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Procollagen C-endopeptidase enhancer 2, known as PCPE2 or PCOC2 (gene name, PCOLCE2) is a glycoprotein that resides in the extracellular matrix, and is similar in domain organization to PCPE1/PCPE, PCOC1 (PCOLCE1/PCOLCE). Due to the many similarities between the two related proteins, PCPE2 has been assumed to have biological functions similar to PCPE. PCPE is a well-established enhancer of procollagen processing activating the enzyme, BMP-1. However, reports show that PCPE2 has a strikingly different tissue expression profile compared to PCPE. With that in mind and given the paucity of published studies on PCPE2, this review examines the current literature citing PCPE2 and its association with specific cell types and signaling pathways. Additionally, this review will present a brief history of PCPE2's discovery, highlighting structural and functional similarities and differences compared to PCPE. Considering the widespread use of RNA sequencing techniques to examine associations between cell-specific gene expression and disease states, we will show that PCPE2 is repeatedly found as a differentially regulated gene (DEG) significantly associated with a number of cellular processes, well beyond the scope of procollagen fibril processing., Competing Interests: Conflict of interests The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Introduction: Cancer is the leading cause of death in Norway. In this nationwide study we describe the number and causes of hospital admissions and treatment in the final year of life for patients who died of cancer, as well as the associations to age and socioeconomic status (SES)., Materials and Methods: From nationwide registries covering 2010-2014, we identified all patients who were diagnosed with cancer 12-60 months before death and had cancer as their reported cause of death. We examined the number of overnight hospital stays, causes of admission, and treatment (chemotherapy, radiotherapy, surgical procedures) offered during the last year of life by individual (age, sex, comorbidity), cancer (type, stage, months since diagnosis), and socioeconomic variables (co-residential status, income, education)., Results: The analytical sample included 17,669 patients; 8,247 (47%) were female, mean age was 71.7 years (standard deviation 13.7). At diagnosis, 31% had metastatic disease, while 29% had an intermediate or high comorbidity burden. Altogether, 94% were hospitalized during their final year, 82% at least twice, and 33% six times or more. Patients spent a median of 23 days in hospital (interquartile range 11-41), and altogether 38% died there. Younger age, bladder and ovarian cancer, not living alone, and higher income were associated with having ≥6 hospitalizations. Cancer-related diagnoses were the main causes of hospitalizations (65%), followed by infections (11%). Around 51% had ≥1 chemotherapy episode, with large variations according to patient age and SES; patients who were younger, did not live alone, had high education, and high income received more chemotherapy. Radiotherapy was received by 15% and declined with age, and the variation according to SES characteristics was minor. Of the 12,940 patients with a cancer type where surgery is a main treatment modality, only 835 (6%) underwent surgical procedures for their primary tumor in the last year of life., Discussion: Most patients who die of cancer are hospitalized multiple times during the last year of life. Hospitalizations and treatment decline with advancing age. Living alone and having low income is associated with fewer hospitalizations and less chemotherapy treatment. Whether this indicates over- or undertreatment across various groups warrants further exploration., Competing Interests: Declaration of Competing Interest Authors report no conflicts of interest., (Copyright © 2023 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Introduction: Specialized palliative care (SPC) is beneficial towards end of life because of its holistic approach to improve quality of life and comfort of patients and their families. Few studies have described how patient age, sex, comorbidities, and socioeconomic status (SES) are associated with SPC use in nonselective populations who die of cancer. This study aimed to evaluate the use of SPC in the year preceding death by all Norwegian individuals with a recent cancer diagnosis who died of cancer., Materials and Methods: From nationwide registries, we identified patients with a recent (<5 years) cancer diagnosis who died during 2010-2014. Using binary logistic regression models, we estimated the probability of receiving hospital-based SPC during the last year of life according to individual (age, sex, comorbidity), cancer (stage, type, and months since diagnosis), and SES (e.g., living alone, household income, and education) characteristics., Results: The analytical sample contained 45,521 patients with a median age at death of 75 years; 46% were women. The probability of receiving hospital-based SPC in the total cohort was 0.43 (95% confidence interval [CI] 0.42-0.43). Use of SPC was higher if patients were younger, female, had limited comorbidity, metastatic disease, had one the following cancer types: colorectal, pancreatic, bladder, kidney, or gastric, were diagnosed more than six months before death, and had higher SES. Adjusted model results suggested that the probability of using SPC in the last year of life for patients aged 80-89 years was 0.31 (95% CI 0.30-0.32), compared to a probability of 0.63 (95% CI 0.61-0.65) for patients aged 50-59 years. For patients ≥90 years, the probability was 0.16 (95% CI 0.15-0.18)., Discussion: Less hospital-based SPC use among older patients, males, and those with lower SES indicates possible under-treatment in these groups. Future studies should be designed to determine the underlying reasons for these observed differences., Competing Interests: Declaration of Competing Interest Authors report no conflicts of interest., (Copyright © 2022 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Cellular membranes are not homogenous mixtures of proteins; rather, they are segregated into microdomains on the basis of preferential association between specific lipids and proteins. These microdomains, called lipid rafts, are well known for their role in receptor signaling on the plasma membrane (PM) and are essential to such cellular functions as signal transduction and spatial organization of the PM. A number of disease states, including atherosclerosis and other cardiovascular disorders, may be caused by dysfunctional maintenance of lipid rafts. Lipid rafts do not occur only in the PM but also have been found in intracellular membranes and extracellular vesicles (EVs). Here, we focus on discussing newly discovered functions of lipid rafts and microdomains in intracellular membranes, including lipid and protein trafficking from the ER, Golgi bodies, and endosomes to the PM, and we examine lipid raft involvement in the production and composition of EVs. Because lipid rafts are small and transient, visualization remains challenging. Future work with advanced techniques will continue to expand our knowledge about the roles of lipid rafts in cellular functioning., (Copyright © 2020 © 2009 ASBMB. Currently published by Elsevier Inc; originally published by American Society for Biochemistry and Molecular Biology. Published by Elsevier Inc. All rights reserved.)

Background: Commonly consumed mushrooms, portobello (PBM) and shiitake (SHM), are abundant in nutrients, soluble dietary fibers, and bioactive compounds that have been implicated as beneficial in reducing inflammation, improving lipid profiles, and ameliorating heart disease and atherosclerosis, an inflammatory disease of the arteries., Objective: The aim of this study was to determine effects of PBM and SHM in preventing atherosclerosis and associated inflammation in an animal model., Methods: Four-week-old Ldlr-/- male mice were divided into 5 dietary groups for 16 wk: a low-fat control (LF-C, 11 kcal% fat), high-fat control (HF-C, 18.9 kcal% fat), HF + 10% (wt:wt) PBM (HF-PBM, 19.5 kcal% fat) or SHM (HF-SHM, 19.7 kcal% fat) powder, and HF + mushroom control mix (MIX-C, 19.6 kcal% fat), a diet best matched to the average macronutrient content of both mushrooms. Body composition was measured using MRI. Aortic tricuspid valves and aortas were collected and stained to quantify plaque formation. Adhesion molecule expression was quantified by immunohistochemistry. Plasma lipid and cytokine concentrations were measured., Results: We found that mice fed a HF-SHM diet had ∼86% smaller aortic lesion area than mice in both HF-C (P < 0.01) and MIX-C (P < 0.01) groups and also expressed 31-48% lower vascular cell adhesion molecule-1 levels (P < 0.05) than all other groups. Similarly, HF-PBM-fed mice displayed a 70% reduction in aortic lesion area in the tricuspid valve only (P < 0.05). Both mushroom-fed groups had lower weight gain and fat mass (P < 0.05) than the control groups., Conclusion: These results suggest that consumption of PBMs and particularly SHMs is effective in preventing development of high-fat diet-induced atherosclerosis in Ldlr-/- mice. Future studies will determine active components in mushrooms responsible for this beneficial effect., (Copyright © American Society for Nutrition 2019.)

Polarization of macrophages to proinflammatory M1 and to antiinflammatory alternatively activated M2 states has physiological implications in the development of experimental hypertension and other pathological conditions. 12/15-Lipoxygenase (12/15-LO) and its enzymatic products 12(S)- and 15(S)-hydroxyeicosatetraenoic acid (HETE) are essential in the process since disruption of the gene encoding 12/15-LO renders the mice unsusceptible to hypertension. The objective was to test the hypothesis that M2 macrophages catabolize 12(S)-HETE into products that are incapable of promoting vasoconstriction. Cultured M2 macrophages metabolized externally added [ 14 C]12(S)-HETE into more polar metabolites, while M1 macrophages had little effect on the catabolism. The major metabolites were identified by mass spectrometry as (ω-1)-hydroxylation and β-oxidation products. The conversion was inhibited by both peroxisomal β-oxidation inhibitor, thioridazine, and cytochrome P450 inhibitors. Quantitative PCR analysis confirmed that several cytochrome P450 enzymes (CYP2E1 and CYP1B1) and peroxisomal β-oxidation markers were upregulated upon M2 polarization. The identified 12,19-dihydroxy-5,8,10,14-eicosatetraenoic acid and 8-hydroxy-6,10-hexadecadienoic acid metabolites were tested on abdominal aortic rings for biological activity. While 12(S)-HETE enhanced vasoconstrictions to angiotensin II from 15% to 25%, the metabolites did not. These results indicate that M2, but not M1, macrophages degrade 12(S)-HETE into products that no longer enhance the angiotensin II-induced vascular constriction, supporting a possible antihypertensive role of M2 macrophages., (Copyright © 2018 by the American Society for Biochemistry and Molecular Biology, Inc.)

HDL normally transports about 50-70% of plasma sphingosine 1-phosphate (S1P), and the S1P in HDL reportedly mediates several HDL-associated biological effects and signaling pathways. The HDL receptor, SR-BI, as well as the cell surface receptors for S1P (S1PRs) may be involved partially and/or completely in these HDL-induced processes. Here we investigate the nature of the HDL-stimulated interaction between the HDL receptor, SR-BI, and S1PR1 using a protein-fragment complementation assay and confocal microscopy. In both primary rat aortic vascular smooth muscle cells and HEK293 cells, the S1P content in HDL particles increased intracellular calcium concentration, which was mediated by S1PR1. Mechanistic studies performed in HEK293 cells showed that incubation of cells with HDL led to an increase in the physical interaction between the SR-BI and S1PR1 receptors that mainly occurred on the plasma membrane. Model recombinant HDL (rHDL) particles formed in vitro with S1P incorporated into the particle initiated the internalization of S1PR1, whereas rHDL without supplemented S1P did not, suggesting that S1P transported in HDL can selectively activate S1PR1. In conclusion, these data suggest that S1P in HDL stimulates the transient interaction between SR-BI and S1PRs that can activate S1PRs and induce an elevation in intracellular calcium concentration.

Improvements in surgical technique and perioperative care have resulted in increased long-term survival for patients with congenital heart disease. As these patients begin to reach their later years, clinicians are challenged with determining optimal management of noncardiac diseases in this complex patient population, including surgically treatable malignancies. We present a case of esophageal cancer in a patient with previously repaired tetralogy of Fallot and right-sided aortic arch, treated with neoadjuvant therapy followed by laparoscopic and left thoracoscopic esophagectomy., (Copyright © 2017 The Society of Thoracic Surgeons. Published by Elsevier Inc. All rights reserved.)

Atherosclerosis is a chronic inflammatory disease of large and medium-sized arteries and the underlying cause of cardiovascular disease, a major cause of mortality worldwide. The over-accumulation of modified cholesterol-containing low-density lipoproteins (e.g. oxLDL) in the artery wall and the subsequent recruitment and activation of macrophages contributes to the development of atherosclerosis. The excessive uptake of modified-LDL by macrophages leads to a lipid-laden "foamy" phenotype and pro-inflammatory cytokine production. Modified-LDLs promote foam cell formation in part by stimulating de novo lipid biosynthesis. However, it is unknown if lipid biosynthesis directly regulates foam cell pro-inflammatory mediator production. Lipin-1, a phosphatidate phosphohydrolase required for the generation of diacylglycerol during glycerolipid synthesis has recently been demonstrated to contribute to bacterial-induced pro-inflammatory responses by macrophages. In this study we present evidence demonstrating the presence of lipin-1 within macrophages in human atherosclerotic plaques. Additionally, reducing lipin-1 levels in macrophages significantly inhibits both modified-LDL-induced foam cell formation in vitro, as observed by smaller/fewer intracellular lipid inclusions, and ablates modified-LDL-elicited production of the pro-atherogenic mediators tumor necrosis factor-α, interleukin-6, and prostaglandin E2. These findings demonstrate a critical role for lipin-1 in the regulation of macrophage inflammatory responses to modified-LDL. These data begin to link the processes of foam cell formation and pro-inflammatory cytokine production within macrophages., (Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.)

DNA and porphyrin based therapeutics are important for anti-cancer treatment. The present studies demonstrate single-stranded DNA (ssDNA) assembles with meso-tetra-4-pyridyl porphine (MTP) forming porphyrin:DNA nano-complexes (PDN) that are stable in aqueous solution under physiologically relevant conditions and undergo dissociation with DNA release in hydrophobic environments, including cell membranes. PDN formation is DNA-dependent with the ratio of porphyrin:DNA being approximately two DNA nucleobases per porphyrin. PDN produce reactive oxygen species (ROS) in a light-dependent manner under conditions that favor nano-complex dissociation in the presence of hydrophobic solvents. PDN induce light-dependent cytotoxicity in vitro and anti-tumor activity towards bladder cancer xenografts in vivo. Light-dependent, PDN-mediated cell death results from ROS-mediated localized membrane damage due to lipid peroxidation with mass spectrometry indicating the generation of the lipid peroxidation products 9- and 13-hydroxy octadecanoic acid. Our results demonstrate that PDN have properties useful for therapeutic applications, including cancer treatment., From the Clinical Editor: In this study, porphyrin-DNA nanocomplexes were investigated as anti-cancer therapeutics inducing ROS production in a light-dependent manner. Efficacy is demonstrated in vitro as well as a in a bladder cancer xenograft model., (© 2014.)

This report details the lipid composition of nascent HDL (nHDL) particles formed by the action of the ATP binding cassette transporter A1 (ABCA1) on apolipoprotein A-I (apoA-I). nHDL particles of different size (average diameters of ∼ 12, 10, 7.5, and <6 nm) and composition were purified by size-exclusion chromatography. Electron microscopy suggested that the nHDL were mostly spheroidal. The proportions of the principal nHDL lipids, free cholesterol, glycerophosphocholine, and sphingomyelin were similar to that of lipid rafts, suggesting that the lipid originated from a raft-like region of the cell. Smaller amounts of glucosylceramides, cholesteryl esters, and other glycerophospholipid classes were also present. The largest particles, ∼ 12 nm and 10 nm diameter, contained ∼ 43% free cholesterol, 2-3% cholesteryl ester, and three apoA-I molecules. Using chemical cross-linking chemistry combined with mass spectrometry, we found that three molecules of apoA-I in the ∼ 9-14 nm nHDL adopted a belt-like conformation. The smaller (7.5 nm diameter) spheroidal nHDL particles carried 30% free cholesterol and two molecules of apoA-I in a twisted, antiparallel, double-belt conformation. Overall, these new data offer fresh insights into the biogenesis and structural constraints involved in forming nascent HDL from ABCA1.

We previously showed that macrophages from macrophage-specific ATP-binding cassette transporter A1 (ABCA1) knockout (Abca1(-M/-M)) mice had an enhanced proinflammatory response to the Toll-like receptor (TLR) 4 agonist, lipopolysaccharide (LPS), compared with wild-type (WT) mice. In the present study, we demonstrate a direct association between free cholesterol (FC), lipid raft content, and hyper-responsiveness of macrophages to LPS in WT mice. Abca1(-M/-M) macrophages were also hyper-responsive to specific agonists to TLR2, TLR7, and TLR9, but not TLR3, compared with WT macrophages. We hypothesized that ABCA1 regulates macrophage responsiveness to TLR agonists by modulation of lipid raft cholesterol and TLR mobilization to lipid rafts. We demonstrated that Abca1(-M/-M) vs. WT macrophages contained 23% more FC in isolated lipid rafts. Further, mass spectrometric analysis suggested raft phospholipid composition was unchanged. Although cell surface expression of TLR4 was similar between Abca1(-M/-M) and WT macrophages, significantly more TLR4 was distributed in membrane lipid rafts in Abca1(-M/-M) macrophages. Abca1(-M/-M) macrophages also exhibited increased trafficking of the predominantly intracellular TLR9 into lipid rafts in response to TLR9-specific agonist (CpG). Collectively, our data suggest that macrophage ABCA1 dampens inflammation by reducing MyD88-dependent TLRs trafficking to lipid rafts by selective reduction of FC content in lipid rafts.

This review discusses how to analyze glycerophospholipids from simple lipid extracts without preliminary chromatographic separation. The analytical method emphasized in the review is electrospray ionization with tandem mass spectrometry.

The purpose of this review is to highlight recent advances toward the refinement of a three-dimensional structure for lipid-bound apolipoprotein A-I (apoA-I) on recombinant HDL. Recently, X-ray crystallography has yielded a new structure for full-length, lipid-free apoA-I. Although this approach has not yet been successful in solving the three-dimensional structure of lipid-bound apoA-I, analysis of the X-ray structures has been of immense help in the interpretation of structural data obtained from other methods that yield structural information. Recent studies emphasize the use of mass spectrometry to unambiguously identify cross-linked peptides or to quantify solvent accessibility using hydrogen-deuterium exchange. The combination of mass spectrometry, molecular modeling, molecular dynamic analysis, and small-angle X-ray diffraction has provided additional structural information on apoA-I folding that complements previous approaches.

Diet-fed low density lipoprotein receptor-deficient/apolipoprotein A-I-deficient (LDLr-/-, apoA-I-/-) mice accumulate a 10-fold greater mass of cholesterol in their skin despite a 1.5- to 2-fold lower plasma cholesterol concentration compared with diet-fed LDLr-/- mice. The accumulation of cholesterol predominantly in the skin has been shown to occur in a growing number of other hypercholesterolemic double knockout mouse models sharing deficits in genes regulating cellular cholesterol homeostasis. Exploring the relationship between cholesterol balance and inflammation, we have examined the time course of cholesterol accumulation in a number of extrahepatic tissues and correlated with the onset of inflammation in diet-fed LDLr-/-, apoA-I-/- mice. After 4 weeks of diet, LDLr-/-, apoA-I-/- mice showed a significant increase in skin cholesterol mass compared with LDLr-/- mice. In addition, after 4 weeks on the diet, cholesterol accumulation in the skin was also found to be associated with macrophage infiltration and accompanied by increases in tumor necrosis factor-alpha, cyclooxygenase-2, and langerin mRNA, which were not seen in the liver. Overall, these data suggest that as early as 4 weeks after starting the diet, the accumulation of skin cholesterol and the onset of inflammation occur concurrently. In summary, the use of hypercholesterolemic LDLr-/-, apoA-I-/- mice may provide a useful tool to investigate the role that apoA-I plays in maintaining cholesterol homeostasis and its relationship to inflammation.

In this report, methods are described to isolate milligram quantities of a mutant apolipoprotein A-I (apoA-I) protein for use in structure-function studies. Expression of the L159R apoA-I mutation in humans reduces the concentration of plasma wild-type apoA-I, thus displaying a dominant negative phenotype in vivo. Earlier attempts to express and isolate this mutant protein resulted in extensive degradation and protein misfolding. Using an Escherichia coli expression system used predominantly for the isolation of soluble apoA-I mutant proteins, we describe the expression and purification of L159R apoA-I (apoA-I(Fin)) from inclusion bodies. In addition, we describe a mass spectrometric method for measuring the L159R-to-wild-type apoA-I ratio in a 1 microl plasma sample. These new methods will facilitate further studies into the mechanism behind the dominant negative phenotype associated with the expression of the L159R apoA-I protein in humans.

Gangliosides are critical in many functions of mammalian cells but present as a minor lipid component with many molecular species of subtle differences. Conventional strategies for profiling gangliosides suffer from poor reproducibility, low sensitivity, and low-throughput capacity. Prior separation of gangliosides by thin-layer chromatography and/or high-performance liquid chromatography not only was laborious and tedious but also could introduce uneven losses of molecular species. We developed a new strategy of using electrospray ionization-tandem mass spectrometry (ESI-MS/MS) to profile gangliosides with high-throughput potential. This strategy involves three new findings: (i) collision-induced fragmentation of gangliosides gave rise to a common ion of m/z 290, a derivative of N-acetylneuraminic acid; (ii) phospholipids exert a profound suppression of ganglioside detection in ESI-MS/MS to prevent a direct detection in total cellular lipid extracts; and (iii) enrichment of gangliosides in the aqueous phase from total cellular lipid extracts eliminates the damping effect of phospholipids and permits direct precursor scan.

We describe an improved assay for platelet-activating factor (PAF; 1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine) using HPLC-tandem mass spectrometry (LC-MS/MS). The present method can readily detect as little as 1 pg (1.9 fmol) of PAF, a significant improvement over previously described LC-MS/MS methods, and gives a linear response up to 1,000 pg of PAF. Our method also overcomes the artifacts from isobaric lipids that have limited the usefulness of certain existing LC-MS/MS assays for PAF. In the course of these studies, we detected three novel lipid species in human neutrophils. One of the novel lipids appears to be a new molecular species of PAF, and the other two have chromatographic and mass spectrometric properties consistent with stearoyl-formyl-glycerophosphocholine and oleoyl-formyl-glycerophosphocholine. These observations identify previously unknown potential interferences in the measurement of PAF by LC-MS/MS. Moreover, our data suggest that the previously described palmitoyl-formyl-glycerophosphocholine is not unique but rather is a member of a new and poorly understood family of formylated lipids.

From a total of 47 known apolipoprotein A-I (apoA-I) mutations, only 18 are linked to low plasma HDL apoA-I concentrations, and 78% of these map to apoA-I helices 6 and 7 (residues 143-186). Gene transfer and transgenic mouse studies have shown that several helix 6 apoA-I mutations have reduced hepatic HDL production. Our objective was to examine the impact of helix 6 modifications on intracellular biosynthetic processing and secretion of apoA-I. Cells were transfected with wild-type or mutant apoA-I, radiolabeled with [(35)S]Met/Cys, and then placed in unlabeled medium for up to 4 h. Results show that >90% of newly synthesized wild-type apoA-I was secreted by 60 min. Over the same length of time, only 20% of helix 6 deletion mutant (Delta 6 apoA-I) was secreted, whereas 80% remained cell associated. Microscopic and biochemical studies revealed that cell-associated Delta 6 apoA-I was located predominantly within the cytoplasm as lipid-protein inclusions, whereas wild-type apoA-I was localized in the endoplasmic reticulum/Golgi. Results using other helix deletions or helix 6 substitution mutations indicated that only complete removal of helix 6 resulted in massive cytoplasmic accumulation. These data suggest that alterations in native apoA-I conformation can lead to aberrant trafficking and accumulation of apolipoprotein-phospholipid structures. Thus, conformation-dependent alterations in intracellular trafficking and turnover may underlie the reduced plasma HDL concentrations observed in individuals harboring deletion mutations within helix 6.

We compared the in vivo metabolism of prebeta HDL particles isolated by anti-human apolipoprotein A-I (apoA-I) immunoaffinity chromatography (LpA-I) in human apoA-I transgenic (hA-I Tg) mice with that of lipid-free apoA-I (LFA-I) and small LpA-I. After injection, prebeta LpA-I were removed from plasma more rapidly than were LFA-I and small LpA-I. Prebeta LpA-I and LFA-I were preferentially degraded by kidney compared with liver; small LpA-I were preferentially degraded by the liver. Five minutes after tracer injection, 99% of LFA-I in plasma was found to be associated with medium-sized (8.6 nm) HDL, whereas only 37% of prebeta tracer remodeled to medium-sized HDL. Injection of prebeta LpA-I doses into C57Bl/6 recipients resulted in a slower plasma decay compared with hA-I Tg recipients and a greater proportion (>60%) of the prebeta radiolabel that was associated with medium-sized HDL. Prebeta LpA-I contained one to four molecules of phosphatidylcholine per molecule of apoA-I, whereas LFA-I contained less than one. We conclude that prebeta LpA-I has two metabolic fates in vivo, rapid removal from plasma and catabolism by kidney or remodeling to medium-sized HDL, which we hypothesize is determined by the amount of lipid associated with the prebeta particle and the particle's ability to bind to medium-sized HDL.

Atherogenic response to dietary fat and cholesterol challenge was evaluated in mice lacking both the LDL receptor (LDLr(-/-)) and apoA-I (apoA-I(-/-)) gene, LDLr(-/-)/apoA-I(-/-) or double-knockout mice. Gender- and age-matched LDLr(-/-)/apoA-I(-/-) mice were fed a diet consisting of 0.1% cholesterol and 10% palm oil for 16 weeks and compared to LDLr(-/-) mice or single-knockout mice. The LDLr(-/-) mice showed a 6- to 7-fold increase in total plasma cholesterol (TPC) compared to their chow-fed mice counterparts, while LDLr(-/-)/apoA-I(-/-) mice showed only a 2- to 3-fold increase in TPC compared to their chow-fed controls. This differential response to the atherogenic diet was unanticipated, since chow-fed LDLr(-/-) and LDLr(-/-)/apoA-I(-/-) mice began the study with similar LDL levels and differed primarily in their HDL concentration. The 6-fold diet-induced increase in TPC observed in the LDLr(-/-) mice occurred mainly in VLDL/LDL and not in HDL. Mid-study plasma samples taken after 8 weeks of diet feeding showed that LDLr(-/-) mice had TPC concentrations approximately 60% of their 16-week level, while the LDLr(-/-)/apoA-I(-/-) mice had reached 100% of their 16-week TPC concentration after only 8 weeks of diet. Male LDLr(-/-) mice showed similar aortic cholesterol levels to male LDLr(-/-)/apoA-I(-/-) mice despite a 4-fold higher VLDL/LDL concentration in the LDLr(-/-) mice. A direct comparison of the severity of aortic atherosclerosis between female LDLr(-/-) and LDLr(-/-)/apoA-I(-/-) mice was compromised due to the loss of female LDLr(-/-)/apoA-I(-/-) mice between 10 and 14 weeks into the study. Diet-fed female and, with time, male LDLr(-/-)/apoA-I(-/-) mice suffered from severe ulcerated cutaneous xanthomatosis. This condition, combined with a complete depletion of adrenal cholesterol, manifested in fatal wasting of the affected mice. In conclusion, LDLr(-/-) and LDLr(-/-)/apoA-I(-/-) mice showed dramatic TPC differences in response to dietary fat and cholesterol challenge, while despite these differences both genotypes accumulated similar levels of aortic cholesterol.

In this study, we asked the question "does alpha-tocopherol supplementation prevent an increase in total plasma cholesterol (TPC) concentration and reduce the deposition of cholesterol in arterial plaques of rabbits fed atherogenic diets?" Isocaloric diets containing 0.1% cholesterol to induce atherosclerosis were enriched in one of three fats: saturated fats (SAT), monounsaturated fats (MONO), or n-6 polyunsaturated fats (POLY). Half of each of the three diets were supplemented with 2,500 IU alpha-tocopherol/kg-diet. Unsupplemented diets contained 25 IU alpha-tocopherol/kg-diet. Rabbits supplemented with alpha-tocopherol had plasma alpha-tocopherol concentrations 10-fold higher and an average TPC concentration 31% lower, P = 0.017, than rabbits fed unsupplemented diets. Among the three fat-fed groups, the difference was greatest for the POLY fat fed group (54%, P = 0.041). POLY fat-fed rabbits without alpha-tocopherol supplementation had plasma HDL cholesterol concentrations that were less than half that of rabbits fed other fats, P < or = 0.0001. In general, differences in mean esterified artery cholesterol concentrations among the three fat-fed groups, with and without alpha-tocopherol supplementation, paralleled differences in TPC concentration among the groups. This study suggests that for rabbits fed high pharmacological doses of alpha-tocopherol, atherosclerosis can be diminished in situations where the plasma cholesterol concentrations are also significantly lower.