Women face a higher lifetime risk of developing neurodegenerative diseases such as Alzheimer's disease and related dementias. The menopausal transition, characterized by a decline in estrogen levels, may affect cholesterol metabolism and neurodegenerative processes. Oxysterols, oxidized cholesterol derivatives, play a role in these pathways, with 24(S)-hydroxycholesterol (24HC) reflecting brain cholesterol turnover and 27-hydroxycholesterol (27HC) linked to systemic cholesterol metabolism. We investigated associations of plasma oxysterols with circulating sex hormones and characteristics of reproductive history in 1,974 postmenopausal women with no history of dementia from the Women's Health Initiative, taking into account APOE4 status and cholesterol-lowering medication. We found that higher levels of bioavailable estradiol were associated with higher 24HC and 27HC levels, and higher estrone was associated with higher 24HC (all P values <0.05). Associations of estradiol with 24HC and 27HC were stronger among APOE4 carriers and those not using cholesterol-lowering medication, with a significant interaction between bioavailable estradiol and APOE4 in relation to 27HC (p for interaction = 0.04). Having an older age at menopause was associated with lower 24HC among those taking cholesterol medication (p for interaction = 0.03). Our findings suggest that 24HC and 27HC may be proxy biomarkers of neuronal health and estrogen status in postmenopausal women. The stronger associations between estradiol and oxysterols among APOE4 carriers and those not using cholesterol medication suggest the need to account for hormonal, genetic, and pharmacological factors when evaluating neurodegenerative risk. Longitudinal studies are warranted to further investigate oxysterols as potential early biomarkers of risk for Alzheimer's disease and related dementias.

Hydrophobic bile acid-mediated hepatobiliary injury is a major driver of cholestasis progression. Most anticholestasis treatments being tested clinically are based on a single agent, which does not always sufficiently alleviate bile acid toxicity to slow disease progression. This study investigates a therapeutic strategy of combining glycine-conjugated β-muricholic acid (Gly-βMCA) and fibroblast growth factor-15 (FGF15) to alleviate bile acid hepatobiliary toxicity in Cyp2c70 KO mice that lack endogenous muricholic acid (MCA) synthesis and have a "humanized" hydrophobic bile acid pool composition. The effects of the single and combination treatments on bile acid metabolism, liver injury, and gut microbiome were investigated in female Cyp2c70 KO mice with progressive cholangiopathy and portal fibrosis. While all three treatments significantly reduced biochemical and histologic features of liver injury, the Gly-βMCA and FGF15 combination achieved a remarkably higher reduction in both bile acid pool size and hydrophobicity than either single treatment. Mechanistically, this resulted from synergistically increased biliary hydrophilic MCA species derived from Gly-βMCA, inhibited intestine endogenous bile acid absorption by Gly-βMCA, and repressed cholesterol 7α-hydroxylase (CYP7A1) by FGF15, which counteracted the undesirable farnesoid X receptor antagonism activity of Gly-βMCA. Furthermore, a hydrophobic bile acid pool in Cyp2c70 KO mice was associated with markedly reduced beneficial Lactobacillaceae family bacteria abundance, which was enriched by Gly-βMCA and the combination treatments. In conclusion, the Gly-βMCA and FGF15 combination shows enhanced efficacy in decreasing humanized bile acid pool size and hydrophobicity and holds potential as a therapeutic strategy to decrease bile acid burden in cholestasis.

The relationship between early pregnancy lipid indicators and gestational diabetes mellitus (GDM) or pre-eclampsia (PE) remains incompletely elucidated. This prospective cohort study explored the associations between seven lipid indicators and GDM and PE among 32,411 pregnant participants. The results suggested that triglycerides (TGs), total cholesterols (TCs), and remnant cholesterols (RCs) were positively associated with both GDM and the composite outcome (GDM/PE). For GDM, compared with the lowest quartile, the highest quartile had odds ratios (ORs) of 1.646 (95% confidence interval [95% CI]: 1.363, 1.988) for TGs, 1.654 (95% CI: 1.241, 2.205) for TCs, and 1.396 (95% CI: 1.189, 1.640) for RCs. For GDM/PE, the corresponding ORs in the highest versus lowest quartile were 1.564 (95% CI: 1.302, 1.877) for TGs, 1.655 (95% CI: 1.253, 2.186) for TCs, and 1.379 (95% CI: 1.180, 1.612) for RCs. Non-HDL-C showed a negative association with GDM and GDM/PE, with ORs of 0.833 (95% CI: 0.758, 0.916) and 0.871 (95% CI: 0.795, 0.954), respectively. TG/HDL-C ratio was positively associated with PE, with an OR of 2.451 (95% CI: 1.369, 4.388). The OR values of the second and third quantiles of HDL-C for PE were 1.706 (95% CI: 1.301, 2.238) and 1.598 (95% CI: 1.170, 2.183), respectively. Nonlinear dose-response relationships were observed for most lipids with the outcomes. Additionally, early pregnancy TG, RC, and TG/HDL-C ratio partially mediated the effect of maternal age on all three outcomes (mediated proportion 2-7%). Non-HDL-C mediated the age-PE pathway (1%). This study simultaneously included multiple lipid parameters for systematic analysis, revealing the impact of dyslipidemia on pregnancy outcomes from a more comprehensive perspective and providing richer evidence for exploring related mechanisms and clinical assessment.

Phospholipase D2 (PLD2) plays critical roles in cellular signaling, membrane dynamics, and cancer progression. Oleate (OA) has been shown to activate PLD2 and promote triple-negative breast cancer (TNBC) cell migration, but the underlying molecular mechanisms remain poorly understood. Using confocal microscopy, lipid raft isolation, and S-acylation assays, we show that OA enhanced PLD2 S-acylation at Cys223 and Cys224, disrupting its lipid raft localization, and consequently increasing its colocalization with PIP-enriched microdomains. Furthermore, we identified PLD2 as a guanine nucleotide exchange factor (GEF) for Cdc42, with its GEF activity regulated by OA-dependent S-acylation and lipid raft dynamics. Mutation of the S-acylation sites or disruption of lipid rafts abolished PLD2-mediated Cdc42 activation and filopodia-like cell protrusion formation. These findings reveal a novel regulatory mechanism by which OA modulates PLD2 activity through S-acylation and membrane microdomain reorganization, providing new insights into the regulation of PLD2 in cell migration and signaling.

Mutations in microtubule-associated protein Tau (MAPT), the gene that codes for the protein Tau, cause frontotemporal lobar degeneration (FTLD) with phenotypes ranging from behavioral changes to cognitive impairment and parkinsonism. Recently, lipid changes have been heavily implicated in synucleinopathies and secondary tauopathies such as Alzheimer's Disease (AD). Whether mutations in MAPT or accumulation of hyperphosphorylated Tau (pTau) can contribute to lipid changes in primary tauopathies is unknown. Here, we examine the effect of the FTLD-associated mutation MAPT P301S on brain lipid metabolism in a Tau transgenic mouse model. We find that the MAPT P301S mutation drives increased levels of diglycerides and hexosyl- and lactosylceramides while reducing triglycerides, specifically those triglyceride species containing monounsaturated fatty acids, but does not affect cholesterol metabolism prior to pTau accumulation. Strikingly, with increasing accumulation of pTau, neutral lipids such as cholesteryl esters and triglycerides start to accumulate in the brain of mutant mice, as also reported in the AD and FTLD brain. Furthermore, with increasing buildup of pTau, we observe a decrease in cholesterol synthesis and turnover to 24S-hydroxycholesterol. Overall our data indicates that Tau mutations strongly affect brain lipid metabolism.

Fatty acids (FAs) play multifaceted roles in neurodegenerative diseases (NDDs), including Alzheimer's disease (AD), Parkinson's disease (PD), and amyotrophic lateral sclerosis (ALS). This review systematically summarizes current understanding of fatty acid metabolism and its diverse implications in NDD pathology. Short-chain fatty acids (SCFAs), primarily generated by gut microbiota, regulate neuroinflammation, gut-brain communication, and blood-brain barrier (BBB) integrity via epigenetic modifications and immune modulation. Medium-chain fatty acids (MCFAs) exhibit therapeutic potential by improving energy metabolism and neuromuscular function, particularly in ALS models. Long-chain polyunsaturated fatty acids (PUFAs), notably docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA), contribute to neuronal membrane integrity, synaptic plasticity, and antioxidant defense, mitigating oxidative stress and neuroinflammation. Conversely, saturated and certain n-6 fatty acids may exacerbate neurodegeneration through pro-inflammatory and oxidative pathways. Emerging evidence highlights fatty acid involvement in key pathological processes such as lipid peroxidation, mitochondrial dysfunction, ferroptosis, and BBB disruption. Therapeutically, targeted supplementation, dietary modification, microbiome manipulation, and advanced nanotechnology-based delivery systems are promising strategies. Nevertheless, precise therapeutic efficacy depends critically on disease stage, dosage, genetic background, and individual metabolic context. Integrating personalized medicine with precision nutritional strategies and novel drug-delivery platforms offers promising avenues to translate fatty acid-based interventions into clinical practice, potentially improving patient outcomes in the aging global population.

Voltage-gated potassium (Kv) channels possess distinct voltage-sensor (VSD) and pore (PD) domains, making it challenging to study domain-specific lipid effects. Here, we examined the functional modification of a prototypical Kv channel, KvAP, by phosphatidylglycerol (POPG) and phosphatidylserine (POPS) in mono-component asymmetric bilayers using the contact bubble bilayer (CBB) method. In these membranes, specific chemical modifications were distinguished from non-specific electrostatic (surface potential) effects by using the channel's gating as an intrinsic probe. No specific effects were observed when charged lipids were in the outer leaflet. When present in the inner leaflet, POPS exerted only a single specific effect: the acceleration of activation kinetics. In contrast, inner-leaflet POPG induced multiple, profound modifications: it also accelerated activation kinetics, but additionally shifted the conductance-voltage (G-V) curve hyperpolarized, attenuated the G-V slope, and accelerated inactivation kinetics. This clear contrast allows a domain-specific interpretation: the shared acceleration of activation is attributed to a general kinetic modulation of the VSD, while POPG's unique effects-impaired electromechanical coupling (attenuated slope) and accelerated inactivation-are attributed to specific chemical interactions with the VSD-PD linker and the PD, respectively. These results reveal a multi-site mechanism of lipid modulation dictated by leaflet asymmetry and headgroup chemistry.

n-3 polyunsaturated fatty acids (n-3 PUFAs) possess numerous health benefits. The FAT-1 desaturase in the model organism Caenorhabditis elegans is a Δ15-desaturase that converts n-6 PUFAs into n-3 PUFAs. Transgenic expression of FAT-1 has been used in organisms such as pigs, mice, and fish to improve n-3 PUFA levels. However, the determination of FAT-1 activity and substrate preference per se remains unclear. AlphaFold structure prediction revealed that FAT-1 is an integral membrane protein located in the endoplasmic reticulum, and it consists of four transmembrane helices (TM1-4) with a functional CYTB5 domain in the N-terminus and a desaturase domain containing three histidine-rich sequences (His boxes) in the C-terminus. A small region in the desaturase domain containing amino acids 210-217, especially G212, G216, and S217, are essential for its activity. FAT-1 can convert all four n-6 PUFAs to corresponding or downstream n-3 PUFAs in both C. elegans and mammalian cells, and may prioritize the conversion of C20:4n6 (arachidonic acid, ARA) to C20:5n3 (eicosapentaenoic acid, EPA). These results uncover the significant mechanism of the activity and substrate preference of the FAT-1 desaturase, providing insights into the transgenic application of FAT-1.

Several oxylipins are lipid mediators derived from the oxidation of polyunsaturated fatty acids (PUFAs). The majority of oxylipins in biological samples occurs esterified in neutral lipids (nLs) and phospholipids (PLs). They are commonly quantified indirectly following alkaline hydrolysis providing excellent sensitivity but the information in which lipid classes the oxylipins occurred in is lost. The direct analysis of oxidized lipids is currently not sensitive enough to detect all esterified oxylipins. Here, a new hydrophilic interaction liquid chromatography (HILIC) based lipid class fractionation using solid-phase extraction (SPE) cartridges was developed separating lipids into nLs and 4 PL fractions using a single column. Esterified oxylipins in the fractions were quantified following alkaline hydrolysis to sensitively pinpoint in which lipid classes they are bound in plasma. The fractionation was extensively characterized for different lipid extracts demonstrating high separation efficiency and recovery using labeled standards and untargeted analysis of endogenous lipids. Esterified oxylipins in the fractions were quantitatively detected. Based on the results from two independent human plasma pools including SRM1950 it is shown that: hydroxy-linoleic acid- and hydroxy-α-linolenic acid-derived oxylipins are preferably bound to nLs whereas long chain hydroxy-PUFAs and PUFAs (i.e. ARA EPA and DHA) are predominantly esterified to phospholipid classes. Supplementation of n3-PUFAs for 12 months led to an increase in EPA- and -DHA-derived oxylipins in all lipid fractions with the highest increase of hydroxy-PUFAs in nLs. This demonstrates a precursor PUFA-dependent binding of oxylipins and a direct effect of diet on esterified oxylipins in plasma.

Matrix metalloproteinase-12 (MMP12) is a proinflammatory macrophage-secreted protein with immunomodulatory functions that affects neutrophil infiltration, cytokine release, macrophage recruitment, and proliferation. We have previously demonstrated that the genetic deletion of MMP12 in a cardiometabolic mouse model ameliorates obesity-induced low-grade inflammation, white adipose tissue dysfunction, and atherosclerosis. Based on the various beneficial metabolic effects of MMP-12 deletion, we hypothesized that loss of MMP-12 also positively affects whole-body energy metabolism and/or brown adipose tissue (BAT) function in a cardiometabolic mouse model. To investigate the effects of MMP12 deletion on whole-body energy metabolism and/or BAT function, we used low-density lipoprotein receptor (Ldlr)/Mmp12 double knockout (DKO) fed a high-fat, sucrose- and cholesterol-enriched diet. DKO mice housed at 22°C showed increased energy expenditure and decreased BAT size and triglyceride (TG) content. Untargeted proteomic analyses revealed the upregulation of proteins and pathways related to mitochondrial function, glucose metabolism, and fatty acid oxidation in the BAT of DKO mice, whereas abundance of proteins and pathways associated with inflammation were reduced. In addition, DKO mice exhibited reduced macrophage infiltration in BAT with the infiltrating macrophages showing lower expression of lipid-associated marker genes. Loss of MMP12 was associated with reduced compactness and sphericity of the mitochondria in the BAT. Following an acute cold exposure, DKO mice had decreased circulating lipid concentrations, especially very low-density lipoprotein-TG and LDL-cholesterol, and increased expression of thermogenic genes. We conclude that MMP12 may play a detrimental role in whole-body energy homeostasis and thermogenesis, as it triggers macrophage infiltration, inflammation, and mitochondria dysfunction in BAT.

Hepatocellular carcinoma (HCC) has a higher incidence in males and is a leading cause of cancer-related deaths, which lacks effective therapies and surveillance markers. Using a murine model of bile acid excess (farnesoid X receptor and small heterodimer partner double knockout, DKO), which phenocopies many aspects of HCC, including sex differences, we investigated the links between sex, bile acid metabolism, and microbial composition. Unexpectedly, the increase in the ratios of carcinogenic deoxycholic acid (DCA) was similar between DKO males with HCC and cancer-resistant DKO females. However, both taurine-conjugated and free-cholic acid (CA), sharply increased in the serum of DKO males with free-CA comprising 65% of the bile acid pool, whereas DKO female serum was mostly comprised of conjugated bile acids. Unlike such sex differences in DKO serum composition, conjugated bile acids were predominant in the hepatic BA pool irrespective of the sex or genotype, as expected. Fecal and cecal microbiota - many of which harbor bile acid transformation/ deconjugation capacity- were altered in DKO mice in a sex-specific manner. Untargeted fecal metabolite analysis showed differences in bile acids, phospholipids, and oxidized fatty acids between the genotypes, with DKO females excreting more sulphated and oxidized CA than tumor-bearing DKO males. Further analysis revealed a direct correlation between unconjugated CA levels with the abundance of the microbial genus Faecalibaculum in the DKO HCC model. These findings suggest distinct sex-specific changes in cecal and fecal microbiota, and BA composition may be leveraged in combination as a potential tool for HCC surveillance.

Understanding the processes and/or the key factors involved in the formation as well as degradation of lipid droplets (LDs) within the adipocytes is of immense importance, especially in the context of health, obesity, cancer, and other diseases. While cold temperature and/or menthol (an edible cooling agent), seem to have diverse and confounding effects on obesity and/or thermogenesis, so far there is no direct evidence that specific pharmacological modulation of the Transient Receptor Potential cation channel subfamily Melastatin member 8 (TRPM8), a cold-temperature-activated ion channel, can indeed affect LD status within the mature adipocytes. Here, we used highly specific antagonists and agonists of TRPM8 to modulate TRPM8 in cultured adipocyte cells in vitro and monitored the expression of TRPM8 as well as other adipogenic functions. Our results indicate that specific activation of TRPM8 in mature adipocytes by a specific agonist, that is, WS12 ((1R∗,2S∗)-N-(4-methoxyphenyl)-5-methyl-2-(1-methylethyl)cyclohexanecarboxamide), results in increased expression of PPARγ protein. However, TRPM8 inhibition by N-(3-aminopropyl)-2-[(3-methylphenyl)methoxy]-N-(2-thienylmethyl)benzamidehydrochloride results in no change in the PPARγ expression, yet causes decreased Oil Red O intensity, a reduction in LD sizes, and an increase in LD numbers. BODIPY (4,4-difluoro-1,3,5,7,8-pentamethyl-4-bora-3a,4a-diaza-s-indacene) labeling in live cells also suggests the same findings. Altogether, data suggest that in the absence of any confounding factors, specific inhibition of TRPM8 results in either less fusion of LDs or enhanced fragmentation of LDs in vitro. These findings may have broad implications in the field of adipogenesis and in cancer.

Hydroxysteroid 17β dehydrogenase 11 (HSD17β11) is a member of the 17β-HSD family with canonical roles in steroid metabolism. Given its predominant localisation on lipid droplets, we investigated HSD17β11's role in lipid metabolism. In patients with metabolic dysfunction-associated fatty liver disease (MASLD), liver HSD17β11 levels are reduced, correlating with liver steatosis severity. HSD17β11 deletion in human cell lines increases lipid droplet size and number. This is associated with triglyceride accumulation due to impaired lipolysis and increased fatty acid uptake. Mechanistically, HSD17β11 facilitates the interaction between PLIN5 and ATGL, enabling efficient protein kinase A (PKA)-stimulated lipolysis. Surprisingly, Hsd17β11 deletion did not affect liver lipid metabolism or MASLD development in lean or obese mice. These findings demonstrate that while HSD17β11 is crucial for efficient PKA-mediated lipolysis in human cells, its deficiency appears redundant for lipid metabolism in mice.

Nonshivering thermogenesis plays an important role in maintaining body temperature and energy homeostasis. Elongation of very long-chain fatty acid protein 3 (ELOVL3), which catalyzes the synthesis of C20-C24 fatty acids, is induced in brown adipose tissue (BAT) by cold exposure and regarded as a thermogenic gene. However, its precise role in thermogenesis remains elusive. BAT-specific KO mice of Elovl3 were generated by the Cre/LoxP approach and phenotypically analyzed under cold exposure. Gene expression changes in BAT were characterized by quantitative RT-PCR and Western blotting, BAT remodeling was evaluated by histological examination, and lipid composition was examined by lipidomic analysis. BAT-specific deletion of the Elovl3 gene resulted in cold intolerance because of impaired BAT thermogenesis, without a significant effect on muscle shivering thermogenesis. Mechanistically, Elovl3 deficiency impaired cold-induced BAT remodeling and Ucp1 expression, with a defect in mitochondrial cristae remodeling. Lipidomics analysis showed a marked reduction in the contents of lysophosphatidylcholine, cardiolipin, and acylcarnitine in BAT in the absence of Elovl3. Taken together, our findings reveal the critical role of ELOVL3 in BAT thermogenesis and provide new ideas for the intervention and treatment of obesity-related diseases.

The mammary gland synthesizes and secretes nutrient-rich milk containing lactose, protein, and lipids, with the complex assortment of lipids providing more than half of the energy and bioactive factors that impact the growth and development of neonates. The birth of neonates initiates the lipogenic capacity of the mammary gland with upregulation in expression of lipogenic genes, including Stearoyl-CoA desaturase (Scd1). SCD1 plays a critical role in lipogenesis, catalyzing the conversion of saturated fatty acids to monounsaturated fatty acids. Previous studies of Scd1 knockout mice revealed that SCD1 impacts several metabolic processes in the liver and adipose tissue, including fat synthesis. However, the role of SCD1 in lactation is not fully understood. Our study aimed to determine the role of SCD1 in lactation and the effects of maternal knockout of Scd1 on the growth of the lactating neonates. We employed second-parity Scd1-deficient female mice (n=7) that we compared with wild-type mice (n=6). To determine lipid and metabolic alterations, mammary gland and milk samples were harvested on lactation day 10. Relative to wild-type mice, mammary gland weight, alveolar area, and milk glycerolipid content were reduced in lactating Scd1 deficient mice. Scd1 deficiency also diminished mammary gland biosynthetic metabolic pathways, such as glycerolipid and phospholipid synthesis, while enhancing catabolic pathways, such as the oxidation of fatty acids. Neonates nursed by Scd1 deficient mice exhibited lower body weights. These findings highlight the critical role of SCD1 in orchestrating metabolic adaptations during lactation to ensure adequate milk synthesis to support the rapidly growing neonates.

Astrocytes comprise approximately 40% of CNS cells and have pivotal roles in brain functions. Under steady-state conditions, astrocytes maintain homeostasis in the CNS through the uptake or release of neurotransmitters. However, in neurodegenerative conditions, astrocytes are activated by inflammatory cytokines, such as interleukin-1alpha (IL-1α) and TNF-α, which are released from activated microglia. Activated astrocytes release several inflammatory cytokines and neurotoxic substances, resulting in neuronal injury. Sphingolipids are a series of bioactive lipids involved in several biological processes, such as apoptosis, inflammatory response, cell cycle, and immune response. SM is a sphingolipid that is a major component of the cellular membrane and is also involved in inflammatory responses. We report that SM promotes IL-1α/TNF-α-induced expressions of representative astrocyte mRNAs and astrocyte activation through the NF-κB pathway. In contrast, reduction of SM by knockdown of sphingomyelin synthase 1 (SMS1) and/or SMS2 suppresses astrocyte activation. Furthermore, removal of SM by the blockade of ceramide transfer protein suppresses astrocyte activation via the induction of histone deacetylase (HDAC) 1 and HDAC3; subsequently, the levels of acetylated p65 (Lys 310) are reduced, leading to the suppression of the NF-κB pathway. Our findings further the understanding of the regulation of astrocyte activation by sphingolipids.

Polyethylene glycol (PEG) is widely used in liposome formulation due to its blocking properties and ability to prolong circulation in vivo, to create biomimetic liposomes and drug delivery devices. Similarly, membrane-embedded DNA nanotechnology is increasingly used to modulate cellular behaviour and communication. However, there is a gap in knowledge in how PEG-lipid formulations can be optimised for both liposome properties and control of selective DNA hybridisation. To address this, we systematically investigated the effect of liposome PEG content on DNA mediated tethering of liposomes to glass surfaces. We formulated liposomes of two different lipid compositions (DOPE/DOPC or DPhPC), with varying amounts of PEGylated lipid (0-50%). We measured the effect of increased PEG content on liposome size and polydispersity through dynamic light scattering (DLS). Small amounts of PEG (0-20%) introduced repulsive forces that reduced size, while large amounts of PEG (30-50%) increased polydispersity. PEG-liposomes were then decorated with cholesterol-DNA strands and labelled with either intercalating lipid dyes or fluorescently labelled lipids. Binding to surfaces via complementary DNA strands was quantified using total internal reflection fluorescence (TIRF) microscopy. We found that PEGylation of DNA-liposomes could either block or enhance surface binding, depending on the amount of PEG. DNA-liposomes with reduced surface binding included DPhPC/DiD with 10% or 20% PEG-lipid. In contrast, DNA-liposome surface binding increased for DOPE/DOPC/DiD with increasing PEG%. This study highlights that while PEG can act to stabilise liposome formulations, its ability to block specific DNA binding interactions on membranes is variable and dependent on membrane composition.

S-palmitoylation is a dynamic and reversible post-translational modification that plays crucial roles in cancer progression. Here, we found the oncogene of metadherin (MTDH) modulates lipid metabolism and ferroptosis by its S-palmitoylation. We demonstrate that MTDH is S-palmitoylated at Cys-75 in the endoplasmic reticulum by ZDHHC1/9 and S-depalmitoylated by APT1. The flexible loop and the α-helix length in the MTDH N-terminus affect its S-palmitoylation level. In addition, metabolomics analysis found that the S-palmitoylated MTDH increases intracellular levels of triglycerides, phosphatidylethanolamines, and phosphatidylcholines. However, the non-S-palmitoylation form of MTDH-CS enhanced the interaction of between MTDH and the ferroptosis enhancer of Acyl-CoA synthetase long-chain family member 4 (ACSL4), thereby reducing ferroptosis sensitivity in breast cancer cell. Taken together, targeting MTDH S-palmitoylation may represent a novel strategy for breast cancer therapy.

Early placentation relies on temporal changes in intrauterine oxygen tension that regulate trophoblast differentiation events. Studies have highlighted the contribution of bioactive sphingolipids to the pathogenesis of placental disorders, characterized by hypoxia. However, it is unknown whether placental sphingolipid metabolism changes during the switch from a hypoxic to an oxygenated environment in the first trimester of gestation and if sustained hypoxia is causative of sphingolipid alterations seen in preeclampsia. Herein, we performed sphingolipid analysis of first-trimester human placentae as well as placentae from conditional (placenta-specific) Phd2 knockout mice (Phd2 cKO) that exhibit preeclampsia-like features, including placental hypoxia. Analysis revealed elevated long chain ceramide (Cer16:0, Cer18:0, Cer20:0 and Cer22:0) and reduced sphingosine-1-phosphate (So-1-P) content in Phd2 cKO placentae. Expression of key regulatory sphingolipid enzymes, acid ceramidase (ASAH1) and sphingosine kinase 1 (SPHK1), was reduced in Phd2 cKO placentae, while that of alkaline ceramidase ACER2 remained unchanged. Human placentae from 5-9 weeks of gestation, when intrauterine oxygen tension is low, exhibited heightened long chain ceramide (Cer14:0, Cer16:0, Cer18:0, Cer 18:1) and sphingosine content and reduced ASAH1 and SPHK1 expression, highlighting the relevance of low oxygen in regulating sphingolipid metabolism under physiological (placental development) and pathological (Phd2 cKO induced preeclampsia) conditions. Ultrastructural analyses of early (5-9 weeks) human and murine Phd2 cKO placentae revealed that increased trophoblast mitochondrial fission events accompanied elevated ceramide. Together, the data support the concept that a chronic low-oxygen environment leads to placental ceramide buildup, which may alter mitochondrial homeostasis and potentially contribute to cell death events characteristic of preeclampsia.