Accumulation Of Droplets Research Articles

Chanarin\u2013Dorfman syndrome: clinical/genetic features and natural history in six Pakistani patients

BackgroundChanarin–Dorfman syndrome (CDS; OMIM # 275630) is a rare neutral lipid storage disorder caused by mutation in ABHD5 (a/b hydrolase domain containing 5″) a cofactor for adipose triglyceride lipase (ATGL) resulting in intracellular accumulation of triacylglycerol (TG) in numerous body tissues. It is an autosomal recessive disorder mutation in ABHD5 that causes the partial or total loss of ATGL activation, leading to the accumulation of TG inside lipid droplets. We aim to assess the clinical and biochemical manifestations, diagnosis, follow-up and genotype–phenotype correlations in six Pakistani pediatric patients with CDS.ResultsSix male patients with mean age 15 months (9–24 months) diagnosed as CDS on the basis of non-bullous ichthyosiform erythroderma, hepatomegaly and Jordans bodies in peripheral smear. We identified two novel mutations in ABHD5 gene (c.338G > T and c.730_731insA). These mutations have a pathogenic and damaging influence on the ABHD5 protein structure and function. During the 2 year clinical follow-up one patient died of severe chest infection; he had severe phenotype. There is no genotype–phenotype correlation in CDS. Therapy with low fat diet, MCT oil, Vit E and ursodeoxycholic acid has promising results in CDS.ConclusionNon-bullous ichthyosiform erythroderma, steatohepatitis and Jordan’s anomaly are consistent findings in all cases of CDS. It is suggested that an accurate diagnosis of CDS should be based on combination of clinical features and pathognomonic ABHD5 mutations. More studies should be carried out to identify population-specific genetic mutations for the rapid and cost-effective diagnosis of CDS.

Reductive Power Generated by Mycobacterium leprae Through Cholesterol Oxidation Contributes to Lipid and ATP Synthesis.

Upon infection, Mycobacterium leprae, an obligate intracellular bacillus, induces accumulation of cholesterol-enriched lipid droplets (LDs) in Schwann cells (SCs). LDs are promptly recruited to M. leprae-containing phagosomes, and inhibition of this process decreases bacterial survival, suggesting that LD recruitment constitutes a mechanism by which host-derived lipids are delivered to intracellular M. leprae. We previously demonstrated that M. leprae has preserved only the capacity to oxidize cholesterol to cholestenone, the first step of the normal cholesterol catabolic pathway. In this study we investigated the biochemical relevance of cholesterol oxidation on bacterial pathogenesis in SCs. Firstly, we showed that M. leprae increases the uptake of LDL-cholesterol by infected SCs. Moreover, fluorescence microscopy analysis revealed a close association between M. leprae and the internalized LDL-cholesterol within the host cell. By using Mycobacterium smegmatis mutant strains complemented with M. leprae genes, we demonstrated that ml1942 coding for 3β-hydroxysteroid dehydrogenase (3β-HSD), but not ml0389 originally annotated as cholesterol oxidase (ChoD), was responsible for the cholesterol oxidation activity detected in M. leprae. The 3β-HSD activity generates the electron donors NADH and NADPH that, respectively, fuel the M. leprae respiratory chain and provide reductive power for the biosynthesis of the dominant bacterial cell wall lipids phthiocerol dimycocerosate (PDIM) and phenolic glycolipid (PGL)-I. Inhibition of M. leprae 3β-HSD activity with the 17β-[N-(2,5-di-t-butylphenyl)carbamoyl]-6-azaandrost-4-en-3one (compound 1), decreased bacterial intracellular survival in SCs. In conclusion, our findings confirm the accumulation of cholesterol in infected SCs and its potential delivery to the intracellular bacterium. Furthermore, we provide strong evidence that cholesterol oxidation is an essential catabolic pathway for M. leprae pathogenicity and point to 3β-HSD as a prime drug target that may be used in combination with current multidrug regimens to shorten leprosy treatment and ameliorate nerve damage.

Temporal correlation of morphological and biochemical changes with the recruitment of different mechanisms of reactive oxygen species formation during human SW872 cell adipogenic differentiation.

Human SW872 preadipocyte conversion to mature adipocytes is associated with time‐dependent changes in differentiation markers' expression and with morphological changes accompanied by the accumulation of lipid droplets (LDs) as well as by increased mitochondriogenesis and mitochondrial membrane potential. Under identical conditions, the formation of reactive oxygen species (ROS) revealed with a general probe was significant at days 3 and 10 of differentiation and bearly detectable at day 6. NADPH oxidase (NOX)‐2 activity determined with an immunocytochemical approach followed a very similar pattern. There was no evidence of mitochondrial ROS (mROS), as detected with a selective fluorescence probe, at days 3 and 6, possibly due to the triggering of the Nrf‐2 antioxidant response. mROS were instead clearly detected at day 10, concomitantly with the accumulation of very large LDs, oxidation of both cardiolipin and thioredoxin 2, and decreased mitochondrial glutathione. In conclusion, the morphological and biochemical changes of differentiating SW872 cells are accompanied by the discontinuous formation of ROS derived from NOX‐2, increasingly implicated in adipogenesis and adipose tissue dysfunction. In addition, mROS formation was significant only in the late phase of differentiation and was associated with mitochondrial dysfunction.

Distinct BTK inhibitors differentially induce apoptosis but similarly suppress chemotaxis and lipid accumulation in mantle cell lymphoma

BackgroundThe more selective second-generation BTK inhibitors (BTKi) Acalabrutinib and Zanubrutinib and the first-generation BTKi Ibrutinib are highlighted by their clinical effectiveness in mantle cell lymphoma (MCL), however, similarities and differences of their biological and molecular effects on anti-survival of MCL cells induced by these BTKi with distinct binding selectivity against BTK remain largely unknown.MethodsAlamarBlue assays were performed to define cytotoxicity of BTKi against MCL cells, Jeko-1 and Mino. Cleaved PARP and caspase-3 levels were examined by immunoblot analysis to study BTKi-induced apoptotic effects. Biological effects of BTKi on MCL-cell chemotaxis and lipid droplet (LD) accumulation were examined in Jeko-1, Mino and primary MCL cells via Transwell and Stimulated Raman scattering imaging analysis respectively. Enzyme-linked immunoassays were used to determine CCL3 and CCL4 levels in MCL-cell culture supernatants. RNA-seq analyses identified BTKi targets which were validated by quantitative RT-PCR (qRT-PCR) and immunoblot analysis.ResultsAcalabrutinib and Zanubrutinib induced moderate apoptosis in Ibrutinib high-sensitive JeKo-1 cells and Ibrutinib low-sensitive Mino cells, which was accompanied by cleaved PARP and caspase-3. Such effects might be caused by the stronger ability of Ibrutinib to upregulate the expression of pro-apoptotic genes, such as HRK, GADD45A, and ATM, in JeKo-1 cells than in Mino cells, and the expression of such apoptotic genes was slightly changed by Acalabrutinib and Zanubrutinib in both JeKo-1 and Mino cells. Further, Acalabrutinib, Zanubrutinib and Ibrutinib reduced MCL-cell chemotaxis with similar efficiency, due to their similar abilities to downmodulate chemokines, such as CCL3 and CCL4. Also, these three BTKi similarly suppressed MCL-cell LD accumulation via downregulating lipogenic factors, DGAT2, SCD, ENPP2 and ACACA without significant differences.ConclusionBTKi demonstrated differential capacities to induce MCL-cell apoptosis due to their distinct capabilities to regulate the expression of apoptosis-related genes, and similar biological and molecular inhibitory effects on MCL-cell chemotaxis and LD accumulation.

\u03b2-catenin-promoted cholesterol metabolism protects against cellular senescence in naked mole-rat cells

The naked mole-rat (NMR; Heterocephalus glaber) exhibits cancer resistance and an exceptionally long lifespan of approximately 30 years, but the mechanism(s) underlying increased longevity in NMRs remains unclear. In the present study, we report unique mechanisms underlying cholesterol metabolism in NMR cells, which may be responsible for their anti-senescent properties. NMR fibroblasts expressed β-catenin abundantly; this high expression was linked to increased accumulation of cholesterol-enriched lipid droplets. Ablation of β-catenin or inhibition of cholesterol synthesis abolished lipid droplet formation and induced senescence-like phenotypes accompanied by increased oxidative stress. β-catenin ablation downregulated apolipoprotein F and the LXR/RXR pathway, which are involved in cholesterol transport and biogenesis. Apolipoprotein F ablation also suppressed lipid droplet accumulation and promoted cellular senescence, indicating that apolipoprotein F mediates β-catenin signaling in NMR cells. Thus, we suggest that β-catenin in NMRs functions to offset senescence by regulating cholesterol metabolism, which may contribute to increased longevity in NMRs.

Olive oil effectively mitigates ovariectomy-induced marrow adiposity assessed by MR spectroscopy in estrogen-deficient rabbits.

Polyphenols in extra virgin olive oil (EVOO) have been found to reduce the expression of PPARγ2, inhibit adipocyte differentiation, and enhance the formation of osteoblasts from bone marrow stem cells. However, the underlying mechanisms of their action remain unknown. To determine the sequential effects of EVOO on marrow fat expansion induced by estrogen deprivation using 3.0-T proton magnetic resonance (MR) spectroscopy in an ovariectomy (OVX) rabbit model of postmenopausal bone loss over a six-month period. A total of 45 female New Zealand rabbits were equally divided into sham-operation, OVX controls, and OVX treated with EVOO for six months. Marrow fat fraction was measured by MR spectroscopy at baseline conditions, and three and six months postoperatively, respectively. Serum bone biomarkers, lumbar and femoral bone mineral density, microtomographic parameters, biomechanical properties, and quantitative parameters of marrow adipocytes were studied. OVX was associated with marrow adiposity in a time-dependent manner, accompanied with increased bone turnover and impaired bone mass and trabecular microarchitecture. In OVX rabbits, EVOO markedly alleviated trabecular bone loss and reduced the accumulation of lipid droplets including adipocyte size, density, and areas of fat deposits in the bone marrow. EVOO prevented such changes in terms of both marrow adiposity and bone remodeling. Early EVOO treatment may exert beneficial effects on bone by modulating marrow adiposity, which would support their protective effect against bone pathologies.

Free fatty acids induce the demethylation of the fructose 1,6-biphosphatase 2 gene promoter and potentiate its expression in hepatocytes.

Obesity is a serious health issue as it is a social burden and the main risk factor for other metabolic diseases. Increasing evidence indicates that a high-fat diet (HFD) is the key factor for the development of obesity, but the key genes and their associated molecular mechanisms are still not fully understood. In this study, we performed integrated bioinformatic analysis and identified that fructose-1,6 biphosphatase 2 (FBP2) was involved in free fatty acids (FFAs)-induced lipid droplet accumulation in hepatocytes and HFD-induced obesity in mice. Our data showed that palmitate (PA) and oleic acid (OA) induced the expression of FBP2 in time- and dose-dependent manners, and accelerated the development of lipid droplets in LO2 human normal liver cells. In HFD-fed C57BL/6 mice, accompanied by insulin resistance and lipid droplet accumulation, the mRNA and protein levels of FBP2 in the livers also increased significantly. The results from the methylation sequencing PCR (MSP) and bisulfite specific PCR (BSP) indicated that PA/OA induced the demethylation of the FBP2 gene promoter in LO2 cells. Moreover, betaine, a methyl donor, attenuated the expression of the FBP2 gene, the accumulation of lipid droplets, and the expression of perilipin-2, a biomarker of lipid droplets, in LO2 cells. All these findings revealed that FBP2 might be involved in HFD-induced obesity, and it is of interest to investigate the role of FBP2 in the treatment and prevention of obesity and its associated complications.

Neutral lipids as early biomarkers of cellular fate: the case of \u03b1-synuclein overexpression

α-synuclein (α-syn) accumulation and aggregation is a common pathological factor found in synucleinopathies, a group of neurodegenerative disorders that includes Parkinson´s disease (PD). It has been proposed that lipid dyshomeostasis is responsible for the occurrence of PD-related processes, however, the precise role of lipids in the onset and progression of neurodegenerative disorders remains unclear. Our aim was to investigate the effect of α-syn overexpression on neutral lipid metabolism and how this impacts on neuronal fate. We found lipid droplet (LD) accumulation in cells overexpressing α-syn to be associated with a rise in triacylglycerol (TAG) and cholesteryl ester (CE) levels. α-syn overexpression promoted diacylglycerol acyltransferase 2 upregulation and acyl-CoA synthetase activation, triggering TAG buildup, that was accompanied by an increase in diacylglycerol acylation. Moreover, the CE increment was associated with higher activity of acyl-CoA:cholesterol acyltransferase. Interestingly, α-syn overexpression increased cholesterol lysosomal accumulation. We observed that sterol regulatory element-binding protein (SREBP)-1 and SREBP-2 were differentially regulated by α-syn overexpression. The latter gave rise to a reduction in SREBP-1 nuclear translocation and consequently in fatty acid synthase expression, whereas it produced an increase in SREBP-2 nuclear localization. Surprisingly, and despite increased cholesterol levels, SREBP-2 downstream genes related to cholesterolgenesis were not upregulated as expected. Notably, phospholipid (PL) levels were diminished in cells overexpressing α-syn. This decrease was related to the activation of phospholipase A2 (PLA2) with a concomitant imbalance of the PL deacylation-acylation cycle. Fatty acids released from PLs by iPLA2 and cPLA2 action were esterified into TAGs, thus promoting a biological response to α-syn overexpression with uncompromised cell viability. When the described steady-state was disturbed under conditions favoring higher levels of α-syn, the response was an enhanced LD accumulation, this imbalance ultimately leading to neuronal death.

PSIX-33 Docosahexaenoic Acid (DHA) but not Eicosapentaenoic Acid (EPA) induces the trans-differentiation of C2C12 into white-like adipocytes phenotype

Abstract Muscle-derived stem cells (MDSCs, or myoblasts) play an important role in myotubes regeneration. However, these cells can differentiate into adipocytes once exposed to EPA and DHA, which are highly suggested during pregnancy. The objective of this study aims at determining the effect of isolated EPA and DHA on C2C12 cells undergoing white and brown adipogenic differentiation. Confluent cultured cells were treated with white and brown adipocyte induction medium (WIM and BIM respectively) with 50µM EPA and 50µM DHA separately. DHA treated groups differentiated into white-like adipocyte by down-regulating the expression of myogenic genes such as MyoD, MyoG, and Mrf4; but promoted white adipocyte marker genes(P < 0.05). Moreover, cells treated with WIM and DHA exhibited a decrease in mitochondrial biogenesis through suppressing PGC1a and TFAM expression (P < 0.05). Also, DHA promoted the expression of lipolysis regulating genes. DHA impaired C2C12 cells browning through reducing the mitochondrial biogenesis by significantly suppressing the expression of COX7a1, PGC1a, and UCP3 genes (P < 0.05). DHA treated groups showed an increased accumulation of lipid droplets and suppressed maximal mitochondrial respiration and spare capacity. EPA treatment reduced myogenesis regulating genes (P < 0.05) but did not affect adipogenic genes (P >0.05). Likewise, EPA suppressed the expression of WAT signature genes (P < 0.05), indicating its antagonism to DHA. EPA and WIM treatment suppressed the expression of TFAM and PGC1a, but did not affect PGC1a protein level. Although mitochondrial biogenic gene expression was reduced in EPA and BIDM treated group, no changes in mitochondrial function were observed. EPA supplementation did not affect the differential route of C2C12 into brown adipocytes. To conclude, EPA and DHA may similarly affect the integrity of muscle tissue, but DHA is a potent adipogenic and lipogenic factor that can change the metabolic profile of the body by increasing intramuscular fat.

The metabolic suppressor 3-iodothyronamine enhances lipolysis in 3T3-L1 adipocytes via activation of the adenosine monophosphate-activated protein kinase/forkhead box O1 signaling pathway.

Obesity is a major health concern in modern societies as it is linked to diverse chronic diseases, such as diabetes, cancer, stroke, and skeletomuscular disorders. This study aimed to investigate the lipolytic potency of the metabolic suppressor 3-iodothyronamine (T1AM) and its molecular mechanism in differentiated 3T3-L1 adipocytes. Cells stained with Oil Red O showed a remarkable accumulation of lipid droplets by 20-d post-differentiation and a plateau at 26 - 30 day. Treatment with 100 μM T1AM for 6 h increased the liberation of free fatty acids (FFAs) and glycerol (P < 0.05) detected in the culture media. However, this stimulatory effect was significantly suppressed by ATGListatin, an inhibitor of adipose triglyceride lipase (ATGL), suggesting that ATGL plays a rate-limiting role in triglyceride (TG) turnover. To understand the lipolytic mechanism, immunoblotting and confocal image analyses of the T1AM-treated and control groups were conducted. The elevated lipolysis was accompanied by increases in the phosphorylation of adenosine monophosphate-activated protein kinase (p-AMPK), nuclear localization of forkhead box O1 (FoxO1), and expression of monoacylglycerol lipase (MGL) protein (P < 0.05). Finally, the treated cells exhibited downregulated expression of acetyl-CoA carboxylase (ACC) relative to p-ACC and increased protein expression of carnitine palmitoyltransferase 1 (CPT1) (P < 0.05). Taken together, T1AM showed lipolytic potency via activation of the AMPK/FoxO1/ATGL/MGL axis for decomposing TGs to FFAs and glycerol and of the AMPK/ACC/CPT1 pathway in facilitating the mobilization of FFAs into the mitochondria, highlighting its therapeutic potential for the treatment of obesity.

Hyperhomocysteinemia and dyslipidemia in point mutation G307S of cystathionine \u03b2-synthase-deficient rabbit generated using CRISPR/Cas9

BackgroundCongenital hyper-homocysteinemia (HHcy) is caused by a defective cystathionine β-synthase (CBS) gene, and is frequently associated with dyslipdemia. The aim of this study was to further elucidate the effect of mutated CBS gene on circulating lipids using a rabbit model harboring a homozygous G307S point mutation in CBS.MethodsCRISPR/Cas9 system was used to edit the CBS gene in rabbit embryos. The founder rabbits were sequenced, and their plasma homocysteine (Hcy) and lipid profile were analyzed.ResultsSix CBS-knockout (CBS-KO) founder lines with biallelic modifications were obtained. Mutation in CBS caused significant growth retardation and high mortality rates within 6 weeks after birth. In addition, the 6-week old CBS-KO rabbits showed higher plasma levels of Hcy, triglycerides (TG), total cholesterol (TC) and low-density lipoprotein cholesterol (LDL-C) compared to the age-matched wild-type (WT) controls. Histological analysis of the mutants showed accumulation of micro-vesicular cytoplasmic lipid droplets in the hepatocytes. However, gastric infusion of vitamin B and betaine complex significantly decreased the plasma levels of TG, TC and LDL-C in the CBS-KO rabbits, and alleviated hepatic steatosis compared to the untreated animals.ConclusionA CBSG307S rabbit model was generated that exhibited severe dyslipidemia when fed on a normal diet, indicating that G307S mutation in the CBS gene is a causative factor for dyslipidemia.

Condensation Enhancement on Hydrophobic Surfaces Using Electrophoretic Method and Hybrid Paint Coating

Condensation heat transfer on stainless steel tube utilizing superhydrophobic coatings was investigated. The electrophoretic deposition and spraying methods were employed to coat the tubes’ outer surface. The mixture that was synthesized for spray coating was a hybrid paint. It incorporated polyurethane matrix and a colloidal suspension containing organic nanoparticles. The hybrid paint had a proper adhesion to the substrate which caused more durability compared to the electrophoretic coating. The agglomeration of nanoparticles in the hybrid paint caused the formation of particles with larger size compared to the particles in the electrophoretic coating. Consequently, contact angle hysteresis over electrophoretic coating was much lesser than spraying method. Dropwise condensation regime was observed over both the coatings as a result of hydrophobicity. Nonetheless, the condensation heat flux declined over the hybrid paint at high subcooling temperatures due to transition to the flooding condensation regime. This was due to the accumulation of ultra-small droplets as a result of high nucleation sites. The continuation of droplets sweeping was hindered by sticking to the substrate. The dropwise condensation was maintained over the electrophoretic coating and the transition to the film-wise condensation happened at larger wall subcooling temperature. The ability of the electrophoretic coating in shedding droplets efficiently was caused by higher hydrophobicity, high roughness density, and minimum droplet pinning.

Aquaporin 7\u2002involved in GINSENOSIDE-RB1-mediated anti-obesity via peroxisome proliferator-activated receptor gamma pathway

BackgroundObesity, characterized by the excessive accumulation of triglycerides in adipocytes and their decreased excretion from adipocytes, is closely related to various health problems. Ginsenoside Rb1 (Rb1), the most active component of the traditional Chinese medicine ginseng, has been reported to have positive effects on lipid metabolism. The aim of the present study was to determine the protective effects of Rb1 on glycolipid metabolism under obesity conditions and its mechanisms and to reveal the signaling pathways involved.MethodsIn our study, male C57BL/6 mice with obesity induced by a high-fat diet (HFD) and mature 3 T3-L1 adipocytes were used to investigate the role of Rb1 in lipid accumulation and explore its possible molecular mechanism in vivo and in vitro, respectively.ResultsRb1 reduced the body weight, fat mass, adipocytes size and serum free fatty acid (FFA) concentration of obese mice. In differentiated 3 T3-L1 adipocytes, Rb1 reduced the accumulation of lipid droplets and stimulated output of triglycerides. Additionally, the expression of peroxisome proliferator-activated receptor gamma (PPARγ), phosphorylated PPARγ (Ser112) and aquaporin 7 (AQP7) was upregulated in adipocytes and adipose tissues upon Rb1 treatment. However, intervention of GW9662, PPARγ antagonist, attenuated Rb1-mediated effects on glycolipid metabolism and AQP7 levels.ConclusionsThese data indicated that Rb1 reduced body weight and improved glycolipid metabolism by upregulating PPARγ and AQP7 protein levels. Our study indicated a potential role for Rb1 in the prevention and treatment of obesity.

Abstract IA16: ER stress responses shape the ovarian cancer immune-microenvironment

Abstract Harnessing the intrinsic ability of our immune system to eliminate malignant cells represents the most promising new anticancer strategy since the development of chemotherapy. While cancer immunotherapy has been designated a scientific breakthrough, tumor-induced immunosuppression represents a major impediment to the success of this new approach in ovarian cancer. We uncovered that ovarian cancer inhibits the development of protective antitumor immunity by inducing a state of persistent “endoplasmic reticulum (ER) stress” in intratumoral dendritic cells (DCs) and T cells. First, we identified that sustained activation of the IRE1-XBP1 arm of the ER stress response in DCs was necessary for the aggressive and accelerated progression of primary and metastatic ovarian cancers in various preclinical models of disease. Mechanistically, intratumoral DCs exhibited constitutive IRE1-XBP1 activation, which was fueled by lipid peroxidation byproducts that caused ER stress by modifying ER-resident chaperones. Persistent XBP1 activation induced a comprehensive triglyceride biosynthetic program promoting accumulation of lipid droplets that subsequently impaired DC antigen presentation to T cells. Conditional deletion or therapeutic silencing of IRE1 or XBP1 in DCs restored their immunogenic function in the tumor and delayed ovarian cancer progression by eliciting protective antitumor T-cell responses. In addition, we recently demonstrated that ovarian cancer also triggers abnormal IRE1-XBP1 activation in T cells to control their mitochondrial function and antitumor activity. XBP1 upregulation in T cells isolated from human ovarian cancer specimens was associated with decreased intratumoral T-cell infiltration and reduced IFNG expression. Ascites fluid from ovarian cancer patients inhibited glucose uptake and caused N-linked protein glycosylation defects in T cells, leading to XBP1-driven mitochondrial dysfunction. Mechanistically, maladaptive XBP1 signaling limited the influx of glutamine necessary to sustain mitochondrial respiration under glucose-deprived conditions. Hence, ovarian cancer hosts lacking XBP1 selectively in T cells demonstrated superior antitumor immunity, delayed malignant progression, and increased overall survival. Taken together, our studies revealed that ovarian cancer provokes persistent IRE1-XBP1 signaling in immune cells to shape the tumor microenvironment and evade immune control. These findings provide a new mechanistic understanding of immunosuppression in ovarian cancer and offer novel avenues for immunotherapeutic intervention. Citation Format: Juan R. Cubillos-Ruiz. ER stress responses shape the ovarian cancer immune-microenvironment [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research; 2019 Sep 13-16, 2019; Atlanta, GA. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(13_Suppl):Abstract nr IA16.

Abstract B095: Tackling chemotherapy resistance of cancer cells: Inhibition of diacylglycerol O-acyltransferase 2 increases ceramide and cancer cell apoptosis

Abstract Cellular stress induced by chemotherapy results in the activation of cell death pathways. The efficacy of chemotherapy can be hindered by the upregulation of proteins that inhibit pro-apoptotic effectors. Ceramide sits at the center of the sphingolipid pathway and Obeid et al demonstrated that exogenous treatment of ceramide induced apoptosis in leukemic cells, whereas cell stressors increase endogenous ceramide levels upstream of caspase activation. The enzyme, diacylglycerol O-acyltransferase 2 (DGAT2), can cause O-acylation of ceramide to 1-O-acylceramide, which is then accumulated in lipid droplets and does not have pro-cell death function. Senkal et al. illustrated that the conversion of ceramide to 1-O-acylceramide causes HCT-116 colon cancer cells to become resistant to chemotherapy 5-fluorouracil (5-FU). Knockdown of the DGAT2 gene showed increased levels of ceramide, decreased levels of 1-O-acylceramides, and sensitized cancer cells to 5FU indicated by a higher caspase 3/7 activation in knockdown cells treated with 5FU than control. Mouse models were employed to demonstrate that a four-week oleate high-fat diet resulted in greater lipid droplet formation and accumulation of acylceramide in the liver versus a control diet, as detected by light chromatography and mass spectrometry (LC-MS). These data demonstrate that the conversion of ceramide to 1-O-acylceramide might act as a survival mechanism and promote chemotherapy resistance and that diet upregulates this process. Thus, we hypothesize that utilizing the commercially available DGAT2 inhibitor (PF-06424439) can increase ceramide in cancer cell lines, hence increase cell death. Treatments of various cancer cell lines, such as MDA-MB-231, MCF-7, and PC3 cells with a chemotherapeutic (cisplatin, doxorubicin, or paclitaxel) were administered alone or paired with PF-06424439 for various time points. Cell viability was quantified via an MTT assay. Oxidoreductase enzymes reduce the MTT dye to insoluble formazan, which exhibits a purple color and represents the reductive activity present in the mitochondria of living cells. Cell death was quantified via lactate dehydrogenase (LDH) release from damaged cells. Greater cell death was observed when both chemotherapy and PF-06424439 were administered in combination versus chemotherapy alone. Ceramide levels were quantified with LC-MS and greater ceramide levels were seen in the combination treatment, hence affirming that ceramide may have a role in mediating cell death. The physiological implications of the combination treatment and factors such as diet will be evaluated via in vivo models in future studies. Citation Format: Rideeta Raquib, Maria J. Hernández-Corbacho, Lina M. Obeid. Tackling chemotherapy resistance of cancer cells: Inhibition of diacylglycerol O-acyltransferase 2 increases ceramide and cancer cell apoptosis [abstract]. In: Proceedings of the Twelfth AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2019 Sep 20-23; San Francisco, CA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2020;29(6 Suppl_2):Abstract nr B095.

Protective effect of adropin against high fat diet-induced obese diabetic wistar rats via nuclear factor erythroid 2-related factor 2 pathway

Introduction: Liver steatosis (fatty liver) is frequently found during the conditions such as diabetes and obesity. The current experimental study was executed the effect of adropin against high fat diet (HFD)-induced obese diabetic Wistar rats via nuclear factor erythroid 2-related factor 2 (Nrf2) pathway. Materials and Methods: Wistar rats were randomly divided into six groups and divided as follows: normal, HFD control, HFD + adropin (2.5, 5 and 10 mg/kg), and HFD + Glibenclamide (2.5 mg/kg), respectively. Lipid and carbohydrate metabolism, hepatic parameters, antioxidant, and proinflammatory cytokines were estimated at the end of the experimental study. Nrf2 transcription and nuclear level were also estimated. In vitro adropin diminished the accumulation of lipid droplets in dose-dependent manner and no effect was observed on the lipolysis. Type II diabetic rats fed with HFD exhibited a marked reduction in hepatic extraction faction and hepatic steatosis after the adropin and glibenclamide treatment. Results: Adropin significantly (P

Hypolipidemic and Hepatoprotective Effects of High-Polydextrose Snack Food on Swiss Albino Mice.

In this study, the hepatoprotective and hypolipidemic effects of high-polydextrose snack food on Swiss albino mice were investigated. The mice were randomly divided into three groups: control diet, high-fat diet, and high-fat and fiber diet groups. Addition of high-polydextrose snack to the high-fat diet resulted in significant reduction in the liver weight, the accumulation of lipid droplets in liver, and the liver damage of hyperlipidemic mice in comparison with the high-fat diet. The high-polydextrose snack also decreased the content of total triglyceride, cholesterol, and low-density lipoprotein cholesterol as well as the alanine aminotransferase and aspartate aminotransferase activities in the mice serum. In addition, the high-polydextrose snack significantly increased the high-density lipoprotein cholesterol content of the hyperlipidemic mice. Consequently, use of high-polydextrose snack generated hepatoprotective and hypolipidemic effects on hyperlipidemic mice.

Western diet induces severe nonalcoholic steatohepatitis, ductular reaction, and hepatic fibrosis in liver CGI-58 knockout mice

Humans and rodents with Comparative Gene Identification-58 (CGI-58) mutations manifest nonalcoholic fatty liver disease (NAFLD). Here we show that liver CGI-58 knockout (LivKO) mice fed a Western diet rapidly develop advanced NAFLD, including nonalcoholic steatohepatitis (NASH) and hepatic fibrosis. After 14 weeks of diet challenge, starting at 6 weeks of age, LivKO mice showed increased inflammatory cell infiltration and proinflammatory gene expression in the liver, which was associated with elevated plasma levels of aminotransferases. Hepatic ductular reactions, pericellular fibrosis, and bridging fibrosis were observed only in the LivKO mice. Consistently, the KO mice had a significant increase in hepatic mRNAs for fibrogenic genes. In addition, LivKO mice displayed massive accumulation of lipid droplets (LDs) in hepatocytes. LDs were also observed in the cholangiocytes of the LivKO mice, but not the floxed controls. Four of the five LD coat proteins, including perilipins 2, 3, 4, and 5, were increased in the CGI-58 KO liver. CRISPR/Cas9-mediated knockout of CGI-58 in Huh7 human hepatoma cells induced LD deposition and perilipin expression, suggesting a cell autonomous effect. Our findings establish the Western diet-fed LivKO mice as an animal model of NASH and hepatic fibrosis. These animals may facilitate preclinical screening of therapeutic agents that counter against NAFLD progression.

Effects of acute exposure to ultra-wideband pulsed electromagnetic fields on the liver and kidneys of mice

ABSTRACTThe biosafety of ultra-wideband (UWB) pulses, which are characterized by simultaneously high power and a high bandwidth ratio, has gained increasing attention. Although there is substantial prior literature on the biological effects of UWB pulses on both cells and animals, an explicit, unequivocal and definite pattern of the corresponding biological responses remains elusive, and the systemic secondary consequences are also still not fully understood. In this study, we found that exposing mice to UWB pulses resulted in the alteration of several biochemical blood parameters, which further prompted us to investigate changes in the liver and kidneys of mice exposed to UWB pulses with different field intensities and different durations. The data demonstrated that exposure to UWB pulses significantly increased the levels of ALT and AST, increased oxidative stress, and could even induce the accumulation of lipid droplets in hepatocytes. The total number of pulses under the tested acute exposure regiment contributed most to the observed hepatic and rental dysfunction. Notably, the physiological and molecular changes recovered approximately 72 hours after exposure. These results imply the potential risk of acute exposure to UWB pulses, and highlight the meaningful targets for further long-term study of chronic exposure.

Assessment of Adrenotoxicity Induced on Prenatal Exposure to Bacterial Endotoxin Lipopolysaccharide: an Age-Related Study in Mice

Preclinical and clinical studies have reported that psychiatric illness stems from exposure to infection during early life. Infection-induced increase in proinflammatory cytokines and further activation of the hypothalamic–pituitary–adrenal (HPA) axis are considered the main factors behind mental illness. The present study elucidated the impairment of HPA axis of female mice prenatally challenged with bacterial endotoxin lipopolysaccharide (LPS). The impairment was evaluated through detailed investigation of adrenal histopathology, the end target endocrine gland of HPA axis, plasma levels of pituitary adrenocorticotropic hormone (ACTH) and corticosterone (CORT). In view of the role of pituitary hormone prolactin (PRL) in modulation of HPA axis under infection, plasma level of PRL was also assessed. The evaluation was done at three different life stages of adulthood, i.e., postnatal day (PND) 85, PND 120 and PND 200. Distinct histopathological alterations in adrenal were revealed: distorted arrangement of zona fasciculata cells, localized cytoplasmic vacuolization and accumulation of lipid droplets were some of the disruptions of the adrenal cortex. Medulla showed distinct vascular congestion and disruptions of chromaffin cells with pyknotic nuclei. Plasma levels of ACTH and CORT were significantly elevated, but PRL was lowered. Bacterial endotoxin LPS might have interfered at different levels of hypothalamus, pituitary and directly on adrenal as well acting through its receptors to induce adrenotoxicity. An age-dependent HPA axis impairment was noted; the persistence of the adrenal toxicity was to a greater degree at PND 200 than PND 120 and PND 85 suggesting the reason for the susceptibility of old age to mental illness.