Lipid Metabolism Pathways Research Articles

#752 Ponatinib exacerbates renal damage in MRL/lpr lupus model mice through the lipid metabolism pathway

Abstract Background and Aims The role of tyrosine kinase inhibitors (TKIs) in lupus nephritis is controversial. Ponatinib is a third-generation tyrosine kinase inhibitor that targets multiple receptors. This study aims to explore the effect of ponatinib in lupus nephritis, with the hope of providing insights for the clinical application of ponatinib. Method At 8 weeks of age, mice were randomly divided into four groups. One group received no treatment, while the other three groups were respectively administered Ponatinib, prednisolone acetate, and a combination of Ponatinib and prednisolone acetate. The treatment was concluded when the mice reached 16 weeks of age. ELISA was employed to measure the levels of anti-dsDNA IgG, ANA, and C3 in mouse serum. DEGs and DELs were identified using the DESeq2 package in R. Pathway and gene ontology enrichment analyses were performed using the MetaCore database. Results Compared to the negative control, those mice receiving ponatinib showed a significant increase in the urinary protein/creatinine ratio (P < 0.01), along with notable elevations in blood creatinine and blood urea nitrogen. Pathological examination of kidney tissue suggested predominant acute lesions, including transparent thrombi, cellulose-like necrosis, cellular/fibrous crescents, and interstitial inflammation. Interestingly, serum testing in MRL/lpr mice revealed no significant differences in lupus activity-related indicators, including autoantibodies ANA, double-stranded DNA, and complement C3, between the negative control group and the ponatinib group. Furthermore, it was discovered that the kidney damage caused by ponatinib in MRL/lpr mice could be reversed by prednisone. RNA-Seq of kidney tissue from MRL/lpr mice showed a total of 44 differentially expressed genes when comparing the negative control with mice receiving ponatinib, including 23 upregulated and 21 downregulated genes. Pathway analysis revealed that the top four pathways were all related to lipid metabolism. Conclusion Ponatinib significantly exacerbated kidney damage in MRL/lpr mice and did not induce changes in lupus activity.

#1320 Differential microRNA expression profile in obesity-induced kidney disease driven by high-fat diet in mice

Abstract Background and Aims Obesity is one of the main causes of chronic kidney disease; however, the precise molecular mechanisms leading to the onset of kidney dysfunction in obesity-induced kidney disease (OIKD) remain unclear. MicroRNAs (miRNAs) have been implicated in the development and progression of obesity, and several candidates have been identified as critical regulators of adipogenesis and lipid metabolism, while their dysregulation contribute to obesity-related metabolic abnormalities. The aim of the present study was to decipher the kidney microRNA (miRNA) expression profile in a mouse model of obesity-induced kidney disease characterized by systemic metabolic disturbances and kidney injury that closely resemble those observed in human disease. Method Male C57BL/6J mice were randomly divided and placed either a standard diet (STD) or a high fat diet (HFD) for 10 weeks. Serum triglycerides and total, HDL and LDL cholesterol were determined by standard clinical methods. In the kidney sections, histopathological assessments were done by using PAS and Oil-Red-O staining's. Expression of renal MCP1, TNFα, α-SMA, Fibronectin, miR-146a, miR-551b and miR-5099 was assessed by real time PCR. In order to elucidate the role of differentially expressed miRNAs, a search for target genes was done using an online computational resource DIANA-miRPath v4.0. Results High fat diet (HFD)-induced obesity led to evident structural alterations in tubular and glomerular regions of the kidney, increased renal expression of adiposity-related proinflammatory and profibrotic genes, as well as an elevated renal expression of genes involved in cellular lipid metabolism. The miRNA sequencing analysis identified a set of 9 miRNAs differentially expressed in the kidney upon HFD feeding, with miR-5099, miR-551b-3p, miR-223-3p, miR-146a-3p and miR-21a-3p showing most significant differential expression between standard diet (STD) and HFD mice. Consistently, Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) enrichment analyses revealed that 3 validated miRNAs (miR-5099, miR-551b-3p and miR-146a-3p) modulated target genes involved in metabolic and adipocytokine pathways, fatty acid and lipid metabolism, inflammatory, senescence and profibrotic pathways. Conclusion Our results propose that differentially expressed miRNAs play pivotal roles in the intricated pathophysiology of obesity-associated kidney disease and could potentially create novel treatment strategies to counteract the deleterious effects of obesity on kidney function.

#911 A mice model of SHPT with bone abnormalities in CKD and potential functional role of sphingolipid in renal osteodystrophy

Abstract Background and Aims Renal osteodystrophy (ROD) is the skeletal disease of chronic kidney disease associated mineral and bone disorder (CKD-MBD). ROD is characterized by abnormal bone mineralization and impaired bone remodeling, leading to increased risk of fractures and mortality in patients with CKD. To date, the pathology of ROD remains poorly understood due to the lack of a suitable animal model. Hence, we successfully established a stable mouse model of ROD using an optimized adenine diet and explored the possible signaling based on RNA-seq expression analyses. Method Blood biochemical analysis, micro-computed tomography (micro-CT) and bone histomorphometry were performed to analyze the skeletal characteristics. RNA sequencing (RNA-seq) was used to investigate the mechanisms involved in ROD. The functions of differentially expressed genes (DEGs) were evaluated by Gene Ontology (GO) analyses, Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses and Gene Set Enrichment Analyses (GSEA). DEGs were validated by quantitative real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR). Results The mice with ROD induced by optimized adenine diet showed severe abnormalities in serum creatinine (Scr) and blood urea nitrogen (BUN) along with marked hyperparathyroidism and hyperphosphatemia. The bone mineral density (BMD) of femurs was significantly lower in the ROD mice than that in the control (CTL) mice. The trabecular and cortical bone parameters of femurs significantly showed bone loss and severe cortical porosity. ROD mice showed high bone turnover with increased osteoid, eroded, osteoblast, and osteoclast perimeters as well as increased bone formation rate and mineral apposition rate. Transcriptomic profiling identified 1286 genes were upregulated in the femurs of ROD mice compared to the CTL mice and 333 genes that were downregulated. GO, KEGG and GSEA pathway analyses indicated that these genes were highly enriched in functions related to lipid metabolism pathway, sphingolipid signaling pathway, among others. The differential expression of alkaline ceramidase 1(Acer1), ceramide synthase 4 (Cers4) sphingomyelin phosphodiesterase 3 (Smpd3), sphingosine-1-phosphate phosphatase 2 (Sgpp2) in skeletal tissue and serum of CKD patients were validated by qRT-PCR. Since sphingolipid pathways have been recently implicated in bone metabolism, these genes are potential target genes in the settings of ROD. Conclusion In conclusion, our findings demonstrated that the CKD mice with bone abnormalities induced by optimized adenine diet may serve as a useful model for skeletal analysis in ROD. Based on the RNA-seq, we identified that sphingolipid might play a crucial role in the pathogenesis of ROD and provided new therapeutic targets for future study.

Effects of dietary lipid levels on growth, lipid metabolism, fatty acid composition and antioxidant capacity of juvenile greasyback shrimp (Metapenaeus ensis)

The optimal dietary lipid requirement and underlying lipid metabolic mechanisms remain elusive in greasyback shrimp (Metapenaeus ensis), presenting significant obstacles to the advancement of their culture development. Five isonitrogenous diets (38.39% crude protein) containing lipid levels of 2.25%, 4.72%, 7.50%, 10.18%, and 12.71% were formulated to evaluate the effects of dietary lipid levels on growth, lipid metabolism, fatty acid composition and antioxidant capacity of juvenile greasyback shrimp. The results indicated that shrimps fed a diet with 7.50% lipid exhibited significantly higher percent weight gain (161.36%) and specific growth rate (1.72%/d) than those fed other diets (P < 0.05). As the dietary fat content increased, the lipid levels of the whole shrimp, muscle, hepatopancreas, and serum exhibited similar elevations. Concurrently, H&E staining clearly revealed the occurrence of hepatocyte swelling and hepatocyte vacuolization in the 10.18% and 12.71% groups, and hepatic lipid accumulation was further observed in these groups by Oil red O staining. In addition, with the increase in dietary fat levels, saturated fatty acids (SFA) decreased and polyunsaturated fatty acids (PUFA) increased in both hepatopancreas and muscle. The expression of genes involved in lipogenesis (fas, acc and dgat) was initially up-regulated and subsequently down-regulated with increasing dietary lipid levels, whereas the expression trends of lipolysis genes (hsl, atgl) remained similar. The mRNA levels of genes associated with long-chain PUFA biosynthesis (elovl5, lxrβ, srebp1) and fatty acid oxidation (cpt1, acox1) in the hepatopancreas of shrimp fed a diet with 7.50% lipid were significantly elevated compared to those in other dietary groups (P < 0.05). With the increase in dietary fat levels, the expression of fatty acid transport genes fatp4 and fabp1 was up-regulated. Antioxidant enzyme activities in the hepatopancreas initially increased with dietary lipid levels, peaking in shrimp fed the 7.50% lipid diet, but subsequently declined. In conclusion, optimal dietary fat levels enhance growth performance, while high lipid levels impede growth, decrease antioxidant enzyme activities, and regulate lipid deposition in greasyback shrimp by modulating lipid metabolic pathways.

The Effect of Ovariectomy and Estradiol Substitution on the Metabolic Parameters and Transcriptomic Profile of Adipose Tissue in a Prediabetic Model.

Menopause brings about profound physiological changes, including the acceleration of insulin resistance and other abnormalities, in which adipose tissue can play a significant role. This study analyzed the effect of ovariectomy and estradiol substitution on the metabolic parameters and transcriptomic profile of adipose tissue in prediabetic females of hereditary hypertriglyceridemic rats (HHTgs). The HHTgs underwent ovariectomy (OVX) or sham surgery (SHAM), and half of the OVX group received 17β-estradiol (OVX+E2) post-surgery. Ovariectomy resulted in weight gain, an impaired glucose tolerance, ectopic triglyceride (TG) deposition, and insulin resistance exemplified by impaired glycogenesis and lipogenesis. Estradiol alleviated some of the disorders associated with ovariectomy; in particular, it improved insulin sensitivity and reduced TG deposition. A transcriptomic analysis of perimetrial adipose tissue revealed 809 differentially expressed transcripts in the OVX vs. SHAM groups, mostly pertaining to the regulation of lipid and glucose metabolism, and oxidative stress. Estradiol substitution affected 1049 transcripts with overrepresentation in the signaling pathways of lipid metabolism. The principal component and hierarchical clustering analyses of transcriptome shifts corroborated the metabolic data, showing a closer resemblance between the OVX+E2 and SHAM groups compared to the OVX group. Changes in the adipose tissue transcriptome may contribute to metabolic abnormalities accompanying ovariectomy-induced menopause in HHTg females. Estradiol substitution may partially mitigate some of these disorders.

Hyperlipidemia-induced hematopoiesis is repressed by MLKL in endothelial cells of the splenic niche.

Dysregulation of the hematopoietic niche during hyperlipidemia facilitates pathologic leukocyte production, driving atherogenesis. Although definitive hematopoiesis occurs primarily in the bone marrow, during atherosclerosis this also occurs in the spleen. Cells of the bone marrow niche, particularly endothelial cells, have been studied in atherosclerosis, although little is known about how splenic endothelial cells respond to the atherogenic environment. Here we show unique dysregulated pathways in splenic compared to bone marrow endothelial cells during atherosclerosis, including perturbations of lipid metabolism and endocytic trafficking pathways. As part of this response, we identify the mixed lineage kinase domain-like (MLKL) protein as a repressor of splenic, but not bone marrow, myelopoiesis. Silencing MLKL in splenic endothelial cells results in inefficient endosomal trafficking and lipid accumulation, ultimately promoting the production of myeloid cells that participate in plaque development. These studies identify endocytic trafficking by MLKL as a key mechanism of splenic endothelial cell maintenance, splenic hematopoiesis and, subsequently, atherosclerosis.

Role of metabolic dysfunction and inflammation along the liver-brain axis in animal models with obesity-induced neurodegeneration.

The interaction between metabolic dysfunction and inflammation is central to the development of neurodegenerative diseases such as Alzheimer's disease and Parkinson's disease. Obesity-related conditions like type 2 diabetes and non-alcoholic fatty liver disease exacerbate this relationship. Peripheral lipid accumulation, particularly in the liver, initiates a cascade of inflammatory processes that extend to the brain, influencing critical metabolic regulatory regions. Ceramide and palmitate, key lipid components, along with lipid transporters lipocalin-2 and apolipoprotein E, contribute to neuroinflammation by disrupting blood-brain barrier integrity and promoting gliosis. Peripheral insulin resistance further exacerbates brain insulin resistance and neuroinflammation. Preclinical interventions targeting peripheral lipid metabolism and insulin signaling pathways have shown promise in reducing neuroinflammation in animal models. However, translating these findings to clinical practice requires further investigation into human subjects. In conclusion, metabolic dysfunction, peripheral inflammation, and insulin resistance are integral to neuroinflammation and neurodegeneration. Understanding these complex mechanisms holds potential for identifying novel therapeutic targets and improving outcomes for neurodegenerative diseases.

Differential Associations of Erythrocyte Membrane Saturated Fatty Acids with Glycemic and Lipid Metabolic Markers in a Chinese Population: A Cross-Sectional Study.

Mounting evidence indicates a complex link between circulating saturated fatty acids (SFAs) and cardiovascular disease (CVD) risk factors, but research on erythrocyte membrane SFA associations with metabolic markers remains limited. Our study sought to investigate the correlations between erythrocyte membrane SFAs and key metabolic markers within glycemic and lipid metabolism in a Chinese population of 798 residents aged 41 to 71 from Guangzhou. Using gas chromatography-mass spectrometry, we assessed the erythrocyte membrane saturated fatty acid profile and performed multiple linear regression to evaluate the relationship between different SFA subtypes and metabolic markers. Our findings revealed that the odd-chain SFA group (C15:0 + C17:0) exhibited negative associations with fasting blood glucose (FBG), homeostatic model assessment for insulin resistance (HOMA-IR), and triglycerides (TG). Conversely, the very-long-chain SFA group (C20:0 + C22:0 + C23:0 + C24:0) exhibited positive associations with fasting insulins (FINS), HOMA-IR, total cholesterol (TC), and low-density lipoprotein cholesterol (LDL-C). Furthermore, there was no evidence supporting an association between the even-chain group (C14:0 + C16:0 + C18:0) and metabolic markers. Our findings suggest that different subtypes of SFAs have diverse effects on glycemic and lipid metabolic markers, with odd-chain SFAs associated with a lower metabolic risk. However, the results concerning the correlations between even-chain SFAs and very-long-chain SFAs with markers of glycemic and lipid metabolism pathways are confusing, highlighting the necessity for further exploration and investigation.

Comparative analysis of circulating metabolomic profiles identifies shared metabolic alterations across distinct multistressor military training exercises.

Military training provides insight into metabolic responses under unique physiological demands that can be comprehensively characterized by global metabolomic profiling to identify potential strategies for improving performance. This study identified shared changes in metabolomic profiles across three distinct military training exercises, varying in magnitude and type of stress. Blood samples collected before and after three real or simulated military training exercises were analyzed using the same untargeted metabolomic profiling platform. Exercises included a 2-wk survival training course (ST, n = 36), a 4-day cross-country ski march arctic training (AT, n = 24), and a 28-day controlled diet- and exercise-induced energy deficit (CED, n = 26). Log2-fold changes of greater than ±1 in 191, 121, and 64 metabolites were identified in the ST, AT, and CED datasets, respectively. Most metabolite changes were within the lipid (57-63%) and amino acid metabolism (18-19%) pathways and changes in 87 were shared across studies. The largest and most consistent increases in shared metabolites were found in the acylcarnitine, fatty acid, ketone, and glutathione metabolism pathways, whereas the largest decreases were in the diacylglycerol and urea cycle metabolism pathways. Multiple shared metabolites were consistently correlated with biomarkers of inflammation, tissue damage, and anabolic hormones across studies. These three studies of real and simulated military training revealed overlapping alterations in metabolomic profiles despite differences in environment and the stressors involved. Consistent changes in metabolites related to lipid metabolism, ketogenesis, and oxidative stress suggest a potential common metabolomic signature associated with inflammation, tissue damage, and suppression of anabolic signaling that may characterize the unique physiological demands of military training.NEW & NOTEWORTHY The extent to which metabolomic responses are shared across diverse military training environments is unknown. Global metabolomic profiling across three distinct military training exercises identified shared metabolic responses with the largest changes observed for metabolites related to fatty acids, acylcarnitines, ketone metabolism, and oxidative stress. These changes also correlated with alterations in markers of tissue damage, inflammation, and anabolic signaling and comprise a potential common metabolomic signature underlying the unique physiological demands of military training.

Identification of potential hub genes and regulatory networks of smoking-related endothelial dysfunction in atherosclerosis using bioinformatics analysis.

Endothelial dysfunction, the earliest stage of atherosclerosis, can be caused by smoking, but its molecular mechanism requires further investigation. This study aimed to use bioinformatics analysis to identify potential mechanisms involved in smoking-related atherosclerotic endothelial dysfunction. The transcriptome data used for this bioinformatics analysis were obtained from the Gene Expression Omnibus (GEO) database. The GSE137578 and GSE141136 datasets were used to identify common differentially expressed genes (co-DEGs) in endothelial cells treated with oxidized low-density lipoprotein (ox-LDL) and tobacco. The co-DEGs were annotated using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomics (KEGG) databases. Additionally, a protein-protein interaction (PPI) network was constructed to visualize their interactions and screen for hub genes. GSE120521 dataset was used to verify the expression of hub genes in unstable plaques. The miRNA expression profile GSE137580 and online databases (starBase 2.0, TargetScan 8.0 and DGIdb v4.2.0) were used to predict the related non-coding RNAs and drugs. A total of 232 co-DEGs were identified, including 113 up-regulated genes and 119 down-regulated genes. These DEGs were primarily enriched in detrimental autophagy, cell death, transcription factors, and cytokines, and were implicated in ferroptosis, abnormal lipid metabolism, inflammation, and oxidative stress pathways. Ten hub genes were screened from the constructed PPI network, including up-regulated genes such as FOS, HMOX1, SQSTM1, PTGS2, ATF3, DDIT3, and down-regulated genes MCM4, KIF15, UHRF1, and CCL2. Importantly, HMOX1 was further up-regulated in unstable plaques (p= 0.034). Finally, a regulatory network involving lncRNA/circRNA-miRNA-hub genes and drug-hub genes was established. Atherosclerotic endothelial dysfunction is associated with smoking-induced injury. Through bioinformatics analysis, we identified potential mechanisms and provided potential therapeutic targets.

Accumulation of Lipid Droplets Induced by Listeria monocytogenes in Macrophages: Implications for Survival and Evasion of Innate Immunity.

Listeriosis, caused by Listeria monocytogenes (L.m.), poses a significant public health concern as one of the most severe foodborne diseases. The pathogenesis of L.m. involves critical steps such as phagosome rupture and escape upon internalization. Throughout infection, L.m. influences various host processes, including lipid metabolism pathways, yet the role of lipid droplets (LDs) remains unclear. Here, we reported a rapid, time-dependent increase in LD formation in macrophages induced by L.m. LD biogenesis was found to be dependent on L.m. viability and virulence genes, particularly on the activity of the pore-forming protein listeriolysin O (LLO). The prevention of LD formation by inhibiting diacylglycerol O-acyltransferase 1 (DGAT1) and cytosolic phospholipase A2 (cPLA2) significantly reduced intracellular bacterial survival, impaired prostaglandin E2 (PGE2) synthesis, and decreased IL-10 production. Additionally, inhibiting LD formation led to increased levels of TNF-α and IFN-β. Collectively, our data suggest a role for LDs in promoting L.m. cell survival and evasion within macrophages.

Metabolomics in Osteoarthritis Knee: A Systematic Review of Literature.

Osteoarthritis (OA) is a common degenerative disorder of the synovial joints and is usually an age-related disease that occurs due to continuous wear and tear of the cartilage in the joints. Presently, there is no proven medical management to halt the progression of the disease in the early stages. The purpose of our systematic review is to analyze the possible metabolites and metabolic pathways that are specifically involved in OA pathogenesis and early treatment of the disease. The articles were collected from PubMed, Cochrane, Google Scholar, Embase, and Scopus databases. "Knee", "Osteoarthritis", "Proteomics", "Lipidomics", "Metabolomics", "Metabolic Methods", and metabolic* were employed for finding the articles. Only original articles with human or animal OA models with healthy controls were included. From the initial screening, a total of 458 articles were identified from the 5 research databases. From these, 297 articles were selected in the end for screening, of which 53 papers were selected for full-text screening. Finally, 50 articles were taken for the review based on body fluid: 6 urine studies, 15 plasma studies, 16 synovial fluid studies, 11 serum studies, 4 joint tissue studies, and 1 fecal study. Many metabolites were found to be elevated in OA. Some of these metabolites can be used to stage the OA Three pathways that were found to be commonly involved are the TCA cycle, the glycolytic pathway, and the lipid metabolism. All these studies showed a vast array of metabolites and metabolic pathways associated with OA. Metabolites like lysophospholipids, phospholipids, arginine, BCCA, and histidine were identified as potential biomarkers of OA but a definite association was not identified, Three pathways (glycolytic pathway, TCA cycle, and lipid metabolic pathways) have been found as highly significant in OA pathogenesis. These metabolic pathways could provide novel therapeutic targets for the prevention and progression of the disease. The online version contains supplementary material available at 10.1007/s43465-024-01169-5.

Long-term thermal stress induces hepatic injury and alters the thermotolerance response in Hong Kong catfish (Clarias fuscus)

Global climate change has a significant impact on fish survival and the aquaculture industry. Even highly adaptable organisms like Siluriformes fish are susceptible to these effects. This study investigates the consequences of extreme high temperature on Hong Kong catfish (Clarias fuscus), a species known for its long-term habitation in tropical and subtropical regions. In this study, C. fuscus were cultivated for 90 days at high temperature (HT, 34 °C) and normal temperature (NT, 26 °C), followed by subjecting the two groups to high temperature stress (34 °C) and temperature recovery (26 °C). The hepatic histology, biochemical, and transcriptomic characteristics of the species were examined before acute high temperature stress (NT-C, HT-C), after acute high temperature stress (NT-T, HT-T), and after temperature recovery (NT-R, HT-R). The histological analysis revealed that long-term heat stress damaged the liver tissue of C. fuscus. Furthermore, significant damage was observed in the liver tissues of the NT group under acute high temperature stress, while no further damage was observed in the HT group. The biochemical analysis showed that 90 days of heat stress altered the body's immune and oxidative balance, resulting in an increase in alkaline phosphatase (ALP), alanine aminotransferase (ALT), catalase (CAT), superoxide dismutase (SOD) and glutathione peroxidase (GPX) activities, and a decrease in malondialdehyde (MDA) content. Upon acute heat stress, the liver function of the NT group was disordered and unable to produce enough total-bilirubin (T-BIL), while the aspartate aminotransferase (AST) activity of the HT group was more susceptible to being affected. Long-term heat stress led to significant changes in gene expression, with the NT group showing three times more differentially expressed genes (DEGs) identified in the liver compared to the HT group under acute heat stress conditions. Analysis of DEG enrichment during the same treatment period in both groups revealed significant differences in amino acid metabolism and lipid metabolism pathways. These results suggest that long-term heat stress caused liver tissue damage in C. fuscus, altering immune and oxidative balance, enzyme activity sensitivity, as well as the body's response to acute heat stress pathways and intensity. This study lays the groundwork for future investigations into fish adaptability to sudden temperature changes and aquaculture strategies to mitigate heat stress in fish facing extreme temperatures.

Gefitinib metabolism-related lncRNAs for the prediction of prognosis, tumor microenvironment and drug sensitivity in lung adenocarcinoma

The complete compound of gefitinib is effective in the treatment of lung adenocarcinoma. However, the effect on lung adenocarcinoma (LUAD) during its catabolism has not yet been elucidated. We carried out this study to examine the predictive value of gefitinib metabolism-related long noncoding RNAs (GMLncs) in LUAD patients. To filter GMLncs and create a prognostic model, we employed Pearson correlation, Lasso, univariate Cox, and multivariate Cox analysis. We combined risk scores and clinical features to create nomograms for better application in clinical settings. According to the constructed prognostic model, we performed GO/KEGG and GSEA enrichment analysis, tumor immune microenvironment analysis, immune evasion and immunotherapy analysis, somatic cell mutation analysis, drug sensitivity analysis, IMvigor210 immunotherapy validation, stem cell index analysis and real-time quantitative PCR (RT-qPCR) analysis. We built a predictive model with 9 GMLncs, which showed good predictive performance in validation and training sets. The calibration curve demonstrated excellent agreement between the expected and observed survival rates, for which the predictive performance was better than that of the nomogram without a risk score. The metabolism of gefitinib is related to the cytochrome P450 pathway and lipid metabolism pathway, and may be one of the causes of gefitinib resistance, according to analyses from the Gene Set Enrichment Analysis (GSEA), Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG). Immunological evasion and immunotherapy analysis revealed that the likelihood of immune evasion increased with risk score. Tumor microenvironment analysis found most immune cells at higher concentrations in the low-risk group. Drug sensitivity analysis found 23 sensitive drugs. Twenty-one of these drugs exhibited heightened sensitivity in the high-risk group. RT-qPCR analysis validated the characteristics of 9 GMlncs. The predictive model and nomogram that we constructed have good application value in evaluating the prognosis of patients and guiding clinical treatment.

RAR-related orphan receptor alpha and the staggerer mice: a fine molecular story.

The retinoic acid-related orphan receptor alpha (RORα) protein first came into the limelight due to a set of staggerer mice, discovered at the Jackson Laboratories in the United States of America by Sidman, Lane, and Dickie (1962) and genetically deciphered by Hamilton etal. in 1996. These staggerer mice exhibited cerebellar defects, an ataxic gait, a stagger along with several other developmental abnormalities, compensatory mechanisms, and, most importantly, a deletion of 160 kilobases (kb), encompassing the RORα ligand binding domain (LBD). The discovery of the staggerer mice and the subsequent discovery of a loss of the LBD within the RORα gene of these mice at the genetic level clearly indicated that RORα's LBD played a crucial role in patterning during embryogenesis. Moreover, a chance study by Roffler-Tarlov and Sidman (1978) noted reduced concentrations of glutamic acid levels in the staggerer mice, indicating a possible role for the essence of a nutritionally balanced diet. The sequential organisation of the building blocks of intact genes, requires the nucleotide bases of deoxyribonucleic acid (DNA): purines and pyrimidines, both of which are synthesized, upon a constant supply of glutamine, an amino acid fortified in a balanced diet and a byproduct of the carbohydrate and lipid metabolic pathways. A nutritionally balanced diet, along with a metabolic "enzymatic machinery" devoid of mutations/aberrations, was essential in the uninterrupted transcription of RORα during embryogenesis. In addition to the above, following translation, a ligand-responsive RORα acts as a "molecular circadian regulator" during embryogenesis and not only is expressed selectively and differentially, but also promotes differential activity depending on the anatomical and pathological site of its expression. RORα is highly expressed in the central nervous system (CNS) and the endocrine organs. Additionally, RORα and the clock genes are core components of the circadian rhythmicity, with the expression of RORα fluctuating in a night-day-night sigmoidal pattern and undoubtedly serves as an endocrine-like, albeit "molecular-circadian regulator". Melatonin, a circadian hormone, along with tri-iodothyronine and some steroid hormones are known to regulate RORα-mediated molecular activity, with each of these hormones themselves being regulated rhythmically by the hypothalamic-pituitary axis (HPA). The HPA regulates the circadian rhythm and cyclical release of hormones, in a self-regulatory feedback loop. Irregular sleep-wake patterns affect circadian rhythmicity and the ability of the immune system to withstand infections. The staggerer mice with their thinner bones, an altered skeletal musculature, an aberrant metabolic profile, the ataxic gait and an underdeveloped cerebellar cortex; exhibited compensatory mechanisms, that not only allowed the survival of the staggerer mice, but also enhanced protection from microbial invasions and resistance to high-fat-diet induced obesity. This review has been compiled in its present form, more than 14years later after a chromatin immunoprecipitation (ChIP) cloning and sequencing methodology helped me identify signal transducer and activator oftranscription 5 (STAT5) target sequences, one of which was mapped to the firstintron of the RORα gene. The 599-base-long sequence containing one consensus TTCNNNGAA (TTCN3GAA) gamma-activated sequence (GAS) and five other non-consensus TTN5AA sequences had been identified from the clones isolated from the STAT5 target sites (fragments) in human phytohemagglutinin-activated CD8+ T lymphocytes, during my doctoral studies between 2006 and 2009. Most importantly, preliminary studies noted a unique RORα expression profile, during a time-course study on the ribonucleic acid (RNA), extracted from human phytohemagglutinin (PHA) activated CD8+ T lymphocytes stimulated with interleukin-2 (IL-2). This review mainly focuses on the "staggerer mice" with one of its first roles materialising during embryogenesis, a molecular-endocrine mediated circadian-like regulatory process.

Abstract PO3-24-04: Altered epigenetic modifications and genome architecture reshape lipid metabolism during secondary trastuzumab resistance formation of HER2-positive breast cancer

Abstract Title Altered epigenetic modifications and genome architecture reshape lipid metabolism during secondary trastuzumab resistance formation of HER2-positive breast cancer Background Secondary trastuzumab resistance seriously affects the treatment of HER2-positive breast cancer. Although studies have demonstrated several key cellular properties that are closely accompanied with trastuzumab resistance formation, we still have poor knowledge on the genetic and epigenetic variations that promote such transformation. Here we suggested that the epigenetic modification changes on histone H3 together with altered chromatin architecture promote lipid metabolism reprogramming during secondary trastuzumab formation. Materials and methods Induced secondary trastuzumab-resistant SKBR3_HR cell line together with the original trastuzumab-sensitive SKBR3 cell line were applied in this study. Total RNA was collected for transcriptome analysis. Anti-H3K4me3 and K27me3 antibodies were chosen for CUT&amp;Tag sequencing library preparation. Total genome DNA was prepared for Micro-C sequencing library preparation. Activity score of metabolism pathway was calculated as relative gene expression value averaged over all genes in this pathway in certain cell types. Results Upregulation of arachidonic acid metabolism and downregulation of unsaturated fatty acid synthesis, which are mainly characterized by the activation of PTGS1 and PTGES genes and the repression of FASN and SCD genes, respectively, were observed during the formation of secondary trastuzumab resistance from SKBR3 cell to SKBR3_HR cells. Variations of H3k4me3 instead of H3k27me3 regulate the expression of these 4 genes. The accumulation of H3k4me3 was detected at the promoters of PTGS1 and PTGES genes while they were removed from the promoters of FASN and SCD genes in SKBR3_HR cells. More intra-chromosomal interactions were constituted during resistance formation. In detail, 4626 and 4394 topological associated domains, 3125 and 5824 DNA loops were founded in SKBR3 and SKBR3_HR cells, respectively. Furthermore, the lost and gained DNA loops between SCD and PTGS1 genes and distant genome regions may indicate the weaken and strengthen interactions with transcriptional regulators located there. Conclusions During trastuzumab resistance formation, altered histone modifications as well as higher genome structure could regulate the expression of key genes in lipid metabolism pathways, which may further affect cell properties and interactions with cells in tumor microenvironment. Citation Format: Ningjun Duan, Yijia Hua, Yongmei Yin. Altered epigenetic modifications and genome architecture reshape lipid metabolism during secondary trastuzumab resistance formation of HER2-positive breast cancer [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO3-24-04.

Abstract PO4-14-08: A gene expression-based classifier for HER2-low breast cancer

Abstract Background: The definition of HER2-low breast cancer from clinical trials of antibody-conjugated drugs (ADCs) relies on immunohistochemistry scoring (IHC). However, in daily-practice the accuracy of IHC is hindered by inter-observer variability in assessing HER2 non-overexpressing status. Here, we aimed to identify breast cancer primary tumors with low HER2 expression by leveraging gene expression profiling. Materials and methods: A discovery approach was applied to gene expression profile of internal INT1 (n= 125) and INT2 (n= 84) datasets. We identified specific differently expressed genes (DEGs) according to HER2 IHC categories 0, 1+, 2+ and 3+. Principal Component Analysis (PCA) was used to generate a HER2-low signature whose performance was confirmed in the independent INT3 (n= 95), and TCGA and GSE20194 publicly available datasets. The association between the HER2-low signature and HER2 IHC categories was evaluated by non-parametric Kruskal-Wallis (KW) test with post hoc pair wise comparisons (i.e. contrasts); the HER2-low signature discriminatory capability was assessed by estimating the area under the receiver operating characteristic (ROC) curve (AUC) with its corresponding 95% Confidence Interval (CI). Gene Ontology and KEGG analyses were performed to enrich the DEGs for functional information. Results: A 20-gene HER2-low signature, consisting of both up-regulated (n=11) and down-regulated (n=9) genes, was computed based on the DEGs according to HER2 IHC categories as follows: 11 genes specific for the 1+ category, 8 genes for the 2+ category, and 1 gene for the HER2-low category. HER2-low signature genes were significantly enriched with lipid and steroid metabolism pathways, peptidase regulation, and humoral immune response. The HER2-low signature levels demonstrated a significant association with HER2 IHC categories (KW p-value &amp;lt; 0.001) and were distributed in a bell-shaped pattern across IHC categories (low values 0 and 3+; high values 1+ and 2+), effectively distinguishing HER2-low from 0 (contrast p-value &amp;lt; 0.001) and 3+ (contrast p-value &amp;lt; 0.001). Notably, the signature levels were significantly higher in tumors scored with 1+ as compared to 0 (contrast p =0.002). The HER2-low signature association with IHC categories and discriminatory capability was confirmed in the independent INT3 and TCGA datasets with higher values in HER2-low compared to both 0 and 3+. The HER2-low signature achieved an AUC value of 0.72 (95%CI 0.62-0.83) in differentiating HER2 0/3+ from HER2 1+/2+ categories, which is worth noting in light of the individual ERBB2 mRNA AUC value of 0.48 (95%CI 0.34-0.62). Conclusions: The 20-gene HER2-low signature was generated by maximizing the differences in gene expression between tumors with different HER2 status according to IHC. In contrast to the previously published single ERBB2 gene expression assessment, our data presents compelling evidence for effectively distinguishing HER2-low tumors, including those scored as 1+ from HER2-0 tumors. Our signature holds potential in selecting novel candidates for ADC therapy. Citation Format: Serena Di Cosimo, Sara Pizzamiglio, Chiara Maura Ciniselli, Valeria Duroni, Vera Cappelletti, Loris de Cecco, Maria Carmen De Santis, Rosaria Orlandi, Marilena Iorio, Marco Silvestri, Giancarlo Pruneri, Paolo Verderio. A gene expression-based classifier for HER2-low breast cancer [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO4-14-08.

Abstract PO1-14-06: Integrated analysis reveals the impact of obesity on triple-negative breast cancer

Abstract Background: Obesity and overweight status, which has been growing rapidly over the past few decades, is considered as a risk factor for many types of cancers including breast cancer. Despite the multi-omics profile of triple-negative breast cancer (TNBC) has been comprehensively characterized, the impact of obesity on molecular features of TNBC is not fully appreciated. Methods: We applied an integrative analysis on clinicopathological data and molecular data (including genomic, transcriptomic, proteomic and metabolomic profiling) using the multi-omics database of TNBC (N = 465) from Fudan University Shanghai Cancer Center (FUSCC) for associations with patient body mass index (BMI). Patients were categorized into overweight/obese (OW/OB, BMI ≥ 24 kg/m2) and normal (NL, BMI &amp;lt; 24 kg/m2) group according to the Chinese criteria of BMI. The clinical and molecular differences between OW/OB and NL patients were systematically explored. We also constructed high-fat diet (HFD)-induced obese mouse tumor models and used single-cell RNA sequencing to investigate the impact of obesity on the tumor microenvironment. Furthermore, we analyzed the efficacy of anti-PD-1 immunotherapy on TNBC tumors in both obese and normal mice. Results: OW/OB patients exhibited higher proportion of metabolic syndrome, more adipose tissue in the breast and worse survival than NL patients. Among most frequently mutated genes, OBSCN showed statistically significantly less mutated in the OW/OB group (3.2% vs 9.6%), while TP53 (68.3% vs 76.9%) and PIK3CA (21.4% vs 14.1%) had tendency to be different. In terms of copy number alterations, we found OW/OB patients had a higher amplified or gained frequency of 13q14.11 (FOXO1) and a lower frequency of deletion or loss of chromosomal region 7p22.1 (FOXK1). We further dissect the expression profile of TNBC. Differentially expressed gene analysis and pathway enrichment analysis demonstrated that immune and metabolic pathways were the major distinction between OW/OB and NL tumors. OW/OB tumors were characterized with elevated inflammation of tumor microenvironment, as well as higher expression of immune checkpoints. Moreover, analyses focusing on metabolic heterogeneity using transcriptomic, proteomic and metabolomic data revealed upregulation of lipid metabolism and reactive oxygen species pathway in OW/OB group. In addition, our in vivo experiments demonstrated that TNBC in the obese mice displayed faster growth rates. Flow cytometry analysis and single-cell RNA sequencing showed that higher proportion of immunosuppressive myeloid cells and exhausted CD8+ T cells and upregulation of lipid metabolism in HFD group. Applying anti-PD-1 immunotherapy in both obese and normal mice displayed that tumors in the obese mice showed more sensitive to anti-PD-1 immunotherapy. Conclusion: Our study systematically revealed that obesity might play a significant role in the molecular heterogeneity of TNBC and showed distinct sensitivities to immunotherapy, which should be taken in account in the field of precision medicine. Keywords: triple-negative breast cancer, obesity, immune, tumor microenvironment, metabolism Citation Format: Yue Gong, Peng Ji, Huai-liang Wu, Li-Hua He, Ming-Liang Jin, Xin Hu, Yi-Zhou Jiang, Zhiming Shao. Integrated analysis reveals the impact of obesity on triple-negative breast cancer [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO1-14-06.

Lipid Identification of Biomarkers in Esophageal Squamous Cell Carcinoma by Lipidomic Analysis

Lipids participate in many important biological functions through energy storage, membrane structure stabilization, signal transduction, and molecular recognition. Previous studies have shown that patients with esophageal squamous cell carcinoma (ESCC) have abnormal lipid metabolism. However, studies characterizing lipid metabolism in ESCC patients through lipidomics are limited. Plasma lipid profiles of 65 ESCC patients and 42 healthy controls (HC) were characterized by lipidomics-based ultraperformance liquid chromatography quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS). Single-factor and multi-factor statistical analysis were used to screen the differences in blood lipids between groups, and combined with component ratio analysis and receiver operating characteristic (ROC) curve diagnostic efficiency assessment, to reveal the potential mechanisms and biomarkers of ESCC. There were significant differences in lipid profiles between the ESCC and HC groups. Thirty-six differential lipids (11 up-regulated and 25 down-regulated) were selected based on the criteria of p < .05 and fold change > 1.3 or < 0.77. Glycerophospholipids were the major differential lipids, suggesting that these lipid metabolic pathways exhibit a significant imbalance that may contribute to the development of esophageal squamous cell carcinoma. Among them, the seven candidate biomarkers for esophageal squamous cell carcinoma with the highest diagnostic value are three phosphatidylserine (PS), three fatty acids (FA) and one phosphatidylcholine (PC).

Dietary starch level regulates the growth and glycolipid metabolism through the insulin-PI3K-AKT signaling pathway of juvenile mud crab (Scylla paramamosain)

A six-week feeding trial was conducted to investigate the effects of dietary starch levels on growth performance, glucose and lipid metabolism and insulin signaling pathway of juvenile Scylla paramamosain. Three isonitrogenous (41.51% ± 0.31%) and isolipidic (8.68% ± 0.53%) experimental diets were formulated to contain 10%, 20% and 30% starch levels, respectively. A total 96 crabs were divided into 3 treatment groups. There were 4 replicates per treatment group and 8 crabs per replicate (initial weight 5.12 ± 0.05 g). The crabs fed the diet with 20% starch showed the best growth performance, with higher weight gain rate (WGR), specific growth rate (SGR), protein efficiency ratio (PER), and feed efficiency (FE) compared to the other diets. Increasing starch levels in the diet resulted in higher levels of glycogen and lipid in the muscles and hepatopancreas. The levels of glucose (GLU), triglyceride (TG), and free fatty acid (FFA) displayed a significant increase in both hemolymph and hepatopancreas with the escalating dietary starch concentrations. In contrast, as the dietary starch levels increased, the concentrations of crustacean hyperglycemic hormone (CHH) and insulin-like peptides (ILPs) in hemolymph and hepatopancreas demonstrated a notable decline. The glycolysis and pentose phosphorylation pathways were significantly promoted, while the gluconeogenic pathway was significantly inhibited, with dietary starch levels increasing from 10% to 30%. Additionally, increasing dietary starch levels led to a significant down-regulation of genes related to the lipid synthesis, lipolysis and insulin signaling pathway. Histological results showed that hepatopancreas fat accumulation increased with the increase of dietary starch level. Overall, the findings of this study suggest that adding 20% starch to the diet can promote growth and improve nutrient utilization. In addition, the level of dietary starch had significant effects on the glucose and lipid metabolism and insulin signaling pathway of mud crabs, thus affecting the deposition of lipid reserves in hepatopancreas.