Intracellular Lipid Research Articles

Unconventional Extraction Methods of Oleaginous Yeast Cell Pretreatment and Disruption

Extraction is one of the most commonly used methods for obtaining and purifying chemical compounds for commercial usage. The aim of this study was to evaluate the effect of unconventional permeabilization and cell disruption methods on the yield of lipid extraction from cells of the oleaginous yeast Yarrowia lipolytica. Batch cultures in a medium with molasses and waste post-frying oil were carried out. The biomass was subjected to pulsed electric field (PEF), high-pressure processing (HPP), ultrasounds (US), and several conventional processing techniques with chemical and mechanical agents (glass beads, acetone, Triton and Tween surfactants). The effectiveness of the applied methods, either on cell permeabilization or cell disruption, was investigated by analyzing the oil and total protein extraction yield and oil leaching efficiency, as well as by using microscope images. The PEF and US treatments proved to be effective permeabilization methods as a step of sample pretreatment for extraction. These unconventional physical methods could efficiently increase intracellular lipid extraction yield in solvent applications.

Simultaneous Lipid and Carotenoid Production via Rhodotorula paludigena CM33 Using Crude Glycerol as the Main Substrate: Pilot-Scale Experiments.

Rhodotorula paludigena CM33 is an oleaginous yeast that has been demonstrated to accumulate substantial quantities of intracellular lipids and carotenoids. In this study, crude glycerol, a by-product of biodiesel production, was used as a carbon source to enhance the accumulation of lipids and carotenoids in the cells. The culture conditions were first optimized using response surface methodology, which revealed that the carotenoid concentration and lipid content improved when the concentration of crude glycerol was 40 g/L. Different fermentation conditions were also investigated: batch, repeated-batch, and fed-batch conditions in a 500 L fermenter. For fed-batch fermentation, the maximum concentrations of biomass, lipids, and carotenoids obtained were 46.32 g/L, 37.65%, and 713.80 mg/L, respectively. A chemical-free carotenoid extraction method was also optimized using high-pressure homogenization and a microfluidizer device. The carotenoids were found to be mostly beta-carotene, which was confirmed by HPLC (high pressure liquid chromatography), LC-MS (liquid chromatography-mass spectrometry), and NMR (nuclear magnetic resonance). The results of this study indicate that crude glycerol can be used as a substrate to produce carotenoids, resulting in enhanced value of this biodiesel by-product.

Nanoplanktonic diatom rapidly alters sinking velocity via regulating lipid content and composition in response to changing nutrient concentrations

Diatom sinking plays a crucial role in the global carbon cycle, accounting for approximately 40% of marine particulate organic carbon export. While oceanic models typically represent diatoms as microphytoplankton (> 20 μm), it is important to recognize that many diatoms fall into the categories of nanophytoplankton (2-20 μm) and picophytoplankton (< 2 μm). These smaller diatoms have also been found to significantly contribute to carbon export. However, our understanding of their sinking behavior and buoyancy regulation mechanisms remains limited. In this study, we investigate the sinking behavior of a nanoplanktonic diatom, Phaeodactylum tricornutum (P. tricornutum), which exhibits rapid changes in sinking behavior in response to varying nutrient concentrations. Our results demonstrate that a higher sinking rate is observed under phosphate limitation and depletion. Notably, in phosphate depletion, the sinking rate of P. tricornutum was 0.79 ± 0.03 m d-1, nearly three times that of the previously reported sinking rates for Skeletonema costatum, Ditylum brightwellii, and Chaetoceros gracile. Furthermore, during the first 6 h of phosphate spike, the sinking rate of P. tricornutum remained consistently high. After 12 h of phosphate spike, the sinking rate decreased to match that of the phosphate repletion phase, only to increase again over the next 12 hours due to phosphate depletion. This rapid sinking behavior contributes to carbon export and potentially allows diatoms to exploit nutrient-rich patches when encountering increased nutrient concentrations. We also observed a significant positive correlation (P< 0.001) between sinking rate and lipid content (R = 0.91) during the phosphate depletion and spike experiment. It appears that P. tricornutum regulates its sinking rate by increasing intracellular lipid content, particularly digalactosyldiacylglycerol, hexosyl ceramide, monogalactosyldiacylglycerol, and triglycerides. Additionally, P. tricornutum replaces phospholipids with more dense membrane sulfolipids, such as sulfoquinovosyldiacylglycerol under phosphate shortage. These findings shed light on the intricate relationship between nutrient availability, sinking behavior, and lipid composition in diatoms, providing insights into their adaptive strategies for carbon export and nutrient utilization.

Rab18 Drift in Lipid Droplet and Endoplasmic Reticulum Interactions of Adipocytes under Obesogenic Conditions.

The adipose tissue stores excess energy in the form of neutral lipids within adipocyte lipid droplets (LDs). The correct function of LDs requires the interaction with other organelles, such as the endoplasmic reticulum (ER) as well as with LD coat-associated proteins, including Rab18, a mediator of intracellular lipid trafficking and ER-LD interaction. Although perturbations of the inter-organelle contact sites have been linked to several diseases, such as cancer, no information regarding ER-LD contact sites in dysfunctional adipocytes from the obese adipose tissue has been published to date. Herein, the ER-LD connection and Rab18 distribution at ER-LD contact sites are examined in adipocytes challenged with fibrosis and inflammatory conditions, which represent known hallmarks of the adipose tissue in obesity. Our results show that adipocytes differentiated in fibrotic conditions caused ER fragmentation, the expansion of ER-LD contact sites, and modified Rab18 dynamics. Likewise, adipocytes exposed to inflammatory conditions favored ER-LD contact, Rab18 accumulation in the ER, and Rab18 redistribution to large LDs. Finally, our studies in human adipocytes supported the suggestion that Rab18 transitions to the LD coat from the ER. Taken together, our results suggest that obesity-related pathogenic processes alter the maintenance of ER-LD interactions and interfere with Rab18 trafficking through these contact sites.

CXCL6 promotes the progression of NAFLD through regulation of PPARα

An increasing number of studies have shown that Nonalcoholic fatty liver disease (NAFLD) is strongly associated with obesity, insulin resistance, dyslipidemia, hypertension and metabolic syndrome, but its specific pathogenesis remains unclear. By analyzing GEO database, we found CXCL6 was upregulated in liver tissues of patients with NAFLD. We also confirmed with qPCR that CXCL6 is highly expressed in serum of patients with NAFLD. To identify the underlying impact of CXCL6 on NAFLD, we established animal and cell models of NAFLD. Similarly, we confirmed by qPCR and Western blot that CXCL6 was upregulated in the NAFLD model in vitro and vivo. After transfecting NAFLD cells with siRNA targeting CXCL6 (si-CXCL6), a series of functional experiments were carried out, and these data indicated that the inhibition of CXCL6 reduced intracellular lipid deposition, decreased AST, ALT and TG level, facilitate cell proliferation and suppress their apoptosis. Furthermore, western blot and qPCR analyses displayed that the suppression of CXCL6 could raise the PPARα expression, but PPAR α inhibitor, GW6471 could partially counteract this effect. What’s more, Oil Red O staining, biochemical analyzer and TG detection kit revealed that GW6471 could reverse the inhibitory effect of si-CXCL6 on NAFLD. In summary, we provide convincing evidence that CXCL6 is markedly elevated in NAFLD, and the CXCL6/PPARα regulatory network mediates disease progression of NAFLD.

Periostin facilitates ovarian cancer recurrence by enhancing cancer stemness

The lethality of epithelial ovarian cancer (OC) is largely due to a high rate of recurrence and development of chemoresistance, which requires synergy between cancer cells and the tumor microenvironment (TME) and is thought to involve cancer stem cells. Our analysis of gene expression microarray data from paired primary and recurrent OC tissues revealed significantly elevated expression of the gene encoding periostin (POSTN) in recurrent OC compared to matched primary tumors (p = 0.015). Secreted POSTN plays a role in the extracellular matrix, facilitating epithelial cell migration and tissue regeneration. We therefore examined how elevated extracellular POSTN, as we found is present in recurrent OC, impacts OC cell functions and phenotypes, including stemness. OC cells cultured with conditioned media with high levels of periostin (CMPOSTNhigh) exhibited faster migration (p = 0.0044), enhanced invasiveness (p = 0.006), increased chemoresistance (p < 0.05), and decreased apoptosis as compared to the same cells cultured with control medium (CMCTL). Further, CMPOSTNhigh-cultured OC cells exhibited an elevated stem cell side population (p = 0.027) along with increased expression of cancer stem cell marker CD133 relative to CMCTL-cultured cells. POSTN-transfected 3T3-L1 cells that were used to generate CMPOSTNhigh had visibly enhanced intracellular and extracellular lipids, which was also linked to increased OC cell expression of fatty acid synthetase (FASN) that functions as a central regulator of lipid metabolism and plays a critical role in the growth and survival of tumors. Additionally, POSTN functions in the TME were linked to AKT pathway activities. The mean tumor volume in mice injected with CMPOSTNhigh-cultured OC cells was larger than that in mice injected with CMCTL-cultured OC cells (p = 0.0023). Taken together, these results show that elevated POSTN in the extracellular environment leads to more aggressive OC cell behavior and an increase in cancer stemness, suggesting that increased levels of stromal POSTN during OC recurrence contribute to more rapid disease progression and may be a novel therapeutic target. Furthermore, they also demonstrate the utility of having matched primary-recurrent OC tissues for analysis and support the need for better understanding of the molecular changes that occur with OC recurrence to develop ways to undermine those processes.

Abstract A104: ADAR1-associated metabolic vulnerabilities in triple-negative breast cancer

Abstract Triple-negative breast cancer (TNBC), lacking expression of estrogen, progesterone, and HER2 receptors, is aggressive and lacks targeted treatment options. Therefore, the mission to identify actionable targets remains the top priority. We recently demonstrated that an RNA editing enzyme, adenosine deaminase acting on RNA 1 (ADAR1), plays important roles in TNBC’s ability to initiate and maintain tumorigenesis. We posit that ADAR1 functions as a homeostatic factor protecting TNBC from stresses when otherwise kept in check, contributing to tumorigenesis. By both targeted and unbiased approaches, we identified two metabolic vulnerabilities, 1) iron-dependent cell death pathway, ferroptosis, and 2) serine addiction, associated with ADAR1 in TNBC. By performing single-reaction monitoring-based liquid chromatography coupled to mass spectrometry (LC-MS) to measure intracellular lipid contents, we showed that ADAR1 loss in TNBC cells increased the abundance of polyunsaturated fatty acid phospholipids (PUFA-PL), of which peroxidation is the primary driver of ferroptosis. Transcriptomic analysis and phenotypic drug screen using a ferroptosis-focused library were performed to identify underlying mechanisms and drug candidates synergizing with ADAR1 loss through ferroptosis, respectively. Meanwhile, an unbiased screen using a FDA-approved compound library revealed that ADAR1 loss also sensitizes TNBC cell line MDA-MB231 to inhibition of the serine biosynthesis pathway. We showed that ADAR1 knockdown leads to altered serine transport, and a metabolic flux analysis using labeled (U-13C) glucose will be performed to determine if ADAR1 loss affects de novo serine biosynthesis and one-carbon metabolism. By testing the hypothesis that ADAR1 regulates the metabolic fitness of TNBC by desensitizing ferroptosis and serine addiction, we aim to leverage these metabolic vulnerabilities to inform basic, pre-clinical, and clinical studies to develop novel therapeutic strategies for TNBC. Citation Format: Che-Pei Kung, Leah P Shriver, Nick Terzich, Ma Xenia G Ilagen, Mike Prinsen, Sua Ryu, Raleigh D Kladney, Gary J Patti, Leonard B Maggi, Jason D Weber. ADAR1-associated metabolic vulnerabilities in triple-negative breast cancer [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr A104.

TFAM-mediated intercellular lipid droplet transfer promotes cadmium-induced mice nonalcoholic fatty liver disease

Cadmium (Cd) is an important environmental pollutant. Herein, we discovered a new way of lipid accumulation, where lipid droplets can be transferred across cells. In this study, mice and AML12 cells were used to establish models of Cd poisoning. After Cd treatment, the level of TFAM was reduced, thereby regulating the reconstitution of the cytosolic actin filament network. MYH9 is a myosin involved in cell polarization, migration, and movement of helper organelles. Rab18 is a member of the Rab GTPase family, which localizes to lipid droplets and regulates lipid drop dynamics. In this study, we found that Cd increases the interaction between MYH9 and Rab18. However, TFAM overexpression alleviated the increase in Cd-induced interaction between MYH9 and Rab18, thereby reducing the transfer of intercellular lipid droplets and the accumulation of intracellular lipids. Through a co-culture system, we found that the transferred lipid droplets can act as a signal to form an inflammatory storm-like effect, and ACSL4 can act as an effector to transfer lipid droplets and promote lipid accumulation in surrounding cells. These results suggest that TFAM can be used as a new therapeutic target for Cd-induced lipid accumulation in the liver.

Clearance of intracellular lipids by lipophagy impaired in APOE ε4 human astrocytes

AbstractBackgroundApolipoprotein E (APOE) ɛ4 is the strongest known genetic risk factor for Alzheimer’s disease. APOE ɛ4 has been found to impair lipid metabolism in human glia causing accumulation of intracellular lipid droplets. We hypothesized that this phenotype in human APOE ɛ4/ɛ4 astrocytes was due to defective clearance via lipophagy.MethodGlobal transcriptomic analyses were performed on astrocytes derived from isogenic and population model human induced pluripotent stem cells (iPSCs). Fluorescent reporters, immunocytochemistry, and western blotting were used to quantify lipid droplets and autophagic machinery in iPSC‐based models.ResultGlobal transcriptomic analyses reveal that APOE ε4 drives autophagy dysregulation in astrocytes from human iPSCs. High‐content imaging of fluorescent autophagy reporters and immunocytochemistry revealed accumulation of lipid droplets and autophagy machinery in APOE ɛ4/ɛ4 astrocytes which was ameliorated by induction of autophagy.ConclusionHuman iPSC‐derived APOE ɛ4/ɛ4 astrocytes exhibit impaired lipophagy.

#67 : Fatty Acid Supplementation into Warming Solutions Improves Outgrowth Competence Human Vitrified Blastocysts

Background and Aims: The vitrification procedure decreased intracellular lipid content and subsequent developmental competence. Furthermore, a recent study reported that fatty acid (FA) supplementation into warming solutions increased intracellular lipid content and improved developmental competence by stimulating the beta-oxidation pathway in mice and bovines. However, in humans, the effects of FA addition to warming solutions on the viability of vitrified blastocysts remain unclear. In this study, we examined the implantation competence of blastocysts warmed by using the FA-supplemented solutions by comparing the in vitro outgrowth. Method: A total of 216 discarded human vitrified blastocysts donated for research by consenting couples were randomly allocated, to be warmed in solutions either with (FA, n = 98) or without FA (control, n = 118). Vitrified blastocysts were cultured for 120 min after warming. The blastocysts were plated on fibronectin-coated dishes and cultured for 96 h to assess the blastocyst competency for adhesion and outgrowth. Results: The embryo characteristics were comparable between the control and FA groups. The adhesion rate at 96 h after the outgrowth culture in the FA group (74.5%) was comparable with that in the control group (74.6%). The 1.3-fold increase in the outgrowth area was observed in the FA group, compared to the control group ([Formula: see text] = 0.0371). Furthermore, the sub-group analysis demonstrated that a more beneficial effect of FA supplementation on blastocyst outgrowth was observed when the trophectoderm was evaluated as grade A ([Formula: see text] = 0.0065). Conclusion: The FA supplementation into warming solutions increased the outgrowth competence of vitrified human blastocysts. These results suggest that the pregnancy outcomes, including the implantation rate, after frozen blastocyst transfers would be improved by using the FA-supplemented solutions.

BAP18 acting as a novel peroxisome proliferator-activated receptor α co-regulator contributes to hepatocellular carcinoma progression

Hepatocellular carcinoma (HCC) is a common malignancy worldwide with a poor prognosis. The therapeutic outcomes of HCC patients are urgently needed to be improved, and predictive biomarkers for the optimal treatment selection remains to be further defined. In the present study, our results showed that BPTF-associated protein of 18 KDa (BAP18) was highly expressed in HCC tissues. In cultured HCC cells, BAP18 regulated a subset of down-stream genes involved in different functions, particularly including peroxisome proliferator-activated receptor (PPAR) pathway and lipid metabolism. Furthermore, BAP18 co-activated PPARα-mediated transactivation and facilitated the recruitment of nucleosome acetyltransferase of H4 (NuA4)/tat interacting protein 60 (TIP60) complex, thereby increasing histone H4 acetylation on stearoyl-CoA desaturase 1 (SCD1) loci. In addition, BAP18 promoted HCC cell proliferation, increased intracellular lipid levels and enhanced cell survival under the metabolic stress conditions, such as glucose limitation or tyrosine kinase inhibitors (TKIs) treatment. Importantly, higher BAP18 expression was positively correlated with the postoperative recurrence and the poor disease-free survival in clinical patients receiving sorafenib treatment. Altogether, we discovered that BAP18 plays an oncogenic role in the survival and proliferation of HCC cells, and BAP18 may serve as a predictive biomarker for adjunct TKIs treatment in patients with HCC, and further facilitate the precise treatment.

An Overview on Lipid Droplets Accumulation as Novel Target for Acute Myeloid Leukemia Therapy.

Metabolic reprogramming is a key alteration in tumorigenesis. In cancer cells, changes in metabolic fluxes are required to cope with large demands on ATP, NADPH, and NADH, as well as carbon skeletons. In particular, dysregulation in lipid metabolism ensures a great energy source for the cells and sustains cell membrane biogenesis and signaling molecules, which are necessary for tumor progression. Increased lipid uptake and synthesis results in intracellular lipid accumulation as lipid droplets (LDs), which in recent years have been considered hallmarks of malignancies. Here, we review current evidence implicating the biogenesis, composition, and functions of lipid droplets in acute myeloid leukemia (AML). This is an aggressive hematological neoplasm originating from the abnormal expansion of myeloid progenitor cells in bone marrow and blood and can be fatal within a few months without treatment. LD accumulation positively correlates with a poor prognosis in AML since it involves the activation of oncogenic signaling pathways and cross-talk between the tumor microenvironment and leukemic cells. Targeting altered LD production could represent a potential therapeutic strategy in AML. From this perspective, we discuss the main inhibitors tested in in vitro AML cell models to block LD formation, which is often associated with leukemia aggressiveness and which may find clinical application in the future.

Injectable alginate hydrogel promotes antitumor immunity through glucose oxidase and Fe3+ amplified RSL3-induced ferroptosis

Ferroptosis induced by RAS-selective lethal small molecule 3 (RSL3) can trigger anti-tumor immune responses by reversing the immunosuppressive tumor microenvironment (TME). However, it is challenging to achieve sufficient ferroptosis in the tumor via RSL3 alone. Because of the excellent reactive oxygen species (ROS) production capacity of glucose oxidase (GOx) and Fe3+, we hypothesized that GOx and Fe3+ could increase intracellular lipid peroxidation (LPO) accumulation, and strengthen RSL3-induced ferroptosis in tumor cells. Herein we designed an in-situ gelation strategy based on sodium alginate (SA) to realize localized transport and specific retention of GOx, RSL3, and Fe3+ in tumor tissues. We loaded hydrophobic RSL3 with the tannic acid (TA)/Fe3+ complexes to form nanoparticles (RTF NPs). GOx diluted in the SA solution was blended with RTF NPs to obtain a homogeneous solution. The solution could form hydrogels in the tumor site (RTFG@SA) upon injection. The retained GOx and Fe3+ amplified the induction of ferroptosis by RSL3, augmented immunogenic cell death (ICD) and promoted antitumor immunity. The RTFG@SA hydrogel presented a significant restraint of tumor growth and metastasis in the 4T1 tumor model. This hydrogel could offer an effective means of co-delivery of hydrophilic drugs, hydrophobic drugs, and metal ions.

Caveolin-1 deficiency alleviates palmitate-induced intracellular lipid accumulation and inflammation in pancreatic β cells.

Lipotoxicity-induced pancreatic β cell damage is a strong predictor of type 2 diabetes mellitus (T2DM). Our previous work showed that Caveolin-1 (Cav-1) depletion decreased β-cell apoptosis and improved β-cell viability. Further microarray analysis indicated significant changes in the expression of genes related to fatty acid metabolism and inflammation. The objective of this study was to explore the role of Cav-1 in intracellular lipid accumulation and inflammation in β cells under lipotoxic conditions. Here, we established a β-cell-specific Cav-1 knockout (β-Cav-1 KO) mouse model and a CAV-1 depleted β cell line (NIT-1). We found that Cav-1 silencing significantly reduced palmitate (PA)-induced intracellular triglyceride (TG) accumulation and decreased proinflammatory factor expression in both the mouse and cell models. Further mechanistic investigation revealed that amelioration of lipid metabolism was achieved through the downregulation of lipogenic markers (SREBP-1c, FAS and ACC) and upregulation of a fatty acid oxidation marker (CPT-1). Meanwhile, decrease of inflammatory cytokines (IL-6, TNF-α, and IL-1β) secretion was found with the involvement of the IKKβ/NF-κB signaling pathways. Our findings suggest that Cav-1 is of considerable importance in regulating lipotoxicity-induced β-cell intracellular lipid accumulation and inflammation.

Smart Ratiometric SERS Nanoprobe for Real-Time Monitoring Hydrogen Peroxide in Living Cells during NADH Treatment Associated with Ferroptosis.

Studying the oxidative stress, especially the reactive oxygen species (ROS) response of ferroptosis, is crucial for the diagnosis and treatment of cancer based on ferroptosis. However, reliable quantitative analysis of intracellular ROS in cancer treatment for drug screening is still a challenge. Herein, a superior ratiometric SERS nanoprobe was developed for in situ, real-time, and highly sensitive detection of content variation of H2O2 within living cells. The SERS nanoprobe was prepared by coassembly of the internal standard molecule p-mercaptobenzonitrile and the reporter molecule p-mercaptophenylboronic acid on the surface of gold nanoparticles, used for synergistic calibration and detection of H2O2, which enables reliable detection of the true content of intracellular H2O2 without the interference of other substances in cells. Based on the nanoprobe, we found that the level of intracellular H2O2 of cancer cells was increased after the nicotinamide adenine dinucleotide (NADH) treatment, with a dose-dependence to the concentration of NADH. High doses of NADH (above 20 mM) can induce cell death by means of ferroptosis associated with the level elevation of intracellular lipid hydroperoxides. This study highlights the potential of the SERS nanoprobe for tracking content variation of cellular H2O2 and understanding its roles in screening new anticancer drugs.

Cimifugin inhibits adipogenesis and TNF-α-induced insulin resistance in 3T3-L1 cells.

To investigate the effects of cimifugin on adipogenesis and tumor necrosis factor (TNF-α)-induced insulin resistance (IR) and inflammation in 3T3-L1 adipocytes. 3T3-L1 adipocytes were treated with 3-isobutyl-1-methyl-xanthine, dexamethasone, and insulin or cimifugin and then Oil Red O staining and intracellular triglyceride content detection were performed to assess adipogenesis. Subsequently, after cimifugin treatment, TNF-α was used to induce IR and inflammation. The results showed that cimifugin reduced intracellular lipids accumulation of 3T3-L1 adipocytes. Cimifugin improved IR of 3T3-L1 adipocytes induced by TNF-α, as reflected in decreased adiponectin, GLUT-4, and IRS-1 mRNA and protein expression. Moreover, cimifugin reduced TNF-α-induced pro-inflammatory factors production and phospho-P65 expression, and MAPK pathway activation in the 3T3-L1 adipocytes. These findings suggested that cimifugin might be useful for the prevention and therapy of obesity-related IR and inflammation.

PI3K-C2β limits mTORC1 signaling and angiogenic growth.

Phosphoinositide 3-kinases (PI3Ks) phosphorylate intracellular inositol lipids to regulate signaling and intracellular vesicular trafficking. Mammals have eight PI3K isoforms, of which class I PI3Kα and class II PI3K-C2α are essential for vascular development. The class II PI3K-C2β is also abundant in endothelial cells. Using in vivo and in vitro approaches, we found that PI3K-C2β was a critical regulator of blood vessel growth by restricting endothelial mTORC1 signaling. Mice expressing a kinase-inactive form of PI3K-C2β displayed enlarged blood vessels without corresponding changes in endothelial cell proliferation or migration. Instead, inactivation of PI3K-C2β resulted in an increase in the size of endothelial cells, particularly in the sprouting zone of angiogenesis. Mechanistically, we showed that the aberrantly large size of PI3K-C2β mutant endothelial cells was caused by mTORC1 activation, which sustained growth in these cells. Consistently, pharmacological inhibition of mTORC1 with rapamycin normalized vascular morphogenesis in PI3K-C2β mutant mice. Together, these results identify PI3K-C2β as a crucial determinant of endothelial signaling and illustrate the importance of mTORC1 regulation during angiogenic growth.

Comprehensive Exploration of the Growth and Lipid Synthesis Phases of T. oleaginosus Cultures Implementing Design of Experiments and Response Surface Methodology.

Trichosporon oleaginosus is an unconventional oleaginous yeast distinguished by its remarkable capacity to accumulate lipids in excess of 70% of its dry weight, particularly when cultivated in nitrogen-restricted conditions with ample carbon sources. A pivotal question that arises pertains to the nutrient dynamics in the culture medium, which give rise to both the excessive lipid content and corresponding lipid concentration. While previous research has predominantly focused on evaluating the impact of the initial carbon-to-nitrogen (C/N) ratio on lipid production, the precise critical thresholds of glucose and ammonium sulfate ((NH4)2SO4) at which growth and intracellular lipid production are either stimulated or impeded remain inadequately defined. This study employs an experimental design and response surface methodology to investigate the complex mechanism of lipid accumulation and its interaction with cellular growth. Application of the aforementioned methodologies resulted in the production of 10.6 g/L of microbial oil in batch cultures under conditions that correspond to a C/N ratio of 76. However, the primary objective is to generate knowledge to facilitate the development of efficient fed-batch cultivation strategies that optimize lipid production exclusively employing inorganic nitrogen sources by finely adjusting carbon and nitrogen levels. The intricate interaction between these levels is comprehensively addressed in the present study, while it is additionally revealed that as glucose levels rise within a non-inhibitory range, lipid-free biomass production decreases while lipid accumulation simultaneously increases. These findings set the stage for further exploration and the potential development of two-stage cultivation approaches, aiming to fully decouple growth and lipid production. This advancement holds the promise of bringing microbial oil production closer to commercial viability.

Identification of a Novel Long Non-Coding RNA G8110 That Modulates Porcine Adipogenic Differentiation and Inflammatory Responses

Long non-coding RNAs (lncRNAs) have been extensively studied, and their crucial roles in adipogenesis, lipid metabolism, and gene expression have been revealed. However, the exact regulatory or other mechanisms by which lncRNAs influence the functioning of mesenteric adipose tissue (MAT) remain largely unknown. In this paper, we report the identification of a new lncRNA, named G8110, from the MAT of Bama pigs. The coordinated expression levels of lncRNA G8110 and NFE2L1 were significantly decreased in the MAT of obese Bama pigs compared with those in the MAT of lean pigs. Using a bone mesenchymal stem cell adipogenic differentiation model, we found that lncRNA G8110 played a role in adipocyte differentiation by positively regulating NFE2L1. We also found that lncRNA G8110 inhibited the formation of intracellular lipid synthesis, promoted lipid metabolism, and inhibited the expression of inflammatory cytokines. Our findings regarding lipid synthesis may further promote the role of lncRNAs in driving adipose tissue remodeling and maintaining metabolic health.

Antifungal activity and mechanism of chitosan against Fusarium solani caused ginger soft rot during postharvest storage

Postharvest rot caused by Fusarium solani (F. solani) results in enormous economic loss to the global vegetables industry. Chitosan has been shown to be effective in control postharvest diseases of vegetables, but the specific mechanism of chitosan on F. solani has not been studied. In this study, we evaluated antifungal activity of chitosan against F. solani caused soft rot in vitro and in vivo. Antifungal assay results showed that chitosan significantly inhibited the mycelial growth and spore germination of F. solani. Simultaneously, chitosan showed disruptive activity on the mycelial morphology and cell membrane, which were achieved by decreasing intracellular sugars, proteins and lipids content. Furthermore, chitosan treatment significantly decreased tricarboxylic acid (TCA)-related enzymes activity, reducing the number of metabolites in the TCA cycle. UPLC-Triple-TOF/MS-based metabolomics analysis showed that chitosan interferes with fungal metabolic pathways, mainly related to energy metabolism, lipid metabolism and sugar metabolism. Exogenous α-D-glucose and linoleic acid treatment promoted mycelia growth, verifying the sugar catabolism and TCA cycle was involved in the inhibition of chitosan against F. solani. The in vivo study showed that chitosan treatment effectively reduced disease rate and severity of postharvest soft rot through activating the biochemical defense response and modulating the expression of phytopathogenic responses LysM genes. These results contribute to explore the underlying mechanisms of chitosan against F. solani.