Promoted Lipid Accumulation Research Articles

Trimethylamine N-oxide promotes oxidative stress and lipid accumulation in macrophage foam cells via the Nrf2/ABCA1 pathway.

Recently, trimethylamine N-oxide (TMAO) has been considered a risk factor for cardiovascular disease and has a proatherogenic effect. Many studies have found that TMAO is involved in plaque oxidative stress and lipid metabolism, but the specific mechanism is still unclear. In our study, meta-analysis and bioinformatic analysis were firstly conducted in the database, and found that the effect of high plasma TMAO levels on promoting atherosclerotic plaque may be related to the expression of key antioxidant genes nuclear factor erytheroid-derived-2-like 2 (NFE2L2/Nrf2) decreased. Next, we assessed the role of Nrf2-mediated signaling pathway in TMAO-treated foam cells. Our results showed that TMAO can inhibit the expression of Nrf2 and its downstream antioxidant response element such as heme oxygenase-1 (HO-1) and glutathione peroxidase4 (GPX4), resulting in increased production of reactive oxygen species and decreased activity of superoxide dismutase, promoting oxidative stress. And TMAO can also promote lipid accumulation in foam cells by inhibiting cholesterol efflux protein expression. In addition, upregulation of Nrf2 expression partially rescues TMAO-induced oxidative stress and reduces ATP-binding cassette A1 (ABCA1)-mediated lipid accumulation. Therefore, TMAO promotes oxidative stress and lipid accumulation in macrophage foam cells through the Nrf2/ABCA1 pathway, which may provide a potential mechanism for the proatherogenic effect of TMAO.

Metabolic interpretation of NaCl stress-induced lipid accumulation in microalgae for promising biodiesel production with saline wastewater

The rising use of saline wastewater for biodiesel production through oleaginous microalgae demands fundamental studies on the metabolic response of microalgal cells to salt stress. Thus, we investigated how salt stress promotes lipid accumulation inside Botryococcus sp. NJD-1 by tracking critical metabolites and proteases as well as analyzing the cellular redox balance. The results showed that the lipid content of microalgal cells followed a negative binomial correlation with intracellular reactive oxygen species (ROS), while the neutral lipid content increased linearly with salinity in range of 0–20 g/L NaCl. The optimal lipid production was achieved as 54.5 % and 110.5 mg/L d-1 at 10 g/L NaCl. Further comparing the metabolomics and proteomics data at 0 and 10 g/L NaCl, the adenosine triphosphate (ATP) and nicotinamide adenine nucleotide phosphate (NADPH) were found significantly increased to quench the Na+-induced ROS. The created NADPH and ATP accord the enhanced activity of the upper Pyruvate-Malate cycle and tricarboxylic acid cycle (TCA) cycle. Therefore, ROS detoxification is proposed to be the essential mechanism for microalgal lipid accumulation under salt stress. The cellular triacylglycerol (TAG) synthesis triggered by salinity is mainly mediated by an acetyl-CoA-dependent Kennedy pathway. These findings will deepen our understanding of the nexus of ambient salt, intracellular ROS and lipid metabolism for microalgae and advance the exploit of saline wastewater for microalgal lipid production.

Desialylation of ATG5 by sialidase (NEU1) promotes macrophages autophagy and exacerbates inflammation under hypoxia

During the process of atherosclerosis (AS), hypoxia induces plaque macrophage inflammation, promoting lipid accumulation. Autophagy is a cell homeostasis process that increases tolerance to stressors like oxidative stress and hypoxia. However, the specific mechanism by which hypoxia initiates autophagy and the inflammation of macrophages remains to be elucidated. Here, we found that hypoxia-induced macrophage inflammation was mediated by autophagy. Then, the effect of hypoxia on autophagy was investigated in terms of post-translational modifications of proteins. The results showed that desialylation of the autophagy protein ATG5 under hypoxic conditions enhanced protein stability by affecting its charge effect and promoted the formation of the ATG5-ATG12-ATG16L complex, further increasing autophagosome formation. And NEU1, a key enzyme in sialic acid metabolism, was significantly up-regulated under hypoxic conditions and was identified as an interacting protein of ATG5, affecting the sialylation of ATG5. In addition, the knockdown or inhibition of NEU1 reversed hypoxia-induced autophagy and inflammatory responses. In conclusion, our data reveal a key mechanism of autophagy regulation under hypoxia involving ATG5 sialylation and NEU1, suggesting that NEU1 may be a potential target for the prevention and treatment of atherosclerosis.

Stearoyl-CoA desaturase 1 inhibition impairs triacylglycerol accumulation and lipid droplet formation in colorectal cancer cells.

Increases in fatty acid (FA) biosynthesis meet the higher lipid demand by intensely proliferating cancer cells and promoting their progression. Stearoyl-CoA desaturase 1 (SCD1) is the key enzyme in FA biosynthesis, converting saturated FA (SFA) into monounsaturated FA (MUFA). Increases in the MUFA/SFA ratio and SCD1 expression have been observed in cancers of various origins and correlate with their aggressiveness. However, much is still unknown about the SCD1-dependent molecular mechanisms that promote specific changes in metabolic pathways of cancer cells. The present study investigated the involvement of SCD1 in shaping glucose and lipid metabolism in colorectal cancer (CRC) cells. Excess FAs that derive from de novo lipogenesis are stored in organelles, called lipid droplets (LDs), mainly in the form of triacylglycerol (TAG) and cholesteryl esters. LD accumulation is associated with key features of cancer development and progression. Consistent with our findings, the pharmacological inhibition of SCD1 activity affects CRC cell viability and impairs TAG accumulation and LD formation in these cells through the activation of lipolytic and lipophagic pathways. We showed that SCD1 suppression affects crucial lipogenic processes that promote lipid accumulation in CRC cells but in a sterol regulatory element-binding protein 1-independent manner. We propose that adenosine monophosphate-activated protein kinase contributes to these changes through the activation of lipolysis and inhibition of TAG synthesis. We also provide evidence of the involvement of SCD1 in the regulation of glucose uptake and utilization in CRC cells. These findings underscore the importance of SCD1 in regulating cellular processes that promote cancer development and progression.

FBXL5 promotes lipid accumulation in alcoholic fatty liver disease by promoting the ubiquitination and degradation of TFEB

BackgroundAlcoholic fatty liver disease (AFLD) is characterized by abnormal lipid droplet accumulation in liver. Epigenetic regulation plays an important role in the pathogenesis of AFLD. Comprehensive bioinformatics analysis revealed that an E3 ubiquitin ligase, F-box and leucine-rich repeats protein 5 (FBXL5), was significantly upregulated in AFLD mice. MethodsThe mouse model of AFLD was established by feeding Lieber-DeCarli liquid diet containing ethanol. An in vitro model of AFLD was established by treating HepG2 cells with ethanol (EtOH). The FBXL5 expression was assessed by quantitative real-time PCR (qRT-PCR) and western blotting assays. The levels of triglyceride (TG), alanine aminotransferase (ALT), aspartate aminotransferase (AST), and lipid accumulation were analyzed by enzyme-linked immunosorbent assay (ELISA) and Nile red staining. ResultsThe FBXL5 expression was markedly up-regulated in in vivo and in vitro models of AFLD compared with controls. Functionally, FBXL5 knockdown alleviated lipid accumulation in EtOH-treated HepG2 cells. Mechanistically, FBXL5 directly interacted with transcription factor EB (TFEB) and accelerated its ubiquitination-mediated degradation. TFEB knockdown reversed the effect of FBXL5 inhibition on decreasing EtOH-induced lipid accumulation. ConclusionOur data suggest that FBXL5 promotes lipid accumulation in AFLD by promoting the ubiquitination and degradation of TFEB.

Ablation of Dual-Specificity Phosphatase 6 Protects against Nonalcoholic Fatty Liver Disease via Cytochrome P450 4A and Mitogen-Activated Protein Kinase

Dual-specificity phosphatase 6 (DUSP6) is a specific phosphatase for mitogen-activated protein kinase (MAPK). This study used a high-fat diet (HFD)-induced murine nonalcoholic fatty liver disease model to investigate the role of DUSP6 in this disease. Wild-type (WT) and Dusp6-haploinsufficiency mice developed severe obesity and liver pathology consistent with nonalcoholic fatty liver disease when exposed to HFD. In contrast, Dusp6-knockout (KO) mice completely eliminated these phenotypes. Furthermore, primary hepatocytes isolated from WT mice exposed to palmitic and oleic acids exhibited abundant intracellular lipid accumulation, whereas hepatocytes from Dusp6-KO mice showed minimal lipid accumulation. Transcriptome analysis revealed significant down-regulation of genes encoding cytochrome P450 4A (CYP4A), known to promote ω-hydroxylation of fatty acids and hepatic steatosis, in Dusp6-KO hepatocytes compared with that in WT hepatocytes. Diminished CYP4A expression was observed in the liver of Dusp6-KO mice compared with WT and Dusp6-haploinsufficiency mice. Knockdown of DUSP6 in HepG2, a human liver-lineage cell line, also promoted a reduction of lipid accumulation, down-regulation of CYP4A, and up-regulation of phosphorylated/activated MAPK. Furthermore, inhibition of MAPK activity promoted lipid accumulation in DUSP6-knockdown HepG2 cells without affecting CYP4A expression, indicating that CYP4A expression is independent of MAPK activation. These findings highlight the significant role of DUSP6 in HFD-induced steatohepatitis through two distinct pathways involving CYP4A and MAPK.

A novel berberine derivative targeting adipocyte differentiation to alleviate TNF-α-induced inflammatory effects and insulin resistance in OP9 cells

Inflammation and insulin resistance play important roles in the development and progression of type 2 diabetes mellitus. The enhancement of adipocyte differentiation can improve insulin sensitivity by increasing glucose uptake, improving insulin signaling, and reducing inflammation. However, only a few adipogenic agents have shown clinical success in patients with type 2 diabetes mellitus. The therapeutic potential of berberine in type 2 diabetes mellitus was confirmed in terms of the target gene-disease relationship using a network pharmacology database prior to synthesizing the derivatives. Novel berberine derivatives were synthesized, and compound 3b promoted adipocyte differentiation and improvement of insulin resistance in OP9 cells. Compound 3b significantly increased the expression of key adipogenic markers including peroxisome proliferator-activated receptor γ (PPARγ) and CCAAT/enhancer binding protein β (C/EBPβ) and promoted lipid accumulation without cytotoxicity. Furthermore, tumor necrosis factor α (TNF-α)-induced inhibition of adipocyte differentiation and the elevation of inflammatory responses were reversed by compound 3b. Subsequently glucose uptake level through insulin sensitivity improvement was enhanced by compound 3b. Mechanistically, TNF-α activated mitogen-activated protein kinases (MAPKs): ERK, JNK, and p38, whereas compound 3b attenuated phosphorylation of three MAPKs. Finally, in silico molecular docking suggested the possible binding sites of compound 3b on PPARγ. Collectively, the adipogenic and glucose uptake effects of compound 3b were associated with its anti-inflammatory effects and reduced phosphorylation of MAPKs. These findings suggest that the berberine derivative compound 3b may be a potent antidiabetic agent.

The m6A modification-mediated OGDHL exerts a tumor suppressor role in ccRCC by downregulating FASN to inhibit lipid synthesis and ERK signaling

Metabolic reprogramming is a hallmark of cancer, and the impact of lipid metabolism as a crucial aspect of metabolic reprogramming on clear cell renal cell carcinoma (ccRCC) progression has been established. However, the regulatory mechanisms underlying the relationship between metabolic abnormalities and ccRCC progression remain unclear. Therefore, this study aimed to identify key regulatory factors of metabolic reprogramming in ccRCC and provide potential therapeutic targets for ccRCC patients. Potential metabolic regulatory factors in ccRCC were screened using bioinformatics analysis. Public databases and patient samples were used to investigate the aberrant expression of Oxoglutarate dehydrogenase-like (OGDHL) in ccRCC. The function of OGDHL in ccRCC growth and metastasis was evaluated through in vitro and in vivo functional experiments. Mechanistic insights were obtained through luciferase reporter assays, chromatin immunoprecipitation, RNA methylation immunoprecipitation, and mutagenesis studies. OGDHL mRNA and protein levels were significantly downregulated in ccRCC tissues. Upregulation of OGDHL expression effectively inhibited ccRCC growth and metastasis both in vitro and in vivo. Furthermore, FTO-mediated OGDHL m6A demethylation suppressed its expression in ccRCC. Mechanistically, low levels of OGDHL promoted TFAP2A expression by inhibiting ubiquitination levels, which then bound to the FASN promoter region and transcriptionally activated FASN expression, thereby promoting lipid accumulation and ERK pathway activation. Our findings demonstrate the impact of OGDHL on ccRCC progression and highlight the role of the FTO/OGDHL/TFAP2A/FASN axis in regulating ccRCC lipid metabolism and progression, providing new targets for ccRCC therapy.

The identification of a novel orally available ferroptosis inducer for the treatment of clear cell renal carcinoma

Abstract Background Background: Clear cell renal cell carcinoma (ccRCC), the most common form of kidney cancer, is typically initiated by inactivation of the von Hippel Lindau (VHL) gene resulting in constitutive activation of the hypoxia inducible factors, HIF-1α and HIF-2α. HIF-2α is a central driver of kidney cancer progression and is a well validated therapeutic target for ccRCC. Additionally, VHL loss and HIF activation promote lipid accumulation that increases the sensitivity of ccRCC cells to ferroptosis, a form of cell death characterized by iron dependent lipid peroxidation. However, classical inducers of ferroptosis, which induce ferroptosis in vitro, show limited activity in vivo due to possible redundancies in the extracellular space, suggesting a need for novel molecular targets for the induction of ferroptosis in vivo for therapeutic benefit. Methods Method: We performed a high-throughput screen to identify small molecule inhibitors of HIF-2α and performed additional validation and structure activity relationship studies to identify compounds with improved potency of HIF-2α inhibition. The molecular target of these inhibitors was determined by mass spectrometry using the Drug Affinity Responsive Target Stability (DARTS) assay, and target validation was confirmed using siRNA knockdown and NMR. In vivo anti-tumor efficacy was verified using syngeneic RENCA cells injected into Balb/C mice. Exposure to KD061 was verified using LC-MS/MS detection of KD061 in the plasma and tumors of treated mice. Results Results: We identified a novel molecule, KD061, which decreases levels of HIF-1/2α and induces ferroptosis in vivo through oral administration by targeting Iron Sulfur Cluster Assembly 2 (ISCA2). ISCA2 is a component of the late mitochondrial Iron Sulfur Cluster (L-ISC) assembly complex and ISCA2 inhibition either pharmacologically or using siRNA triggers the iron starvation response, resulting in iron/metals overload and death via ferroptosis. Cell death induced by KD061 was decreased by re-expression of VHL, suggesting synthetic lethality in the context of VHL loss. ISCA2 inhibition also decreased HIF-2α protein levels by blocking iron-responsive element (IRE)-dependent translation, and at higher concentrations, also decreased HIF-1α translation through unknown mechanisms. Treatment of tumor-bearing RENCA-Balb/C mice with KD061 significantly reduced tumor growth in vivo, decreased HIF-α levels and increased lipid peroxidation, suggesting increased ferroptosis in vivo. Treatment with KD061 was well tolerated at the therapeutic dose with no detectable effects on blood chemistry and blood counts. Conclusions Conclusions: Here, we describe the first orally available inducer of ferroptosis with anti-tumor efficacy in vivo. The targeting of ISCA2 using KD061 is a promising therapeutic strategy to inhibit HIF-1/2α and to induce ferroptosis in pVHL deficient cells. CDMRP DOD Funding: yes

Downregulation of 15-PGDH enhances MASH-HCC development via fatty acid-induced T-cell exhaustion

Hepatocellular carcinoma (HCC) mainly develops from chronic hepatitis. Metabolic dysfunction-associated steatohepatitis (MASH) has gradually become the main pathogenic factor for HCC given the rising incidence of obesity and metabolic diseases. 15-Hydroxyprostaglandin dehydrogenase (15-PGDH) degrades prostaglandin 2 (PGE2), which is known to exacerbate inflammatory responses. However, the role of PGE2 accumulation caused by 15-PGDH downregulation in the development of MASH-HCC has not been determined. We utilised the steric animal model to establish a MASH-HCC model using wild-type and 15-Pgdh+/- mice to assess the significance of PGE2 accumulation in the development of MASH-HCC. Additionally, we analysed clinical samples obtained from patients with MASH-HCC. PGE2 accumulation in the tumour microenvironment induced the production of reactive oxygen species in macrophages and the expression of cell growth-related genes and antiapoptotic genes. Conversely, the downregulation of fatty acid metabolism in the background liver promoted lipid accumulation in the tumour microenvironment, causing a decrease in mitochondrial membrane potential and CD8+ T-cell exhaustion, which led to enhanced development of MASH-HCC. 15-PGDH downregulation inactivates immune surveillance by promoting the proliferation of exhausted effector T cells, which enhances hepatocyte survival and proliferation and leads to the development of MASH-HCC. The suppression of PGE2-related inflammation and subsequent lipid accumulation leads to a reduction in the severity of MASH and inhibition of subsequent progression toward MASH-HCC.

Abstract IA007: A novel strategy to trigger ferroptosis in kidney cancer

Abstract Clear cell renal cell carcinoma (ccRCC), the most common form of kidney cancer, is typically initiated by inactivation of the von Hippel Lindau (VHL) gene resulting in constitutive activation of the hypoxia inducible factors, HIF-1α and HIF-2α. VHL loss and HIF activation promote lipid accumulation that increases the sensitivity of ccRCC cells to ferroptosis. Here, we reveal Iron Sulfur Cluster Assembly 2 (ISCA2) as a novel molecular target whose inhibition decreases HIF-1/2α levels and induces ferroptosis specifically in VHL-deficient cells. ISCA2 is a component of the late mitochondrial Iron Sulfur Cluster (L-ISC) assembly complex and ISCA2 inhibition either pharmacologically or using siRNA triggers the iron starvation response, resulting in iron/metals overload and death via ferroptosis. ISCA2 inhibition also decreases HIF-2α protein levels by blocking iron-responsive element (IRE)-dependent translation, and at higher concentrations, also decreases HIF-1α translation through unknown mechanisms. Strikingly, pharmacological inhibition of ISCA2 using a novel, orally available ISCA2 inhibitor significantly reduced ccRCC xenograft growth in vivo, decreased HIF-α levels and increased lipid peroxidation, suggesting increased ferroptosis in vivo. Thus, the targeting of ISCA2 may be a promising therapeutic strategy to inhibit HIF-1/2α and to induce ferroptosis in pVHL deficient cells. Citation Format: Mei Yee Koh. A novel strategy to trigger ferroptosis in kidney cancer [abstract]. In: Proceedings of the AACR Special Conference: Advances in Kidney Cancer Research; 2023 Jun 24-27; Austin, Texas. Philadelphia (PA): AACR; Cancer Res 2023;83(16 Suppl):Abstract nr IA007.

Secreted frizzled-related protein 5: A promising therapeutic target for metabolic diseases via regulation of Wnt signaling

Metabolic diseases pose a significant global health challenge, characterized by an imbalance in metabolism and resulting in various complications. Secreted frizzled-related protein 5 (SFRP5), an adipokine known for its anti-inflammatory properties, has gained attention as a promising therapeutic target for metabolic diseases. SFRP5 acts as a key regulator in the Wnt signaling pathway, exerting its influence on critical cellular functions including proliferation, differentiation, and migration. Its significance extends to the realm of adipose tissue biology, where it plays a central role in regulating inflammation, insulin resistance, adipogenesis, lipid metabolism, glucose homeostasis, and energy balance. By inhibiting Wnt signaling, SFRP5 facilitates adipocyte growth, promotes lipid accumulation, and contributes to a decrease in oxidative metabolism. Lifestyle interventions and pharmacological treatments have shown promise in increasing SFRP5 levels and protecting against metabolic abnormalities. SFRP5 is a pivotal player in metabolic diseases and presents itself as a promising therapeutic target. An overview of SFRP5 and its involvement in metabolic disorders and metabolism is provided in this comprehensive review. By elucidating these aspects, valuable insights can be gained to foster the development of effective strategies in combating metabolic diseases.

In vitro acne disease model from inertial focusing effect for studying the interactions between sebocyte glands and macrophages.

Acne is one of the most widespread skin diseases. The acne mechanism is intricate, involving interactions between different types of cells (i.e., sebocytes and macrophages). One of the challenges in studying the mechanism of acne is that current in vitro culture methods cannot reflect the 3D cellular environment in the tissue, including inflammatory stimuli and cellular interactions especially the interactions between sebocytes and immune cells. To solve this issue, we generated an in vitro acne disease model consisting of 3D artificial sebocyte glands and macrophages through the inertial focusing effect method. Using this model, we produced a controllable inflammatory environment similar to the acne pathogenetic process in the skin. The 3D artificial sebocyte glands and macrophages can be separated for analyzing each cell type, assisting the in-depth understanding of the acne mechanism. This study indicates that proinflammatory macrophages promote lipid accumulation and induce oxidative stress in sebocyte glands. Additionally, in an inflammatory environment, sebocyte glands induce macrophage polarization into the M1 phenotype. Employing this model for drug screening, we also demonstrated that, cannabidiol (CBD), a clinically investigated drug, is effective in restoring lipid accumulation, oxidative stress, inflammatory cytokines and macrophage polarization in the acne disease.

Gene cloning and sequence analysis of the RPL29 gene and its effect on lipogenesis in goat intramuscular adipocytes

The aim of this study was to clone the goat RPL29 gene and analyze its effect on lipogenesis in intramuscular adipocytes. Using Jianzhou big-eared goats as the object, the goat RPL29 gene was cloned by reverse transcription-polymerase chain reaction (RT-PCR), the gene structure and expressed protein sequence were analyzed by bioinformatics, and the mRNA expression levels of RPL29 in various tissues and different differentiation stages of intramuscular adipocytes of goats were detected by quantitative real-time PCR (qRT-PCR). The RPL29 overexpression vector pEGFP-N1-RPL29 constructed by gene recombination was used to transfect into goat intramuscular preadipocytes and induce differentiation. Subsequently, the effect of overexpression of RPL29 on fat droplet accumulation was revealed morphologically by oil red O and Bodipy staining, and changes in the expression levels of genes related to lipid metabolism were detected by qRT-PCR. The results showed that the length of the goat RPL29 was 507 bp, including a coding sequence (CDS) region of 471 bp which encodes 156 amino acid residues. It is a positively charged and stable hydrophilic protein mainly distributed in the nucleus of cells. Tissue expression profiling showed that the expression level of this gene was much higher in subcutaneous adipose tissue and inter-abdominal adipose tissue of goats than in other tissues (P < 0.05). The temporal expression profile showed that the gene was expressed at the highest level at 84 h of differentiation in goat intramuscular adipocytes, which was highly significantly higher than that in the undifferentiated period (P < 0.01). Overexpression of RPL29 promoted lipid accumulation in intramuscular adipocytes, and the optical density values of oil red O staining were significantly increased (P < 0.05). In addition, overexpression of RPL29 was followed by a highly significant increase in ATGL and ACC gene expression (P < 0.01) and a significant increase in FASN gene expression (P < 0.05). In conclusion, the goat RPL29 may promote intra-muscular adipocyte deposition in goats by up-regulating FASN, ACC and ATGL.

Loss of hepatic FTCD promotes lipid accumulation and hepatocarcinogenesis by upregulating PPARγ and SREBP2

Exploiting key regulators responsible for hepatocarcinogenesis is of great importance for the prevention and treatment of hepatocellular carcinoma (HCC). However, the key players contributing to hepatocarcinogenesis remain poorly understood. We explored the molecular mechanisms underlying the carcinogenesis and progression of HCC for the development of potential new therapeutic targets. The Cancer Genome Atlas-Liver Hepatocellular Carcinoma (TCGA-LIHC) and Genotype-Tissue Expression (GTEx) databases were used to identify genes with enhanced expression in the liver associated with HCC progression. A murine liver-specific Ftcd knockout (Ftcd-LKO) model was generated to investigate the role of formimidoyltransferase cyclodeaminase (FTCD) in HCC. Multi-omics analysis of transcriptomics, metabolomics, and proteomics data were applied to further analyse the molecular effects of FTCD expression on hepatocarcinogenesis. Functional and biochemical studies were performed to determine the significance of loss of FTCD expression and the therapeutic potential of Akt inhibitors in FTCD-deficient cancer cells. FTCD is highly expressed in the liver but significantly downregulated in HCC. Patients with HCC and low levels of FTCD exhibited worse prognosis, and patients with liver cirrhosis and low FTCD levels exhibited a notable higher probability of developing HCC. Hepatocyte-specific knockout of FTCD promoted both chronic diethylnitrosamine-induced and spontaneous hepatocarcinogenesis in mice. Multi-omics analysis showed that loss of FTCD affected fatty acid and cholesterol metabolism in hepatocarcinogenesis. Mechanistically, loss of FTCD upregulated peroxisome proliferator-activated receptor (PPAR)γ and sterol regulatory element-binding protein 2 (SREBP2) by regulating the PTEN/Akt/mTOR signalling axis, leading to lipid accumulation and hepatocarcinogenesis. Taken together, we identified a FTCD-regulated lipid metabolic mechanism involving PPARγ and SREBP2 signaling in hepatocarcinogenesis and provide a rationale for therapeutically targeting of HCC driven by downregulation of FTCD. Exploiting key molecules responsible for hepatocarcinogenesis is significant for the prevention and treatment of HCC. Herein, we identified formimidoyltransferase cyclodeaminase (FTCD) as the top enhanced gene, which could serve as a predictive and prognostic marker for patients with HCC. We generated and characterised the first Ftcd liver-specific knockout murine model. We found loss of FTCD expression upregulated peroxisome proliferator-activated receptor (PPAR)γ and sterol regulatory element-binding protein 2 (SREBP2) by regulating the PTEN/Akt/mTOR signalling axis, leading to lipid accumulation and hepatocarcinogenesis, and provided a rationale for therapeutic targeting of HCC driven by downregulation of FTCD.

Engineering multicomponent tissue by spontaneous adhesion of myogenic and adipogenic microtissues cultured with customized scaffolds

The integration of intramuscular fat—or marbling—into cultured meat will be critical for meat texture, mouthfeel, flavor, and thus consumer appeal. However, culturing muscle tissue with marbling is challenging since myocytes and adipocytes have different media and scaffold requirements for optimal growth and differentiation. Here, we present an approach to engineer multicomponent tissue using myogenic and adipogenic microtissues. The key innovation in our approach is the engineering of myogenic and adipogenic microtissues using scaffolds with customized physical properties; we use these microtissues as building blocks that spontaneously adhere to produce multicomponent tissue, or marbled cultured meat. Myocytes are grown and differentiated on gelatin nanofiber scaffolds with aligned topology that mimic the aligned structure of skeletal muscle and promotes the formation of myotubes in both primary rabbit skeletal muscle and murine C2C12 cells. Pre-adipocytes are cultured and differentiated on edible gelatin microbead scaffolds, which are customized to have a physiologically-relevant stiffness, and promote lipid accumulation in both primary rabbit and murine 3T3-L1 pre-adipocytes. After harvesting and stacking the individual myogenic and adipogenic microtissues, we find that the resultant multicomponent tissues adhere into intact structures within 6–12 h in culture. The resultant multicomponent 3D tissue constructs show behavior of a solid material with a Young’s modulus of ∼ 2 ± 0.4 kPa and an ultimate tensile strength of ∼ 23 ± 7 kPa without the use of additional crosslinkers. Using this approach, we generate marbled cultured meat with ∼ mm to ∼ cm thickness, which has a protein content of ∼ 4 ± 2 g/100 g that is comparable to a conventionally produced Wagyu steak with a protein content of ∼ 9 ± 4 g/100 g. We show the translatability of this layer-by-layer assembly approach for microtissues across primary rabbit cells, murine cell lines, as well as for gelatin and plant-based scaffolds, which demonstrates a strategy to generate edible marbled meats derived from different species and scaffold materials.

Lipid accumulation and gene expression changes induced by tributyltin exposure in primary hepatocytes of lined seahorse (Hippocampus erectus)

Tributyltin (TBT), an antifouling biocide frequently detected in aquatic systems, is generally considered to be an environmental obesogen. However, alterations in lipid metabolism in aquatic animals that are exposed to TBT are scarcely known. This study examined the effects of in vitro exposure to TBT on hepatic lipid homeostasis in the lined seahorse (Hippocampus erectus). Primary seahorse hepatocyte cultures were established for the first time. TBT exposure (100 and 500 nM for 24 h) significantly promoted lipid accumulation in seahorse hepatocytes and drastically reduced the number of active intracellular lysosomes. Furthermore, exposure to TBT significantly upregulated the gene expression of lipogenic enzymes and transcription factors but downregulated that of genes involved in the catabolism of lipid droplets in seahorse hepatocytes. These results indicate that TBT disrupts hepatic lipid homeostasis by simultaneously promoting lipid synthesis and inhibiting lipid droplet breakdown in seahorses. The present study extends our understanding of the utilization of primary hepatocytes from marine animals for toxicological research, and the molecular evidence of the effects of TBT on hepatic lipid homeostasis in teleost fishes.

AIP4 regulates adipocyte differentiation by targeting C/EBPα for ubiquitin-mediated proteasomal degradation.

Adipogenesis, that is, the formation of terminally differentiated adipocytes is intricately regulated by transcription factors where CCAAT/enhancer binding protein alpha (C/EBPα) plays a key role. In the current study, we demonstrate that E3 ubiquitin ligase AIP4 negatively regulates C/EBPα protein stability leading to reduced adipogenesis. While AIP4 overexpression in 3T3-L1 cells preadipocytes inhibited lipid accumulation when treated with differentiation inducing media (MDI), AIP4 depletion was sufficient to partially promote lipid accumulation even in the absence of MDI. Mechanistically, overexpression of AIP4 inhibited protein levels of both ectopically expressed as well as endogenous C/EBPα while catalytically inactive AIP4 failed. On the contrary, AIP4 depletion profoundly enhanced endogenous C/EBPα protein levels. The observation that AIP4 levels decrease with concomitant increase in C/EBPα levels during adipocyte differentiation further indicated that AIP4 negatively regulates C/EBPα levels. We further show that AIP4 physically interacts with C/EBPα and ubiquitinates it leading to its proteasomal degradation. AIP4 promoted K48-linked ubiquitination of C/EBPα while catalytically inactive AIP4-C830A failed. Taken together, our data demonstrate that AIP4 inhibits adipogenesis by targeting C/EBPα for ubiquitin-mediated proteasome degradation.

Biotechnological product potential of Auxenochlorella protothecoides including biologically active compounds (BACs) under nitrogen stress conditions.

Nitrogen stress can influence microalgae's growth characteristics, and microalgae grown in nitrogen-deficient conditions may produce higher or lower levels of biotechnological products as a result of metabolic changes. In photoautotrophic and heterotrophic cultures, nitrogen limitation has been proven effective in promoting lipid accumulation. In spite of this, no study has demonstrated a significant correlation between lipid content and other biotechnological products such as bioactive compounds (BACs).This research examines a strategy for lipid accumulation as well as the potential production of BACs with antibacterial properties in parallel with that strategy. This concept involved the treatment of the microalga Auxenochlorella protothecoides with low and high concentrations of ammonium (NH4+). This particular experiment reached a maximum lipid content of 59.5% using a 0.8 mM NH4+ concentration, resulting in the yellowing of the chlorophyll levels. Agar diffusion assays were conducted to determine the antibacterial activity of different extracts derived from the biomass when stressed with different levels of nitrogen.Algal extracts prepared by a variety of solvents showed different levels of antibacterial activity against representative strains of both gram-negative (Escherichia coli) and gram-positive (Staphylococcus aureus) bacteria. Among the extracts tested, 500mg/L ethyl acetate extract had the greatest antibacterial activity against Escherichia coli. In order to identify the components responsible for the extract's antibacterial activity, fatty acid methyl ester (FAME) analysis was performed. It has been suggested that the lipid fraction may be a valuable indicator of these activities since some lipid components are known to possess antimicrobial properties. In this regard, it was found that the amount of polyunsaturated fatty acid (PUFA) significantly decreased by 53.4% under the conditions with the highest antibacterial activity observed.

PEMT Mediates Hepatitis C Virus-Induced Steatosis, Explains Genotype-Specific Phenotypes and Supports Virus Replication.

The hepatitis C virus (HCV) relies on cellular lipid pathways for virus replication and also induces liver steatosis, but the mechanisms involved are not clear. We performed a quantitative lipidomics analysis of virus-infected cells by combining high-performance thin-layer chromatography (HPTLC) and mass spectrometry, using an established HCV cell culture model and subcellular fractionation. Neutral lipid and phospholipids were increased in the HCV-infected cells; in the endoplasmic reticulum there was an ~four-fold increase in free cholesterol and an ~three-fold increase in phosphatidyl choline (p < 0.05). The increase in phosphatidyl choline was due to the induction of a non-canonical synthesis pathway involving phosphatidyl ethanolamine transferase (PEMT). An HCV infection induced expression of PEMT while knocking down PEMT with siRNA inhibited virus replication. As well as supporting virus replication, PEMT mediates steatosis. Consistently, HCV induced the expression of the pro-lipogenic genes SREBP 1c and DGAT1 while inhibiting the expression of MTP, promoting lipid accumulation. Knocking down PEMT reversed these changes and reduced the lipid content in virus-infected cells. Interestingly, PEMT expression was over 50% higher in liver biopsies from people infected with the HCV genotype 3 than 1, and three times higher than in people with chronic hepatitis B, suggesting that this may account for genotype-dependent differences in the prevalence of hepatic steatosis. PEMT is a key enzyme for promoting the accumulation of lipids in HCV-infected cells and supports virus replication. The induction of PEMT may account for virus genotype specific differences in hepatic steatosis.