Innovative technologies for the fabrication of 3D/4D smart hydrogels and its biomedical applications - A comprehensive review.

Repairing and regenerating damaged tissues or organs, and restoring their functioning has been the ultimate aim of medical innovations. 'Reviving healthcare' blends tissue engineering with alternative techniques such as hydrogels, which have emerged as vital tools in modern medicine. Additive manufacturing (AM) is a practical manufacturing revolution that uses building strategies like molding as a viable solution for precise hydrogel manufacturing. Recent advances in this technology have led to the successful manufacturing of hydrogels with enhanced reproducibility, accuracy, precision, and ease of fabrication. Hydrogels continue to metamorphose as the vital compatible bio-ink matrix for AM. AM hydrogels have paved the way for complex 3D/4D hydrogels that can be loaded with drugs or cells. Bio-mimicking 3D cell cultures designed via hydrogel-based AM is a groundbreaking in-vivo assessment tool in biomedical trials. This brief review focuses on preparations and applications of additively manufactured hydrogels in the biomedical spectrum, such as targeted drug delivery, 3D-cell culture, numerous regenerative strategies, biosensing, bioprinting, and cancer therapies. Prevalent AM techniques like extrusion, inkjet, digital light processing, and stereo-lithography have been explored with their setup and methodology to yield functional hydrogels. The perspectives, limitations, and the possible prospects of AM hydrogels have been critically examined in this study.

Increased risk of chronic fatigue syndrome following infection: a 17-year population-based cohort study.

BACKGROUND: Previous serological studies have indicated an association between viruses and atypical pathogens and Chronic Fatigue Syndrome (CFS). This study aims to investigate the correlation between infections from common pathogens, including typical bacteria, and the subsequent risk of developing CFS. The analysis is based on data from Taiwan's National Health Insurance Research Database. METHODS: From 2000 to 2017, we included a total of 395,811 cases aged 20 years or older newly diagnosed with infection. The cases were matched 1:1 with controls using a propensity score and were followed up until diagnoses of CFS were made. RESULTS: The Cox proportional hazards regression analysis was used to estimate the relationship between infection and the subsequent risk of CFS. The incidence density rates among non-infection and infection population were 3.67 and 5.40 per 1000 person-years, respectively (adjusted hazard ratio [HR] = 1.5, with a 95% confidence interval [CI] 1.47-1.54). Patients infected with Varicella-zoster virus, Mycobacterium tuberculosis, Escherichia coli, Candida, Salmonella, Staphylococcus aureus and influenza virus had a significantly higher risk of CFS than those without these pathogens (p < 0.05). Patients taking doxycycline, azithromycin, moxifloxacin, levofloxacin, or ciprofloxacin had a significantly lower risk of CFS than patients in the corresponding control group (p < 0.05). CONCLUSION: Our population-based retrospective cohort study found that infection with common pathogens, including bacteria, viruses, is associated with an increased risk of developing CFS.

Trajectory in biological metal-organic frameworks: Biosensing and sustainable strategies-perspectives and challenges.

The ligand attribute of biomolecules to form coordination bonds with metal ions led to the discovery of a novel class of materials called biomolecule-associated metal-organic frameworks (Bio-MOFs). These biomolecules coordinate in multiple ways and provide versatile applications. Far-spread bio-ligands include nucleobases, amino acids, peptides, cyclodextrins, saccharides, porphyrins/metalloporphyrin, proteins, etc. Low-toxicity, self-assembly, stability, designable and selectable porous size, the existence of rigid and flexible forms, bio-compatibility, and synergistic interactions between metal ions have led Bio-MOFs to be commercialized in industries such as sensors, food, pharma, and eco-sensing. The rapid growth and commercialization are stunted by absolute bio-compatibility issues, bulk morphology that makes it rigid to alter shape/porosity, longer reaction times, and inadequate research. This review elucidates the structural vitality, biocompatibility issues, and vital sensing applications, including challenges for incorporating bio-ligands into MOF. Critical innovations in Bio-MOFs' applicative spectrum, including sustainable food packaging, biosensing, insulin and phosphoprotein detection, gas sensing, CO2 capture, pesticide carriers, toxicant adsorptions, etc., have been elucidated. Emphasis is placed on biosensing and biomedical applications with biomimetic catalysis and sensitive sensor designing.

Advanced Detection Method for Dengue NS1 Protein Using Ultrasensitive ELISA with Thio-NAD Cycling.

Dengue fever, a mosquito-borne disease in tropical and subtropical climates caused by the dengue virus (DENV), has become a major social and economic burden in recent years. However, current primary detection methods are inadequate for early diagnosis of DENV because they are either time-consuming, expensive, or require training. Non-structural protein 1 (NS1) is secreted during DENV infection and is thus considered a suitable biomarker for the development of an early detection method. In the present study, we developed a detection method for the NS1 protein based on a previously reported thio-NAD cycling ELISA (i.e., ultrasensitive ELISA) and successfully achieved a LOD of 1.152 pg/mL. The clinical diagnosis potential of the detection system was also evaluated by using 85 patient specimens, inclusive of 60 DENV-positive and 25 DENV-negative specimens confirmed by the NAAT method. The results revealed 98.3% (59/60) sensitivity and 100% (25/25) specificity, which was in almost perfect agreement with the NAAT data with a kappa coefficient of 0.972. The present study demonstrates the diagnostic potential of using an ultrasensitive ELISA as a low-cost, easy-to-use method for the detection of DENV compared with NAAT and could be of great benefit in low-income countries.

SPECT/CT imaging for tracking subendothelial retention of electronegative low-density lipoprotein in vivo.

The fifth subfraction of low-density lipoprotein (L5 LDL) can be separated from human LDL using fast-protein liquid chromatography with an anion exchange column. L5 LDL induces vascular endothelial injury both in vitro and in vivo through the lectin-like oxidized LDL receptor-1 (LOX-1). However, no in vivo evidence shows the tendency of L5 LDL deposition on vascular endothelium and links to dysfunction. This study aimed to investigate L5 LDL retention in vivo using SPECT/CT imaging, with Iodine-131 (131I)-labeled and injected into six-month-old apolipoprotein E knockout (apoE-/-) mice through tail veins. Besides, we examined the biodistribution of L5 LDL in tissues and analyzed the intracellular trafficking in human aortic endothelial cells (HAoECs) by confocal microscopy. The impacts of L5 LDL on HAoECs were analyzed using electron microscopy for mitochondrial morphology and western blotting for signaling. Results showed 131I-labeled-L5 was preferentially deposited in the heart and vessels compared to L1 LDL. Furthermore, L5 LDL was co-localized with the mitochondria and associated with mitofusin (MFN1/2) and optic atrophy protein 1 (OPA1) downregulation, leading to mitochondrial fission. In summary, L5 LDL exhibits a propensity for subendothelial retention, thereby promoting endothelial dysfunction and the formation of atherosclerotic lesions.

Untargeted Lipidomic Profiling Reveals Lysophosphatidylcholine and Ceramide as Atherosclerotic Risk Factors in apolipoprotein E Knockout Mice.

Despite the availability and use of numerous cholesterol-lowering drugs, atherosclerotic cardiovascular disease (ASCVD) remains the leading cause of mortality globally. Many researchers have focused their effort on identifying modified lipoproteins. However, lipid moieties such as lysophosphatidylcholine (LPC) and ceramide (CER) contribute to atherogenic events. LPC and CER both cause endothelial mitochondrial dysfunction, leading to fatty acid and triglyceride (TG) accumulation. In addition, they cause immune cells to differentiate into proinflammatory phenotypes. To uncover alternative therapeutic approaches other than cholesterol- and TG-lowering medications, we conducted untargeted lipidomic investigations to assess the alteration of lipid profiles in apolipoprotein E knockout (apoE-/-) mouse model, with or without feeding a high-fat diet (HFD). Results indicated that, in addition to hypercholesterolemia and hyperlipidemia, LPC levels were two to four times higher in apoE-/- mice compared to wild-type mice in C57BL/6 background, regardless of whether they were 8 or 16 weeks old. Sphingomyelin (SM) and CER were elevated three- to five-fold in apoE-/- mice both at the basal level and after 16 weeks when compared to wild-type mice. After HFD treatment, the difference in CER levels elevated more than ten-fold. Considering the atherogenic properties of LPC and CER, they may also contribute to the early onset of atherosclerosis in apoE-/- mice. In summary, the HFD-fed apoE-/- mouse shows elevated LPC and CER contents and is a suitable model for developing LPC- and CER-lowering therapies.

Elucidating the Implications of Norovirus N- and O-Glycosylation, O-GlcNAcylation, and Phosphorylation.

Norovirus is the most common cause of foodborne gastroenteritis, affecting millions of people worldwide annually. Among the ten genotypes (GI-GX) of norovirus, only GI, GII, GIV, GVIII, and GIX infect humans. Some genotypes reportedly exhibit post-translational modifications (PTMs), including N- and O-glycosylation, O-GlcNAcylation, and phosphorylation, in their viral antigens. PTMs have been linked to increased viral genome replication, viral particle release, and virulence. Owing to breakthroughs in mass spectrometry (MS) technologies, more PTMs have been discovered in recent years and have contributed significantly to preventing and treating infectious diseases. However, the mechanisms by which PTMs act on noroviruses remain poorly understood. In this section, we outline the current knowledge of the three common types of PTM and investigate their impact on norovirus pathogenesis. Moreover, we summarize the strategies and techniques for the identification of PTMs.

The Critical Role of Galectin-12 in Modulating Lipid Metabolism in Sebaceous Glands.

Sebaceous glands play an important role in maintaining the skin barrier function by producing lipids. Dysregulated lipid production in these glands may contribute to the pathogenesis of human skin diseases. Galectin-12, a member of the ß-galactoside‒binding lectin family, is preferentially expressed in adipocytes, where it regulates adipogenesis and functions as an intrinsic negative regulator of lipolysis. It is also expressed by sebocytes and contributes to the proliferation of this cell type. In this study, we show the association between galectin-12 expression and sebocyte differentiation. Galectin-12 knockdown in a human sebocyte cell line reduced lipogenesis and decreased the production of cholesteryl esters, triglycerides, free fatty acids, and cholesterol. Metabolomic analysis of skin surface lipids showed that the levels of the lipids mentioned earlier decreased in sebaceous gland‒specific galectin-12‒knockout mice compared with that in wild-type mice. In addition, galectin-12 positively regulated peroxisome proliferator‒activated receptor-γ transcriptional activity in sebocytes stimulated with fatty acids. Downregulating galectin-12 suppressed the expression of peroxisome proliferator‒activated receptor-γ target genes-acetyl-coenzyme A synthetase 2 gene ACS2 and diacylglycerol O-acyltransferase 1 gene DGAT1-that are required for fatty acid activation and cholesterol and triglyceride biosynthesis. In conclusion, galectin-12 is a positive regulator of sebaceous lipid metabolism with a potential role in the maintenance of skin homeostasis.

Galectin-12 modulates sebocyte proliferation and cell cycle progression by regulating cyclin A1 and CDK2.

Enhanced sebocyte proliferation is associated with the pathogenesis of human skin diseases related to sebaceous gland hyperfunction and androgens, which are known to induce sebocyte proliferation, are key mediators of this process. Galectin-12, a member of the ß-galactoside-binding lectin family that is preferentially expressed by adipocytes and functions as an intrinsic negative regulator of lipolysis, has been shown to be expressed by human sebocytes. In this study, we identified galectin-12 as an important intracellular regulator of sebocyte proliferation. Galectin-12 knockdown in the human SZ95 sebocyte line suppressed cell proliferation, and its overexpression promoted cell cycle progression. Inhibition of galectin-12 expression reduced the androgen-induced SZ95 sebocyte proliferation and growth of sebaceous glands in mice, respectively. The mRNA expression of the key cell cycle regulators cyclin A1 (CCNA1) and cyclin-dependent kinase 2CDK2 was reduced in galectin-12 knockdown SZ95 sebocytes, suggesting a pathway of galectin-12 regulation of sebocyte proliferation. Further, galectin-12 enhanced peroxisome proliferator-activated receptor gamma (PPARγ) expression and transcriptional activity in SZ95 sebocytes, consistent with our previous studies in adipocytes. Rosiglitazone, a PPARγ ligand, induced CCNA1 levels, suggesting that galectin-12 may upregulate CCNA1 expression via PPARγ. Our findings suggest the possibility of targeting galectin-12 to treat human sebaceous gland hyperfunction and androgen-associated skin diseases.

Mass Spectrometry-Based System for Identifying and Typing Norovirus Major Capsid Protein VP1.

Norovirus-associated diseases are the most common foodborne illnesses worldwide. Polymerase chain reaction-based methods are the primary diagnostics for clinical samples; however, the high mutation rate of norovirus makes viral amplification and genotyping challenging. Technological advances in mass spectrometry (MS) make it a promising tool for identifying disease markers. Besides, the superior sensitivity of MS and proteomic approaches may enable the detection of all variants. Thus, this study aimed to establish an MS-based system for identifying and typing norovirus. We constructed three plasmids containing the major capsid protein VP1 of the norovirus GII.4 2006b, 2006a, and 2009a strains to produce virus-like particles for use as standards. Digested peptide signals were collected using a nano-flow ultra-performance liquid chromatography mass spectrometry (nano-UPLC/MSE) system, and analyzed by ProteinLynx Global SERVER and TREE-PUZZLE software. Results revealed that the LC/MSE system had an excellent coverage rate: the system detected more than 94% of amino acids of 3.61 femtomole norovirus VP1 structural protein. In the likelihood-mapping analysis, the proportions of unresolved quartets were 2.9% and 4.9% in the VP1 and S domains, respectively, which is superior to the 15.1% unresolved quartets in current PCR-based methodology. In summary, the use of LC/MSE may efficiently monitor genotypes, and sensitively detect structural and functional mutations of noroviruses.

Electronegative very-low-density lipoprotein induces brain inflammation and cognitive dysfunction in mice.

Epidemiologic studies have indicated that dyslipidemia may facilitate the progression of cognitive dysfunction. We previously showed that patients with metabolic syndrome (MetS) had significantly higher plasma levels of electronegative very-low-density lipoprotein (VLDL) than did healthy controls. However, the effects of electronegative-VLDL on the brain and cognitive function remain unclear. In this study, VLDL isolated from healthy volunteers (nVLDL) or patients with MetS (metVLDL) was administered to mice by means of tail vein injection. Cognitive function was assessed by using the Y maze test, and plasma and brain tissues were analyzed. We found that mice injected with metVLDL but not nVLDL exhibited significant hippocampus CA3 neuronal cell loss and cognitive dysfunction. In mice injected with nVLDL, we observed mild glial cell activation in the medial prefrontal cortex (mPFC) and hippocampus CA3. However, in mice injected with metVLDL, plasma and brain TNF-α and Aß-42 levels and glial cell activation in the mPFC and whole hippocampus were higher than those in control mice. In conclusion, long-term exposure to metVLDL induced levels of TNF-α, Aß-42, and glial cells in the brain, contributing to the progression of cognitive dysfunction. Our findings suggest that electronegative-VLDL levels may represent a new therapeutic target for cognitive dysfunction.

Adiponectin forms a complex with atherogenic LDL and inhibits its downstream effects.

Adiponectin, an adipocyte-derived protein, has antiatherogenic and antidiabetic effects, but how it confers the atherogenic effects is not well known. To study the antiatherogenic mechanisms of adiponectin, we examined whether it interacts with atherogenic low density lipoprotein (LDL) to attenuate LDL's atherogenicity. L5, the most electronegative subfraction of LDL, induces atherogenic responses similarly to copper-oxidized LDL (oxLDL). Unlike the native LDL endocytosed via the LDL receptor, L5 and oxLDL are internalized by cells via the lectin-like oxidized LDL receptor-1 (LOX-1). Using enzyme-linked immunosorbent assays (ELISAs), we showed that adiponectin preferentially bound oxLDL but not native LDL. In Chinese hamster ovary (CHO) cells transfected with the LOX-1 or LDL receptor, adiponectin selectively inhibited the uptake of oxLDL but not of native LDL, respectively. Furthermore, adiponectin suppressed the internalization of oxLDL in human coronary artery endothelial cells (HCAECs) and THP-1-derived macrophages. Western blot analysis of human plasma showed that adiponectin was abundant in L5 but not in L1, the least electronegative subfraction of LDL. Sandwich ELISAs with anti-adiponectin and anti-apolipoprotein B antibodies confirmed the binding of adiponectin to L5 and oxLDL. In LOX-1-expressing CHO cells, adiponectin inhibited cellular responses to oxLDL and L5, including nuclear factor-κB activation and extracellular signal-regulated kinas phosphorylation. In HCAECs, adiponectin inhibited oxLDL-induced endothelin-1 secretion and extracellular signal-regulated kinase phosphorylation. Conversely, oxLDL suppressed the adiponectin-induced activation of adenosine monophosphate-activated protein kinase in COS-7 cells expressing adiponectin receptor AdipoR1. Our findings suggest that adiponectin binds and inactivates atherogenic LDL, providing novel insight into the antiatherogenic mechanisms of adiponectin.

Molecular and Cellular Mechanisms of Electronegative Lipoproteins in Cardiovascular Diseases.

Dysregulation of glucose and lipid metabolism increases plasma levels of lipoproteins and triglycerides, resulting in vascular endothelial damage. Remarkably, the oxidation of lipid and lipoprotein particles generates electronegative lipoproteins that mediate cellular deterioration of atherosclerosis. In this review, we examined the core of atherosclerotic plaque, which is enriched by byproducts of lipid metabolism and lipoproteins, such as oxidized low-density lipoproteins (oxLDL) and electronegative subfraction of LDL (LDL(-)). We also summarized the chemical properties, receptors, and molecular mechanisms of LDL(-). In combination with other well-known markers of inflammation, namely metabolic diseases, we concluded that LDL(-) can be used as a novel prognostic tool for these lipid disorders. In addition, through understanding the underlying pathophysiological molecular routes for endothelial dysfunction and inflammation, we may reassess current therapeutics and might gain a new direction to treat atherosclerotic cardiovascular diseases, mainly targeting LDL(-) clearance.

An Increased Plasma Level of ApoCIII-Rich Electronegative High-Density Lipoprotein May Contribute to Cognitive Impairment in Alzheimer's Disease.

High-density lipoprotein (HDL) plays a vital role in lipid metabolism and anti-inflammatory activities; a dysfunctional HDL impairs cholesterol efflux pathways. To understand HDL's role in patients with Alzheimer's disease (AD), we analyzed the chemical properties and function. HDL from AD patients (AD-HDL) was separated into five subfractions, H1-H5, using fast-protein liquid chromatography equipped with an anion-exchange column. Subfraction H5, defined as the most electronegative HDL, was increased 5.5-fold in AD-HDL (23.48 ± 17.83%) in comparison with the control HDL (4.24 ± 3.22%). By liquid chromatography mass spectrometry (LC/MSE), AD-HDL showed that the level of apolipoprotein (apo)CIII was elevated but sphingosine-1-phosphate (S1P)-associated apoM and anti-oxidative paraoxonase 1 (PON1) were reduced. AD-HDL showed a lower cholesterol efflux capacity that was associated with the post-translational oxidation of apoAI. Exposure of murine macrophage cell line, RAW 264.7, to AD-HDL induced a vibrant expression of ganglioside GM1 in colocalization with apoCIII on lipid rafts alongside a concomitant increase of tumor necrosis factor-α (TNF-α) detectable in the cultured medium. In conclusion, AD-HDL had a higher proportion of H5, an apoCIII-rich electronegative HDL subfraction. The associated increase in pro-inflammatory (apoCIII, TNF-α) components might favor Amyloid ß assembly and neural inflammation. A compromised cholesterol efflux capacity of AD-HDL may also contribute to cognitive impairment.

Disruption of retinoid homeostasis induces RBP4 overproduction in diabetes: O-GlcNAcylation involved.

BACKGROUND: Retinol-binding protein 4 (RBP4) is elevated and associated with inflammation in metabolic diseases. Disruption of the retinol cascade and O-GlcNAcylation of the RBP4 receptor (STRA6) are found in diabetic kidneys. OBJECTIVES: We investigated whether the disruption of the retinol cascade induces RBP4 overproduction and if O-linked GlcNAc modification targets RBPR2 and contributes to the disruption of retinol cascades in diabetic livers. METHODS: Western blot or immunohistochemistry for RBPR2, CRBP1, LRAT, RALDH, RARα, RARγ, RXRα, RBP4, GFAT, OGT, OGA and inflammatory markers, as well as ELISA for RBP4, were performed in livers of db/db and ob/ob mice and high glucose-cultured hepatocytes. Immunoprecipitation and dual fluorescence staining were used to explore O-GlcNAc-modified RBPR2 and RBP4 binding activity on RBPR2. Transfection of the CRBP1 gene was done to verify whether a disrupted retinol cascade induces RBP4 overproduction. OGT silencing was done to investigate the association of O-GlcNAcylation with the disruption of retinol cascade. RESULTS: Disruption of retinol cascade, RBP4 overproduction, O-GlcNAcylation of RBPR2, decreased RBP4 binding activity on RBPR2 and inflammation were found in livers of db/db and ob/ob mice and high glucose-cultured hepatocytes. CRBP1 gene transfection reversed the suppression of the cellular retinol cascade and simultaneously attenuated the RBP4 overproduction and inflammation in high glucose-treated hepatocytes. The silencing of OGT reversed the disruption of the cellular retinol cascade, RBP4 overproduction and inflammation induced by high glucose in hepatocytes. CONCLUSIONS: This study indicates that the disruption of cellular retinol cascade is strongly associated with RBP4 overproduction and inflammation in diabetic livers. RBPR2 is one target for high glucose-mediated O-linked GlcNAc modification, which causes liver retinol dyshomeostasis.

The role of postprandial very-low-density lipoprotein in the development of atrial remodeling in metabolic syndrome.

BACKGROUND: Negatively charged very-low-density lipoprotein (VLDL-χ) in metabolic syndrome (MetS) patients exerts cytotoxic effects on endothelial cells and atrial myocytes. Atrial cardiomyopathy, manifested by atrial remodeling with a dilated diameter, contributes to atrial fibrillation pathogenesis and predicts atrial fibrillation development. The correlation of VLDL-χ with atrial remodeling is unknown. This study investigated the association between VLDL-χ and remodeling of left atrium. METHODS: Consecutively, 87 MetS and 80 non-MetS individuals between 23 and 74 years old (50.6% men) without overt cardiovascular diseases were included in the prospective cohort study. Blood samples were collected while fasting and postprandially (at 0.5, 1, 2, and 4 h after a unified meal). VLDL was isolated by ultracentrifugation; the percentile concentration of VLDL-χ (%) was determined by ultra-performance liquid chromatography. The correlations of left atrium diameter (LAD) with variables including VLDL-χ, LDL-C, HDL-C, triglycerides, glucose, and blood pressure, were analyzed by multiple linear regression models. A hierarchical linear model was conducted to test the independencies of each variable's correlation with LAD. RESULTS: The mean LAD was 3.4 ± 0.5 cm in non-MetS subjects and 3.9 ± 0.5 cm in MetS patients (P < 0.01). None of the fasting lipid profiles were associated with LAD. VLDL-χ, BMI, waist circumference, hip circumference, and blood pressure were positively correlated with LAD (all P < 0.05) after adjustment for age and sex. Significant interactions between VLDL-χ and blood pressure, waist circumference, and hip circumference were observed. When adjusted for obesity- and blood pressure-related variables, 2-h postprandial VLDL-χ (mean 1.30 ± 0.61%) showed a positive correlation with LAD in MetS patients. Each 1% VLDL-χ increase was estimated to increase LAD by 0.23 cm. CONCLUSIONS: Postprandial VLDL-χ is associated with atrial remodeling particularly in the MetS group. VLDL-χ is a novel biomarker and may be a therapeutic target for atrial cardiomyopathy in MetS patients. TRIAL REGISTRATION: ISRCTN 69295295 . Retrospectively registered 9 June 2020.

Clinical Significance of Electronegative Low-Density Lipoprotein Cholesterol in Atherothrombosis.

Despite the numerous risk factors for atherosclerotic cardiovascular diseases (ASCVD), cumulative evidence shows that electronegative low-density lipoprotein (L5 LDL) cholesterol is a promising biomarker. Its toxicity may contribute to atherothrombotic events. Notably, plasma L5 LDL levels positively correlate with the increasing severity of cardiovascular diseases. In contrast, traditional markers such as LDL-cholesterol and triglyceride are the therapeutic goals in secondary prevention for ASCVD, but that is controversial in primary prevention for patients with low risk. In this review, we point out the clinical significance and pathophysiological mechanisms of L5 LDL, and the clinical applications of L5 LDL levels in ASCVD can be confidently addressed. Based on the previously defined cut-off value by receiver operating characteristic curve, the acceptable physiological range of L5 concentration is proposed to be below 1.7 mg/dL. When L5 LDL level surpass this threshold, clinically relevant ASCVD might be present, and further exams such as carotid intima-media thickness, pulse wave velocity, exercise stress test, or multidetector computed tomography are required. Notably, the ultimate goal of L5 LDL concentration is lower than 1.7 mg/dL. Instead, with L5 LDL greater than 1.7 mg/dL, lipid-lowering treatment may be required, including statin, ezetimibe or PCSK9 inhibitor, regardless of the low-density lipoprotein cholesterol (LDL-C) level. Since L5 LDL could be a promising biomarker, we propose that a high throughput, clinically feasible methodology is urgently required not only for conducting a prospective, large population study but for developing therapeutics strategies to decrease L5 LDL in the blood.

Effect of acyl and alkyl analogs of platelet-activating factor on inflammatory signaling.

Platelet-activating factor (PAF), a bioactive ether phospholipid with significant pro-inflammatory properties, was identified almost half a century ago. Despite extensive study of this autocoid, therapeutic strategies for targeting its signaling components have not been successful, including the recent clinical trials with darapladib, a drug that targets plasma PAF-acetylhydrolase (PAF-AH). We recently provided experimental evidence that the previously unrecognized acyl analog of PAF, which is concomitantly produced along with PAF during biosynthesis, dampens PAF signaling by acting both as a sacrificial substrate for PAF-AH and probably as an endogenous PAF-receptor antagonist/partial agonist. If this is the scenario in vivo, PAF-AH needs to catalyze the selective hydrolysis of alkyl-PAF and not acyl-PAF. Accordingly, different approaches are needed for treating inflammatory diseases in which PAF signaling is implicated. The interplay between acyl-PAF, alkyl-PAF, PAF-AH, and PAF-R is complex, and the outcome of this interplay has not been previously appreciated. In this review, we discuss this interaction based on our recent findings. It is very likely that the relative abundance of acyl and alkyl-PAF and their interactions with PAF-R in the presence of their hydrolyzing enzyme PAF-AH may exert a modulatory effect on PAF signaling during inflammation.

Increased APOE glycosylation plays a key role in the atherogenicity of L5 low-density lipoprotein.

Low-density lipoprotein (LDL) is heterogeneous, composed of particles with variable atherogenicity. Electronegative L5 LDL exhibits atherogenic properties in vitro and in vivo, and its levels are elevated in patients with increased cardiovascular risk. Apolipoprotein E (APOE) content is increased in L5, but what role APOE plays in L5 function remains unclear. Here, we characterized the contributions of APOE posttranslational modification to L5's atherogenicity. Using two-dimensional electrophoresis and liquid chromatography-mass spectrometry, we studied APOE's posttranslational modification in L5 from human plasma. APOE structures with various glycan residues were predicted. Molecular docking and molecular dynamics simulation were performed to examine the functional changes of APOE resulting from glycosylation. We also examined the effects of L5 deglycosylation on endothelial cell apoptosis. The glycan sequence N-acetylgalactosamine, galactose, and sialic acid was consistently expressed on serine 94, threonine 194, and threonine 289 of APOE in L5 and was predicted to contribute to L5's negative surface charge and hydrophilicity. The electrostatic force between the negatively charged sialic acid-containing glycan residue of APOE and positively charged amino acids at the receptor-binding area suggested that glycosylation interferes with APOE's attraction to receptors, lipid-binding ability, and lipid transportation and metabolism functions. Importantly, L5 containing glycosylated APOE induced apoptosis in cultured endothelial cells through lectin-like oxidized LDL receptor-1 (LOX-1) signaling, and glycosylation removal from L5 attenuated L5-induced apoptosis. APOE glycosylation may contribute to the atherogenicity of L5 and be a useful biomarker for rapidly quantifying L5.

Identifying the Therapeutic Significance of Mesenchymal Stem Cells.

The pleiotropic behavior of mesenchymal stem cells (MSCs) has gained global attention due to their immense potential for immunosuppression and their therapeutic role in immune disorders. MSCs migrate towards inflamed microenvironments, produce anti-inflammatory cytokines and conceal themselves from the innate immune system. These signatures are the reason for the uprising in the sciences of cellular therapy in the last decades. Irrespective of their therapeutic role in immune disorders, some factors limit beneficial effects such as inconsistency of cell characteristics, erratic protocols, deviating dosages, and diverse transfusion patterns. Conclusive protocols for cell culture, differentiation, expansion, and cryopreservation of MSCs are of the utmost importance for a better understanding of MSCs in therapeutic applications. In this review, we address the immunomodulatory properties and immunosuppressive actions of MSCs. Also, we sum up the results of the enhancement, utilization, and therapeutic responses of MSCs in treating inflammatory diseases, metabolic disorders and diabetes.