV) virion assembly, LDs are vital to viral infection. In addition, the hepatic LD metabolism can be disturbed by non-structural HCV proteins, such as NS5A, but the details are still inexplicit. METHODS: HCV NS5A was overexpressed in the livers and hepatocytes of wild-type and Plin5-null mice. BODIPY 493/503 and oil red O staining were used to detect the lipid content in mouse livers and hepatocytes. The levels of lipids, lipid peroxidation and inflammation biomarkers were further determined. Immunofluorescence assay and co-immunoprecipitation assay were performed to investigate the relationship of Plin5 and NS5A. RESULTS: One week after adenovirus injection, livers expressing NS5A showed more inflammatory cell aggregation and more severe hepatic injuries in Plin5-null mice than in control mice, which was consistent with the increased serum levels of IL-2 and TNF-α (P < 0.05) observed in Plin5-null mice. Moreover, Plin5 deficiency in the liver and hepatocytes aggravated the elevation of MDA and 4-HNE levels induced by NS5A expression (P < 0.01). The triglyceride (TG) content was increased approximately 25% by NS5A expression in the wild-type liver and hepatocytes but was unchanged in the Plin5-null liver and hepatocytes. More importantly, Plin5 deficiency in the liver and hepatocytes exacerbated the elevation of non-esterified fatty acids (NEFAs) stimulated by NS5A expression (P < 0.05 and 0.01 respectively). Using triacsin C to block acyl-CoA biosynthesis, we found that Plin5 deficiency aggravated the NS5A-induced lipolysis of TG. In contrast, Plin5 overexpression in HepG2 cells ameliorated the NS5A-induced lipolysis and lipotoxic injuries. Immunofluorescent staining demonstrated that NS5A expression stimulated the targeting of Plin5 to the surface of the LDs in hepatocytes without altering the protein levels of Plin5. By co-IP, we found that the N-terminal domain (aa 32-128) of Plin5 was pivotal for its binding with NS5A. CONCLUSIONS: Our data highlight a protective role of Plin5 against hepatic lipotoxic injuries induced by HCV NS5A, which is helpful for understanding the steatosis and injuries in liver during HCV infection.

Perilipin 5 Deletion in Hepatocytes Remodels Lipid Metabolism and Causes Hepatic Insulin Resistance in Mice.

Defects in hepatic lipid metabolism cause nonalcoholic fatty liver disease and insulin resistance, and these pathologies are closely linked. Regulation of lipid droplet metabolism is central to the control of intracellular fatty acid fluxes, and perilipin 5 (PLIN5) is important in this process. We examined the role of PLIN5 on hepatic lipid metabolism and systemic glycemic control using liver-specific Plin5-deficient mice (Plin5LKO ). Hepatocytes isolated from Plin5LKO mice exhibited marked changes in lipid metabolism characterized by decreased fatty acid uptake and storage, decreased fatty acid oxidation that was associated with reduced contact between lipid droplets and mitochondria, and reduced triglyceride secretion. With consumption of a high-fat diet, Plin5LKO mice accumulated intrahepatic triglyceride, without significant changes in inflammation, ceramide or diglyceride contents, endoplasmic reticulum stress, or autophagy. Instead, livers of Plin5LKO mice exhibited activation of c-Jun N-terminal kinase, impaired insulin signal transduction, and insulin resistance, which impaired systemic insulin action and glycemic control. Re-expression of Plin5 in the livers of Plin5LKO mice reversed these effects. Together, we show that Plin5 is an important modulator of intrahepatic lipid metabolism and suggest that the increased Plin5 expression that occurs with overnutrition may play an important role in preventing hepatic insulin resistance.

Perilipin 5 promotes hepatic steatosis in dairy cows through increasing lipid synthesis and decreasing very low density lipoprotein assembly.

Fatty liver is a common metabolic disorder in dairy cows during the transition period. Perilipin 5 (PLIN5), a lipid droplet coat protein, plays important roles in the development of hepatic steatosis in mice and humans. Whether PLIN5 plays a role in the development of fatty liver in dairy cows is unknown. An in vivo study consisting of 10 healthy and 10 cows with fatty liver was performed to harvest liver tissue and blood samples. In addition, hepatocytes isolated from calves were infected with PLIN5 overexpression adenovirus for 48 h; treated with 0, 0.6, 1.2, or 2.4 mM nonesterified fatty acids (NEFA) for 24 h; or infected with PLIN5 silencing adenovirus for 48 h and then treated with 1.2 mM NEFA for 24 h. Serum concentrations of NEFA and ß-hydroxybutyrate were greater in cows with fatty liver. Milk production and plasma glucose concentrations were lower in cows with fatty liver. The results revealed that PLIN5 is highly expressed in steatotic liver and localized to lipid droplets. The abundance of fatty acid and triacylglycerol (TAG) synthesis-related proteins including sterol regulatory element binding protein-1c, fatty acid synthase, acetyl-coA carboxylase 1, diacylglycerol acyltransferase 1, and diacylglycerol acyltransferase 2 was greater in the liver of cows with fatty liver. In contrast, the abundance of microsomal triglyceride transfer protein (MTP), apolipoprotein B100, and apolipoprotein E was lower in the liver of cows with fatty liver. Consequently, cows with fatty liver exhibited severe hepatic TAG accumulation and lower blood concentration of very low density lipoprotein apolipoprotein B (VLDL-ApoB). Overexpression of PLIN5 and exogenous NEFA in cultured hepatocytes increased the abundance of sterol regulatory element binding protein-1, fatty acid synthase, acetyl-coA carboxylase 1, diacylglycerol acyltransferase 1, and diacylglycerol acyltransferase 2 but decreased the abundance of microsomal triglyceride transfer protein, apolipoprotein B100, and apolipoprotein E, which promoted TAG synthesis and inhibited VLDL-ApoB assembly, inducing lipid accumulation. Importantly, knockdown of PLIN5 attenuated the upregulation of TAG synthesis and downregulation of VLDL-ApoB assembly induced by NEFA. Overall, these data suggest that NEFA activate PLIN5, leading to TAG accumulation and inhibition of VLDL assembly. As such, these mechanisms explain in part the development of hepatic steatosis in dairy cows.

The Effects of Resveratrol, Metformin, Cold and Strength Training on the Level of Perilipin 5 in the Heart, Skeletal Muscle and Brown Adipose Tissues in Mouse.

The high accumulation of lipid droplets in the cell is related to metabolic disorders, such as obesity. Perilipin 5 (Plin5), plays an important role in triglyceride hydrolysis in the lipid droplets. In this study, this protein has been evaluated in different tissues and conditions in mice. Fifty male mice were divided into 5 groups and treated for 45 days with Resveratrol, Metformin, strength training, and 4 °C cold. Brown adipose tissue (BAT), gastrocnemius skeletal muscle and heart were isolated for RNA extraction. The Plin5 gene expression was evaluated, using Real-Time PCR, and the plin5 was analyzed at the protein level, using western blot. In BAT, Resveratrol significantly reduced the plin5 protein level and gene expression (p < 0.05). In heart tissue, Resveratrol and strength training, decreased (p < 0.05) the plin5 expression, but Metformin increased the gene expression (p < 0.05). In skeletal muscle, resveratrol, strength training, cold and Metformin significantly increased the plin5 expression at the gene and protein level (p < 0.05). In BAT, Resveratrol has a greater effect in decreasing lipid deposits, compared with the strength training and cold; thus, it can play a better role in preventing lipid accumulation. In heart tissue, Resveratrol probably decreases insulin resistance, due to the increased expression of plin5 in skeletal muscle.

Perilipin 5 and liver fatty acid binding protein function to restore quiescence in mouse hepatic stellate cells.

Hepatic stellate cell (HSC) activation occurs along with decreased Perilipin5 (Plin5) and liver fatty acid-binding protein (L-Fabp) expression and coincident lipid droplet (LD) depletion. Conversely, the activated phenotype is reversible in WT HSCs upon forced expression of Plin5. Here, we asked if L-Fabp expression is required for Plin5-mediated rescue of the quiescent phenotype. Lentiviral Plin5 transduction of passaged L-Fabp-/- HSCs failed to reverse activation markers or restore lipogenic gene expression and LD formation. However, adenoviral L-Fabp infection of lentiviral Plin5 transduced L-Fabp-/- HSCs restored both the quiescent phenotype and LD formation, an effect also mediated by adenoviral intestine-Fabp or adipocyte-Fabp. Expression of exogenous Plin5 in activated WT HSCs induced a transcriptional program of lipogenic gene expression including endogenous L-Fabp, but none of the other FABPs. We further demonstrated that selective, small molecule inhibition of endogenous L-Fabp also eliminated the ability of exogenous Plin5 to rescue LD formation and reverse activation of WT HSCs. This functional coordination of L-Fabp with Plin5 was 5'-AMP-activated protein kinase (AMPK)-dependent and was eliminated by AMPK inhibition. Taken together, our results indicate that L-Fabp is required for Plin5 to activate a transcriptional program that restores LD formation and reverses HSC activation.

Plin5 alleviates myocardial ischaemia/reperfusion injury by reducing oxidative stress through inhibiting the lipolysis of lipid droplets.

Myocardial ischaemia-reperfusion (I/R) injury is a complex pathophysiological process. Current research has suggested that energy metabolism disorders, of which the abnormal consumption of fatty acids is closely related, compose the main pathological basis for myocardial I/R injury. Lipid droplets (LD) are critical regulators of lipid metabolism by LD-associated proteins. Among the lipid droplet proteins, the perilipin family members regulate lipolysis and lipogenesis through different mechanisms. Plin5, an important perilipin protein, promotes LD generation and lowers fatty acid oxidation, thus protecting the myocardium from lipotoxicity. This study investigated the protective effects of Plin5 in I/R myocardium. Our results indicated that Plin5 deficiency exacerbated the myocardial infarct area, aggravated left ventricular systolic dysfunction, reduced lipid storage, and elevated free fatty acids. Plin5-deficient myocardium exhibited severely damaged mitochondria, elevated reactive oxygen species (ROS) and malondialdehyde (MDA) levels, and decreased superoxide dismutase (SOD) activity. Furthermore, the decreased phosphorylation of PI3K/Akt in Plin5-null cardiomyocytes might contribute to I/R injury aggravation. In conclusion, Plin5, a new regulator of myocardial lipid metabolism, decreases free fatty acid peroxidation by inhibiting the lipolysis of intracellular lipid droplets, thus providing cardioprotection against I/R injury and shedding new light on therapeutic solutions for I/R diseases.