Foam fractionation studies of recombinant human apolipoprotein A-I.

Apolipoprotein A-I (apoA-I), the primary protein component of plasma high-density lipoproteins (HDL), is comprised of two structural regions, an N-terminal amphipathic α-helix bundle domain (residues 1-184) and a hydrophobic C-terminal domain (residues 185-243). When a recombinant fusion protein construct [bacterial pelB leader sequence - human apoA-I (1-243)] was expressed in Escherichia coli shaker flask cultures, apoA-I was recovered in the cell lysate. By contrast, when the C-terminal domain was deleted from the construct, large amounts of the truncated protein, apoA-I (1-184), were recovered in the culture medium. Consequently, following pelB leader sequence cleavage in the E. coli periplasmic space, apoA-I (1-184) was secreted from the bacteria. When the pelB-apoA-I (1-184) fusion construct was expressed in a 5 L bioreactor, substantial foam production (~30 L) occurred. Upon foam collection and collapse into a liquid foamate, SDS-PAGE revealed that apoA-I (1-184) was the sole major protein present. Incubation of apoA-I (1-184) with phospholipid vesicles yielded reconstituted HDL (rHDL) particles that were similar in size and cholesterol efflux capacity to those generated with full-length apoA-I. Mass spectrometry analysis confirmed that pelB leader sequence cleavage occurred and that foam fractionation did not result in unwanted protein modifications. The facile nature and scalability of bioreactor-based apolipoprotein foam fractionation provide a novel means to generate a versatile rHDL scaffold protein.

Photophysical Characterization and In Vitro Evaluation of α-Mangostin-Loaded HDL Mimetic Nano-Complex in LN-229 Glioblastoma Spheroid Model.

Cytotoxic activity has been reported for the xanthone α-mangostin (AMN) against Glioblastoma multiforme (GBM), an aggressive malignant brain cancer with a poor prognosis. Recognizing that AMN's high degree of hydrophobicity is likely to limit its systemic administration, we formulated AMN using reconstituted high-density lipoprotein (rHDL) nanoparticles. The photophysical characteristics of the formulation, including fluorescence lifetime and steady-state anisotropy, indicated that AMN was successfully incorporated into the rHDL nanoparticles. To our knowledge, this is the first report on the fluorescent characteristics of AMN with an HDL-based drug carrier. Cytotoxicity studies in a 2D culture and 3D spheroid model of LN-229 GBM cells and normal human astrocytes showed an enhanced therapeutic index with the rHDL-AMN formulation compared to the unincorporated AMN and Temozolomide, a standard GBM chemotherapy agent. Furthermore, treatment with the rHDL-AMN facilitated a dose-dependent upregulation of autophagy and reactive oxygen species generation to a greater extent in LN-229 cells compared to astrocytes, indicating the reduced off-target toxicity of this novel formulation. These studies indicate the potential therapeutic benefits to GBM patients via selective targeting using the rHDL-AMN formulation.

Thermal Remodeling of Human HDL Particles Reveals Diverse Subspecies.

High-density lipoproteins (HDL) are micelle-like particles consisting of a core of triglycerides and cholesteryl esters surrounded by a shell of phospholipid, cholesterol, and apolipoproteins. HDL is considered "good" cholesterol, and its concentration in plasma is used clinically in assessing cardiovascular health. However, these particles vary in structure, composition, and therefore function, and thus can be resolved into subpopulations, some of which have specific cardioprotective properties. Mass measurements of HDL by charge detection mass spectrometry (CD-MS) previously revealed seven distinct subpopulations which could be delineated by mass and charge [Lutomski, C. A. et al. Anal. Chem. 2018]. Here, we investigate the thermal stabilities of these subpopulations; upon heating, the particles within each subpopulation undergo structural rearrangements with distinct transition temperatures. In addition, we find evidence for many new families of structures within each subpopulation; at least 15 subspecies of HDL are resolved. These subspecies vary in size, charge, and thermal stability. While this suggests that these new subspecies have unique molecular compositions, we cannot rule out the possibility that we have found evidence for new structural forms within the known subpopulations. The ability to resolve new subspecies of HDL particles may be important in understanding and delineating the role of unique particles in cardiovascular health and disease.

Negatively charged nanodiscs for the reduction of toxicity and enhanced efficacy of polymyxin B against Acinetobacter baumannii sepsis.

The treatment of sepsis caused by multidrug-resistant (MDR) Gram-negative bacterial infections remains challenging. With these pathogens exhibiting resistance to carbapenems and new generation cephalosporins, the traditional antibiotic polymyxin B (PMB) has reemerged as a critical treatment option. However, its severe neurotoxicity and nephrotoxicity greatly limit the clinical application. Therefore, we designed negatively charged high-density lipoprotein (HDL) mimicking nanodiscs as a PMB delivery system, which can simultaneously reduce toxicity and enhance drug efficacy. The negative charge prevented the PMB release in physiological conditions and binding to cell membranes, significantly reducing toxicity in mammalian cells and mice. Notably, nanodisc-PMB exhibits superior efficacy than free PMB in sepsis induced by carbapenem-resistant Acinetobacter baumannii (CRAB) strains. Nanodisc-PMB shows promise as a treatment for carbapenem-resistant Gram-negative bacterial sepsis, especially caused by Acinetobacter baumannii, and the nanodiscs could be repurposed for other toxic antibiotics as an innovative delivery system. STATEMENT OF SIGNIFICANCE: Multidrug-resistant Gram-negative bacteria, notably carbapenem-resistant Acinetobacter baumannii, currently pose a substantial challenge due to the scarcity of effective treatments, rendering Polymyxins a last-resort antibiotic option. However, their therapeutic application is significantly limited by severe neurotoxic and nephrotoxic side effects. Prevailing polymyxin delivery systems focus on either reducing toxicity or enhancing bioavailability yet fail to simultaneously achieve both. In this scenario, we have developed a distinctive HDL-mimicking nanodisc for polymyxin B, which not only significantly reduces toxicity but also improves efficacy against Gram-negative bacteria, especially in sepsis caused by CRAB. This research offers an innovative drug delivery system for polymyxin B. Such advancement could notably improve the therapeutic landscape and make a significant contribution to the arsenal against these notorious pathogens.

Microvortex-induced turbulent mixing for reconstitution of high-density lipoprotein-mimicking nanoparticles with aggregation-prone phosphatidylcholine.

Lipid nanoparticles often contain a phosphatidylcholine with a long chain fatty acid, 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC). However, their preparation often encounters difficulties such as the inability to yield <20 nm nanoparticles due to the aggregation-prone behavior of DSPC. High-density lipoproteins (HDLs) are ∼10 nm protein-bound lipid nanoparticles in our body, and microfluidic preparations of HDL-mimicking nanoparticles (µHDL) have been reported. Herein, we report a new microfluidic mixing mode that enables preparation of µHDL with DSPC in high yield (≥90% on a protein basis). The critical mechanism of this mode is a spontaneous asymmetric distribution of the ethanol flow injected in a symmetric manner followed by turbulent mixing in a simple rectangular parallelepiped-shaped chip.

Constructing myosin/high-density lipoprotein composite emulsions: Roles of pH on emulsification stability, rheological and structural properties.

The emulsification activity of myosin plays a significant role in affecting quality of emulsified meat products. High-density lipoprotein (HDL) possesses strong emulsification activity and stability due to its structural characteristics, suggesting potential for its utilization in developing functional emulsified meat products. In order to explore the effect of HDL addition on emulsification stability, rheological properties and structural features of myosin (MS) emulsions, HDL-MS emulsion was prepared by mixing soybean oil with isolated HDL and MS, with pH adjustments ranging from 3.0 to 11.0. The results found that emulsification activity and stability in two emulsion groups consistently improved as pH increased. Under identical pH, HDL-MS emulsion exhibited superior emulsification behavior as compared to MS emulsion. The HDL-MS emulsion under pH of 7.0-11.0 formed a viscoelastic protein layer at the interface, adsorbing more proteins and retarding oil droplet diffusion, leading to enhanced oxidative stability, compared to the MS emulsion. Raman spectroscopy analysis showed more flexible conformational changes in the HDL-MS emulsion. Microstructural observations corroborated these findings, showing a more uniform distribution of droplet sizes in the HDL-MS emulsion with smaller particle sizes. Overall, these determinations suggested that the addition of HDL enhanced the emulsification behavior of MS emulsions, and the composite emulsions demonstrated heightened responsiveness under alkaline conditions. This establishes a theoretical basis for the practical utilization of HDL in emulsified meat products.

Synergistic Anti-Inflammatory Activity of Lipid-Free Apolipoprotein (apo) A-I and CIGB-258 in Acute-Phase Zebrafish via Stabilization of the apoA-I Structure to Enhance Anti-Glycation and Antioxidant Activities.

CIGB-258, a 3 kDa peptide from heat shock protein 60, exhibits synergistic anti-inflammatory activity with apolipoprotein A-I (apoA-I) in reconstituted high-density lipoproteins (rHDLs) via stabilization of the rHDL structure. This study explored the interactions between CIGB-258 and apoA-I in the lipid-free state to assess their synergistic effects in the structural and functional enhancement of apoA-I and HDL. A co-treatment of lipid-free apoA-I and CIGB-258 inhibited the cupric ion-mediated oxidation of low-density lipoprotein (LDL) and a lowering of oxidized species in the dose-responsive manner of CIGB-258. The co-presence of CIGB-258 caused a blue shift in the wavelength of maximum fluorescence (WMF) of apoA-I with protection from proteolytic degradation. The addition of apoA-I:CIGB-258, with a molar ratio of 1:0.1, 1:0.5, and 1:1, to HDL2 and HDL3 remarkably enhanced the antioxidant ability against LDL oxidation up to two-fold higher than HDL alone. HDL-associated paraoxonase activities were elevated up to 28% by the co-addition of apoA-I and CIGB-258, which is linked to the suppression of Cu2+-mediated HDL oxidation with the slowest electromobility. Isothermal denaturation by a urea treatment showed that the co-presence of CIGB-258 attenuated the exposure of intrinsic tryptophan (Trp) and increased the mid-points of denaturation from 2.33 M for apoA-I alone to 2.57 M for an apoA-I:CIGB-258 mixture with a molar ratio of 1:0.5. The addition of CIGB-258 to apoA-I protected the carboxymethyllysine (CML)-facilitated glycation of apoA-I with the prevention of Trp exposure. A co-treatment of apoA-I and CIGB-258 synergistically safeguarded zebrafish embryos from acute death by CML-toxicity, suppressing oxidative stress and apoptosis. In adult zebrafish, the co-treatment of apoA-I+CIGB-258 exerted the highest anti-inflammatory activity with a higher recovery of swimming ability and survivability than apoA-I alone or CIGB-258 alone. A co-injection of apoA-I and CIGB-258 led to the lowest infiltration of neutrophils and interleukin (IL)-6 generation in hepatic tissue, with the lowest serum triglyceride, aspartate transaminase, and alanine transaminase levels in plasma. In conclusion, the co-presence of CIGB-258 ameliorated the beneficial functionalities of apoA-I, such as antioxidant and anti-glycation activities, by enhancing the structural stabilization and protection of apoA-I. The combination of apoA-I and CIGB-258 synergistically enforced the anti-inflammatory effect against CML toxicity in embryos and adult zebrafish.

Development of Recombinant High-Density Lipoprotein Platform with Innate Adipose Tissue-Targeting Abilities for Regional Fat Reduction.

As an escalating public health issue, obesity and overweight conditions are predispositions to various diseases and are exacerbated by concurrent chronic inflammation. Nonetheless, extant antiobesity pharmaceuticals (quercetin, capsaicin, catecholamine, etc.) manifest constrained efficacy alongside systemic toxic effects. Effective therapeutic approaches that selectively target adipose tissue, thereby enhancing local energy expenditure, surmounting the limitations of prevailing antiobesity modalities are highly expected. In this context, we developed a temperature-sensitive hydrogel loaded with recombinant high-density lipoprotein (rHDL) to achieve targeted delivery of resveratrol, an adipose browning activator, to adipose tissue. rHDL exhibits self-regulation on fat cell metabolism and demonstrates natural targeting toward scavenger receptor class B type I (SR-BI), which is highly expressed by fat cells, thereby achieving a synergistic effect for the treatment of obesity. Additionally, the dispersion of rHDL@Res in temperature-sensitive hydrogels, coupled with the regulation of their degradation and drug release rate, facilitated sustainable drug release at local adipose tissues over an extended period. Following 24 days' treatment regimen, obese mice exhibited improved metabolic status, resulting in a reduction of 68.2% of their inguinal white adipose tissue (ingWAT). Specifically, rHDL@Res/gel facilitated the conversion of fatty acids to phospholipids (PA, PC), expediting fat mobilization, mitigating triglyceride accumulation, and therefore facilitating adipose tissue reduction. Furthermore, rHDL@Res/gel demonstrated efficacy in attenuating obesity-induced inflammation and fostering angiogenesis in ingWAT. Collectively, this engineered local fat reduction platform demonstrated heightened effectiveness and safety through simultaneously targeting adipocytes, promoting WAT browning, regulating lipid metabolism, and controlling inflammation, showing promise for adipose-targeted therapy.

Detection of atherosclerotic plaques with HDL-like porphyrin nanoparticles using an intravascular dual-modality optical coherence tomography and fluorescence system.

Atherosclerosis is the build-up of fatty plaques within blood vessel walls, which can occlude the vessels and cause strokes or heart attacks. It gives rise to both structural and biomolecular changes in the vessel walls. Current single-modality imaging techniques each measure one of these two aspects but fail to provide insight into the combined changes. To address this, our team has developed a dual-modality imaging system which combines optical coherence tomography (OCT) and fluorescence imaging that is optimized for a porphyrin lipid nanoparticle that emits fluorescence and targets atherosclerotic plaques. Atherosclerosis-prone apolipoprotein (Apo)e-/- mice were fed a high cholesterol diet to promote plaque development in descending thoracic aortas. Following infusion of porphyrin lipid nanoparticles in atherosclerotic mice, the fiber-optic probe was inserted into the aorta for imaging, and we were able to robustly detect a porphyrin lipid-specific fluorescence signal that was not present in saline-infused control mice. We observed that the nanoparticle fluorescence colocalized in areas of CD68+ macrophages. These results demonstrate that our system can detect the fluorescence from nanoparticles, providing complementary biological information to the structural information obtained from simultaneously acquired OCT.

High-density lipoprotein mimetic nano-therapeutics targeting monocytes and macrophages for improved cardiovascular care: a comprehensive review.

The prevalence of cardiovascular diseases continues to be a challenge for global health, necessitating innovative solutions. The potential of high-density lipoprotein (HDL) mimetic nanotherapeutics in the context of cardiovascular disease and the intricate mechanisms underlying the interactions between monocyte-derived cells and HDL mimetic showing their impact on inflammation, cellular lipid metabolism, and the progression of atherosclerotic plaque. Preclinical studies have demonstrated that HDL mimetic nanotherapeutics can regulate monocyte recruitment and macrophage polarization towards an anti-inflammatory phenotype, suggesting their potential to impede the progression of atherosclerosis. The challenges and opportunities associated with the clinical application of HDL mimetic nanotherapeutics, emphasize the need for additional research to gain a better understanding of the precise molecular pathways and long-term effects of these nanotherapeutics on monocytes and macrophages to maximize their therapeutic efficacy. Furthermore, the use of nanotechnology in the treatment of cardiovascular diseases highlights the potential of nanoparticles for targeted treatments. Moreover, the concept of theranostics combines therapy and diagnosis to create a selective platform for the conversion of traditional therapeutic medications into specialized and customized treatments. The multifaceted contributions of HDL to cardiovascular and metabolic health via highlight its potential to improve plaque stability and avert atherosclerosis-related problems. There is a need for further research to maximize the therapeutic efficacy of HDL mimetic nanotherapeutics and to develop targeted treatment approaches to prevent atherosclerosis. This review provides a comprehensive overview of the potential of nanotherapeutics in the treatment of cardiovascular diseases, emphasizing the need for innovative solutions to address the challenges posed by cardiovascular diseases.

Lipid exchange of apolipoprotein A-I amyloidogenic variants in reconstituted high-density lipoprotein with artificial membranes.

High-density lipoproteins (HDLs) are responsible for removing cholesterol from arterial walls, through a process known as reverse cholesterol transport. The main protein in HDL, apolipoprotein A-I (ApoA-I), is essential to this process, and changes in its sequence significantly alter HDL structure and functions. ApoA-I amyloidogenic variants, associated with a particular hereditary degenerative disease, are particularly effective at facilitating cholesterol removal, thus protecting carriers from cardiovascular disease. Thus, it is conceivable that reconstituted HDL (rHDL) formulations containing ApoA-I proteins with functional/structural features similar to those of amyloidogenic variants hold potential as a promising therapeutic approach. Here we explored the effect of protein cargo and lipid composition on the function of rHDL containing one of the ApoA-I amyloidogenic variants G26R or L174S by Fourier transformed infrared spectroscopy and neutron reflectometry. Moreover, small-angle x-ray scattering uncovered the structural and functional differences between rHDL particles, which could help to comprehend higher cholesterol efflux activity and apparent lower phospholipid (PL) affinity. Our findings indicate distinct trends in lipid exchange (removal vs. deposition) capacities of various rHDL particles, with the rHDL containing the ApoA-I amyloidogenic variants showing a markedly lower ability to remove lipids from artificial membranes compared to the rHDL containing the native protein. This effect strongly depends on the level of PL unsaturation and on the particles' ultrastructure. The study highlights the importance of the protein cargo, along with lipid composition, in shaping rHDL structure, contributing to our understanding of lipid-protein interactions and their behavior.

Structural studies of a serum amyloid A octamer that is primed to scaffold lipid nanodiscs.

Serum amyloid A (SAA) is a highly conserved acute-phase protein that plays roles in activating multiple pro-inflammatory pathways during the acute inflammatory response and is commonly used as a biomarker of inflammation. It has been linked to beneficial roles in tissue repair through improved clearance of lipids and cholesterol from sites of damage. In patients with chronic inflammatory diseases, elevated levels of SAA may contribute to increased severity of the underlying condition. The majority of circulating SAA is bound to lipoproteins, primarily high-density lipoprotein (HDL). Interaction with HDL not only stabilizes SAA but also alters its functional properties, likely through altered accessibility of protein-protein interaction sites on SAA. While high-resolution structures for lipid-free, or apo-, forms of SAA have been reported, their relationship with the HDL-bound form of the protein, and with other possible mechanisms of SAA binding to lipids, has not been established. Here, we have used multiple biophysical techniques, including SAXS, TEM, SEC-MALS, native gel electrophoresis, glutaraldehyde crosslinking, and trypsin digestion to characterize the lipid-free and lipid-bound forms of SAA. The SAXS and TEM data show the presence of soluble octamers of SAA with structural similarity to the ring-like structures reported for lipid-free ApoA-I. These SAA octamers represent a previously uncharacterized structure for lipid-free SAA and are capable of scaffolding lipid nanodiscs with similar morphology to those formed by ApoA-I. The SAA-lipid nanodiscs contain four SAA molecules and have similar exterior dimensions as the lipid-free SAA octamer, suggesting that relatively few conformational rearrangements may be required to allow SAA interactions with lipid-containing particles such as HDL. This study suggests a new model for SAA-lipid interactions and provides new insight into how SAA might stabilize protein-lipid nanodiscs or even replace ApoA-I as a scaffold for HDL particles during inflammation.

Optimizing the formation of myosin/high-density lipoprotein composite gels: PH-dependent effects on heat-induced aggregation.

This study investigated impact of high-density lipoprotein (HDL) on thermal aggregation and gelling behavior of myosin in relation to varied pHs. Results revealed that HDL modified myosin structure before and after heating, with distinct effects observed at varied pH. Under pH 5.0, both myosin and HDL-MS exhibited larger aggregates and altered microstructure; at pH 7.0 and 9.0, HDL inhibited myosin aggregation, resulting in enhanced solubility, reduced turbidity and particle size. Comparative analysis of surface hydrophobicity, free sulfhydryl groups and secondary structure highlighted distinct thermal aggregation behavior between MS and HDL-MS, with the latter showing inhibitory effects under neutral or alkaline conditions. Gelation behavior was enhanced at pH 7.0 with maximum strength, hardness, water-holding capacity and rheological properties. Under acidic pH, excessive protein aggregation resulted in increased whiteness and rough microstructure with granular aggregates. Under alkaline pH, gel network structure was weaker, possibly due to higher thermal stability of protein molecules. Scanning electron microscopy revealed expanded HDL protein particles at pH 7.0, accounting for decreased gel strength and altered rheological properties compared with myosin gel. Overall, the results indicated a positive role of HDL at varied pH in regulating thermal aggregation of myosin and further impacting heat-induced gel characteristics.

An atomistic characterization of high-density lipoproteins and the conserved "LN" region of apoA-I.

The physicochemical characteristics of the various subpopulations of high-density lipoproteins (HDLs) and, in particular, their surface properties determine their ability to scavenge lipids and interact with specific receptors and peptides. Five representative spheroidal HDL subpopulation models were mapped from a previously reported equilibrated coarse-grained (CG) description to an atomistic representation for subsequent molecular dynamics simulation. For each HDL model a range of finer-level analyses was undertaken, including the component-wise characterization of HDL surfaces, the average size and composition of hydrophobic surface patches, dynamic protein secondary structure monitoring, and the proclivity for solvent exposure of the proposed ß-amyloid (Aß) binding region of apolipoprotein A-I (apoA-I), "LN." This study reveals that previously characterized ellipsoidal HDL3a and HDL2a models revert to a more spherical geometry in an atomistic representation due to the enhanced conformational flexibility afforded to the apoA-I protein secondary structure, allowing for enhanced surface lipid packing and lower overall surface hydrophobicity. Indeed, the proportional surface hydrophobicity and apoA-I exposure reduced with increasing HDL size, consistent with previous characterizations. Furthermore, solvent exposure of the "LN" region of apoA-I was exclusively limited to the smallest HDL3c model within the timescale of the simulations, and typically corresponded to a distinct loss in secondary structure across the "LN" region to form part of a significant contiguous hydrophobic patch on the HDL surface. Taken together, these findings provide preliminary evidence for a subpopulation-specific interaction between HDL3c particles and circulating hydrophobic species such as Aß via the exposed "LN" region of apoA-I.

Manipulating Redox Homeostasis of Cancer Stem Cells Overcome Chemotherapeutic Resistance through Photoactivatable Biomimetic Nanodiscs.

Tumor heterogeneity remains a significant obstacle in cancer therapy due to diverse cells with varying treatment responses. Cancer stem-like cells (CSCs) contribute significantly to intratumor heterogeneity, characterized by high tumorigenicity and chemoresistance. CSCs reside in the depth of the tumor, possessing low reactive oxygen species (ROS) levels and robust antioxidant defense systems to maintain self-renewal and stemness. A nanotherapeutic strategy is developed using tumor-penetrating peptide iRGD-modified high-density lipoprotein (HDL)-mimetic nanodiscs (IPCND) that ingeniously loaded with pyropheophorbide-a (Ppa), bis (2-hydroxyethyl) disulfide (S-S), and camptothecin (CPT) by synthesizing two amphiphilic drug-conjugated sphingomyelin derivatives. Photoactivatable Ppa can generate massive ROS which as intracellular signaling molecules effectively shut down self-renewal and trigger differentiation of the CSCs, while S-S is utilized to deplete GSH and sustainably imbalance redox homeostasis by reducing ROS clearance. Simultaneously, the depletion of GSH is accompanied by the release of CPT, which leads to subsequent cell death. This dual strategy successfully disturbed the redox equilibrium of CSCs, prompting their differentiation and boosting the ability of CPT to kill CSCs upon laser irradiation. Additionally, it demonstrated a synergistic anti-cancer effect by concurrently eliminating therapeutically resistant CSCs and bulk tumor cells, effectively suppressing tumor growth in CSC-enriched heterogeneous colon tumor mouse models.

High-density lipoprotein in diabetes: Structural and functional relevance.

Low levels of high-density lipoprotein-cholesterol (HDL-C) is considered a major cardiovascular risk factor. However, recent studies have suggested a more U-shaped association between HDL-C and cardiovascular disease. It has been shown that the cardioprotective effect of HDL is related to the functions of HDL particles rather than their cholesterol content. HDL particles are highly heterogeneous and have multiple functions relevant to cardiometabolic conditions including cholesterol efflux capacity, anti-oxidative, anti-inflammatory, and vasoactive properties. There are quantitative and qualitative changes in HDL as well as functional abnormalities in both type 1 and type 2 diabetes. Non-enzymatic glycation, carbamylation, oxidative stress, and systemic inflammation can modify the HDL composition and therefore the functions, especially in situations of poor glycemic control. Studies of HDL proteomics and lipidomics have provided further insights into the structure-function relationship of HDL in diabetes. Interestingly, HDL also has a pleiotropic anti-diabetic effect, improving glycemic control through improvement in insulin sensitivity and ß-cell function. Given the important role of HDL in cardiometabolic health, HDL-based therapeutics are being developed to enhance HDL functions rather than to increase HDL-C levels. Among these, recombinant HDL and small synthetic apolipoprotein A-I mimetic peptides may hold promise for preventing and treating diabetes and cardiovascular disease.

Apolipoprotein E secreted by astrocytes forms antiparallel dimers in discoidal lipoproteins.

The Apolipoprotein E gene (APOE) is of great interest due to its role as a risk factor for late-onset Alzheimer's disease. ApoE is secreted by astrocytes in the central nervous system in high-density lipoprotein (HDL)-like lipoproteins. Structural models of lipidated ApoE of high resolution could aid in a mechanistic understanding of how ApoE functions in health and disease. Using monoclonal Fab and F(ab')2 fragments, we characterize the structure of lipidated ApoE on astrocyte-secreted lipoproteins. Our results provide support for the "double-belt" model of ApoE in nascent discoidal HDL-like lipoproteins, where two ApoE proteins wrap around the nanodisc in an antiparallel conformation. We further show that lipidated, recombinant ApoE accurately models astrocyte-secreted ApoE lipoproteins. Cryogenic electron microscopy of recombinant lipidated ApoE further supports ApoE adopting antiparallel dimers in nascent discoidal lipoproteins.

Nuclear Magnetic Resonance-Based Metabolomics and Risk of CKD.

RATIONALE & OBJECTIVE: Chronic kidney disease (CKD) leads to lipid and metabolic abnormalities, but a comprehensive investigation of lipids, lipoprotein particles, and circulating metabolites associated with the risk of CKD has been lacking. We examined the associations of nuclear magnetic resonance (NMR)-based metabolomics data with CKD risk in the UK Biobank study. STUDY DESIGN: Observational cohort study. SETTING & PARTICIPANTS: A total of 91,532 participants in the UK Biobank Study without CKD and not receiving lipid-lowering therapy. EXPOSURE: Levels of metabolites including lipid concentration and composition within 14 lipoprotein subclasses, as well as other metabolic biomarkers were quantified via NMR spectroscopy. OUTCOME: Incident CKD identified using ICD codes in any primary care data, hospital admission records, or death register records. ANALYTICAL APPROACH: Cox proportional hazards regression models were used to estimate hazard ratios and 95% confidence intervals. RESULTS: We identified 2,269 CKD cases over a median follow-up period of 13.1 years via linkage with the electronic health records. After adjusting for covariates and correcting for multiple testing, 90 of 142 biomarkers were significantly associated with incident CKD. In general, higher concentrations of very-low-density lipoprotein (VLDL) particles were associated with a higher risk of CKD whereas higher concentrations of high-density lipoprotein (HDL) particles were associated with a lower risk of CKD. Higher concentrations of cholesterol, phospholipids, and total lipids within VLDL were associated with a higher risk of CKD, whereas within HDL they were associated with a lower risk of CKD. Further, higher triglyceride levels within all lipoprotein subclasses, including all HDL particles, were associated with greater risk of CKD. We also identified that several amino acids, fatty acids, and inflammatory biomarkers were associated with risk of CKD. LIMITATIONS: Potential underreporting of CKD cases because of case identification via electronic health records. CONCLUSIONS: Our findings highlight multiple known and novel pathways linking circulating metabolites to the risk of CKD. PLAIN-LANGUAGE SUMMARY: The relationship between individual lipoprotein particle subclasses and lipid-related traits and risk of chronic kidney disease (CKD) in general population is unclear. Using data from 91,532 participants in the UK Biobank, we evaluated the associations of metabolites measured using nuclear magnetic resonance testing with the risk of CKD. We identified that 90 out of 142 lipid biomarkers were significantly associated with incident CKD. We found that very-low-density lipoproteins, high-density lipoproteins, the lipid concentration and composition within these lipoproteins, triglycerides within all the lipoprotein subclasses, fatty acids, amino acids, and inflammation biomarkers were associated with CKD risk. These findings advance our knowledge about mechanistic pathways that may contribute to the development of CKD.

Enhancement of Anti-inflammatory Effects of Synthetic High-Density Lipoproteins by Incorporation of Anionic Lipids.

Phosphatidylserine (PS) is an anionic phospholipid component in endogenous high-density lipoprotein (HDL). With the intrinsic anti-inflammatory effects of PS and the correlation between PS content and HDL functions, it was hypothesized that incorporating PS would enhance the therapeutic effects of HDL mimetic particles. To test this hypothesis, a series of synthetic high-density lipoproteins (sHDLs) were prepared with an apolipoprotein A-I (ApoA-1) mimetic peptide, 1-palmitoyl-2-oleoyl-glycero-3-phosphocholine (POPC), and 1-palmitoyl-2-oleoyl-glycero-3-phospho-l-serine (POPS). Incorporating PS was found to improve the particle stability of sHDLs. Moreover, increasing the PS content in sHDLs enhanced the anti-inflammatory effects on lipopolysaccharide-activated macrophages and endothelial cells. The incorporation of PS had no negative impact on cholesterol efflux capacity, in vivo cholesterol mobilization, and did not affect the pharmacokinetic profiles of sHDLs. Such results suggest the therapeutic potential of PS-containing sHDLs for inflammation resolution in atherosclerosis and other inflammatory diseases.

Effects of CaCl2 on the structure of high-density lipoprotein and low-density lipoprotein isolated from rapidly salted separated egg yolk.

According to previous research, adding CaCl2 to the salting solution improves the quality of salted separated egg yolk. To further understand the improvement mechanism of CaCl2, this paper investigated the effect of CaCl2 on the structure of high-density lipoprotein (HDL) and low-density lipoprotein (LDL) during the salting process. The results indicated that the addition of CaCl2 can affect the composition of HDL and LDL apolipoproteins, improving the orderliness of the HDL structure and the looseness of the LDL structure. It was discovered by atomic force microscopy (AFM) that adding CaCl2 to the salting solution can weaken the aggregation behavior of HDL. Simultaneously, the addition of CaCl2 decreased the relative content of intermolecular ß-sheets in the secondary structure of HDL and LDL, influenced their tertiary conformation, and prevented HDL and LDL from participating in the formation of a three-dimensional gel structure by influencing their hydrogen bonds and hydrophobic interactions.