Designing Efficient Enzymes: Eight Predicted Mutations Convert a Hydroxynitrile Lyase into an Efficient Esterase.

Hydroxynitrile lyase from rubber tree (HNL) shares 45% identical amino acid residues with the homologous esterase from tobacco, SABP2, but the two enzymes catalyze different reactions. The x-ray structures reveal a serine-histidine-aspartate catalytic triad in both enzymes along with several differing amino acid residues within the active site. Previous exchange of three amino acid residues in the active site of HNL with the corresponding amino acid residue in SABP2 (T11G-E79H-K236M) created variant HNL3, which showed low esterase activity toward p-nitrophenyl acetate.

ATP-Binding Cassette Transporter Family C Protein 10 Participates in the Synthesis and Efflux of Hexosylceramides in Liver Cells.

In addition to sphingomyelin and ceramide, sugar derivatives of ceramides, hexosylceramides (HexCer) are the major circulating sphingolipids. We have shown that silencing of ABCA1 transmembrane protein function for instance in cases of loss of function of ABCA1 gene results in low levels of HDL as well as a concomitant reduction in plasma HexCer levels. However, proteins involved in hepatic synthesis and egress of HexCer from cells is not well known although ABCA1 seems to be indirectly controlling the HexCer plasma levels by supporting HDL synthesis.

ATP-binding cassette protein ABCA7 deficiency impairs sphingomyelin synthesis, cognitive discrimination, and synaptic plasticity in the entorhinal cortex.

Sphingomyelin (SM) is an abundant plasma membrane and plasma lipoprotein sphingolipid. We previously reported that ATP-binding cassette family A protein 1 (ABCA1) deficiency in humans and mice decreases plasma SM levels. However, overexpression, induction, downregulation, inhibition, and knockdown of ABCA1 in human hepatoma Huh7 cells did not decrease SM efflux.

Discovery and Characterization of a Novel Aryl Hydrocarbon Receptor Inhibitor, IK-175, and Its Inhibitory Activity on Tumor Immune Suppression.

Aryl hydrocarbon receptor (AHR) is a transcription factor that regulates the activity of multiple innate and adaptive immune cells subsequent to binding to numerous endogenous and exogenous ligands. For example, AHR is activated by the metabolite kynurenine, which is secreted into the tumor microenvironment by cancer cells leading to broad immunosuppression. Therefore, AHR inhibition provides a novel and ideal approach to stimulate immune-mediated recognition and subsequent eradication of tumor cells.

The Significance and Impact of Screening Preservation Fluid Cultures in Renal Transplant Recipients.

Introduction: Infectious complications can be a major cause of morbidity and mortality in solid organ transplant recipients. Preservation fluid is necessary to maintain organ viability but may serve as a vector or infection. The utility of screening preservation fluid routinely for microbial growth and the impact of culture-positive preservation fluid is controversial.

Model systems for studying the assembly, trafficking, and secretion of apoB lipoproteins using fluorescent fusion proteins.

apoB exists as apoB100 and apoB48, which are mainly found in hepatic VLDLs and intestinal chylomicrons, respectively. Elevated plasma levels of apoB-containing lipoproteins (Blps) contribute to coronary artery disease, diabetes, and other cardiometabolic conditions. Studying the mechanisms that drive the assembly, intracellular trafficking, secretion, and function of Blps remains challenging.

ATP binding cassette family A protein 1 determines hexosylceramide and sphingomyelin levels in human and mouse plasma.

Sphingolipids, including ceramide, SM, and hexosylceramide (HxCer), are carried in the plasma by lipoproteins. They are possible markers of metabolic diseases, but little is known about their control. We previously showed that microsomal triglyceride transfer protein (MTP) is critical to determine plasma ceramide and SM, but not HxCer, levels.

Sphingolipids and Lipoproteins in Health and Metabolic Disorders.

Sphingolipids are structurally and functionally diverse molecules with significant physiologic functions and are found associated with cellular membranes and plasma lipoproteins. The cellular and plasma concentrations of sphingolipids are altered in several metabolic disorders and may serve as prognostic and diagnostic markers. Here we discuss various sphingolipid transport mechanisms and highlight how changes in cellular and plasma sphingolipid levels contribute to cardiovascular disease, obesity, diabetes, insulin resistance, and nonalcoholic fatty liver disease (NAFLD).

Hepatic Tm6sf2 overexpression affects cellular ApoB-trafficking, plasma lipid levels, hepatic steatosis and atherosclerosis.

The human transmembrane 6 superfamily member 2 (TM6SF2) gene has been implicated in plasma lipoprotein metabolism, alcoholic and non-alcoholic fatty liver disease and myocardial infarction in multiple genome-wide association studies. To investigate the role of Tm6sf2 in metabolic homeostasis, we generated mice with elevated expression using adeno-associated virus (AAV)-mediated gene delivery. Hepatic overexpression of mouse Tm6sf2 resulted in phenotypes previously observed in Tm6sf2-deficient mice including reduced plasma lipid levels, diminished hepatic triglycerides secretion and increased hepatosteatosis.

Targeting microsomal triglyceride transfer protein and lipoprotein assembly to treat homozygous familial hypercholesterolemia.

Homozygous familial hypercholesterolemia (HoFH) is a polygenic disease arising from defects in the clearance of plasma low-density lipoprotein (LDL), which results in extremely elevated plasma LDL cholesterol (LDL-C) and increased risk of atherosclerosis, coronary heart disease, and premature death. Conventional lipid-lowering therapies, such as statins and ezetimibe, are ineffective at lowering plasma cholesterol to safe levels in these patients. Other therapeutic options, such as LDL apheresis and liver transplantation, are inconvenient, costly, and not readily available.

Structure-function analyses of microsomal triglyceride transfer protein missense mutations in abetalipoproteinemia and hypobetalipoproteinemia subjects.

We describe two new hypolipidemic patients with very low plasma triglyceride and apolipoprotein B (apoB) levels with plasma lipid profiles similar to abetalipoproteinemia (ABL) patients. In these patients, we identified two previously uncharacterized missense mutations in the microsomal triglyceride transfer protein (MTP) gene, R46G and D361Y, and studied their functional effects. We also characterized three missense mutations (H297Q, D384A, and G661A) reported earlier in a familial hypobetalipoproteinemia patient.

Microsomal Triglyceride Transfer Protein Transfers and Determines Plasma Concentrations of Ceramide and Sphingomyelin but Not Glycosylceramide.

Sphingolipids, a large family of bioactive lipids, are implicated in stress responses, differentiation, proliferation, apoptosis, and other physiological processes. Aberrant plasma levels of sphingolipids contribute to metabolic disease, atherosclerosis, and insulin resistance. They are fairly evenly distributed in high density and apoB-containing lipoproteins (B-lps).

Novel Abetalipoproteinemia Missense Mutation Highlights the Importance of the N-Terminal β-Barrel in Microsomal Triglyceride Transfer Protein Function.

Background: The use of microsomal triglyceride transfer protein (MTP) inhibitors is limited to severe hyperlipidemias because of associated hepatosteatosis and gastrointestinal adverse effects. Comprehensive knowledge about the structure-function of MTP might help design new molecules that avoid steatosis. Characterization of mutations in MTP causing abetalipoproteinemia has revealed that the central α-helical and C-terminal β-sheet domains are important for protein disulfide isomerase binding and lipid transfer activity.

Found in transition: applying milestones to three unique discharge curricula.

Introduction. A safe and effective transition from hospital to post-acute care is a complex and important physician competency. Milestones and Entrustable Professional Activities (EPA) form the new educational rubric in Graduate Medical Education Training.

Ankle Dorsiflexion Displacement During Landing is Associated With Initial Contact Kinematics but not Joint Displacement.

The ankle, knee, and hip joints work together in the sagittal plane to absorb landing forces. Reduced sagittal plane motion at the ankle may alter landing strategies at the knee and hip, potentially increasing injury risk; however, no studies have examined the kinematic relationships between the joints during jump landings. Healthy adults (N = 30; 15 male, 15 female) performed jump landings onto a force plate while three-dimensional kinematic data were collected.

Ω-3 fatty acids prevent hepatic steatosis, independent of PPAR-α activity, in a murine model of parenteral nutrition-associated liver disease.

Objectives: ω-3 Fatty acids (FAs), natural ligands for the peroxisome proliferator-activated receptor-α (PPAR-α), attenuate parenteral nutrition-associated liver disease (PNALD). However, the mechanisms underlying the protective role of ω-3 FAs are still unknown. The aim of this study was to determine the effects of ω-3 FAs on hepatic triglyceride (TG) accumulation in a murine model of PNALD and to investigate the role of PPAR-α and microsomal triglyceride transfer protein (MTP) in this experimental setting.

Loss of both phospholipid and triglyceride transfer activities of microsomal triglyceride transfer protein in abetalipoproteinemia.

Mutations in microsomal triglyceride transfer protein (MTP) cause abetalipoproteinemia (ABL), characterized by the absence of plasma apoB-containing lipoproteins. In this study, we characterized the effects of various MTP missense mutations found in ABL patients with respect to their expression, subcellular location, and interaction with protein disulfide isomerase (PDI). In addition, we characterized functional properties by analyzing phospholipid and triglyceride transfer activities and studied their ability to support apoB secretion.

Lower extremity muscle activation and knee flexion during a jump-landing task.

Context: Decreased sagittal-plane motion at the knee during dynamic tasks has been reported to increase impact forces during landing, potentially leading to knee injuries such as anterior cruciate ligament rupture.

Objective: To describe the relationship between lower extremity muscle activity and knee-flexion angle during a jump-landing task.

Design: Cross-sectional study.

Multiple functions of microsomal triglyceride transfer protein.

Microsomal triglyceride transfer protein (MTP) was first identified as a major cellular protein capable of transferring neutral lipids between membrane vesicles. Its role as an essential chaperone for the biosynthesis of apolipoprotein B (apoB)-containing triglyceride-rich lipoproteins was established after the realization that abetalipoproteinemia patients carry mutations in the MTTP gene resulting in the loss of its lipid transfer activity. Now it is known that it also plays a role in the biosynthesis of CD1, glycolipid presenting molecules, as well as in the regulation of cholesterol ester biosynthesis.

Oxidative stress activates ADAM17/TACE and induces its target receptor shedding in platelets in a p38-dependent fashion.

Aims: Oxidative stress accompanies inflammatory and vascular diseases. The objective of this study was to explore whether reactive oxygen species can activate shedding of platelet receptors and thus suppress platelet function.

Methods And Results: Hydrogen peroxide and glucose oxidase were chosen to model oxidative stress in vitro.

The effect of C1 inhibitor on intestinal ischemia and reperfusion injury.

Complement activation and neutrophil stimulation are two major components in events leading to ischemia and reperfusion (IR) injury. C1 inhibitor (C1INH) inhibits activation of each of the three pathways of complement activation and of the contact system. It is also endowed with anti-inflammatory properties that are independent of protease inhibition.

ADAMTS13: a new link between thrombosis and inflammation.

von Willebrand factor (VWF) levels are elevated and a disintegrin-like and metalloprotease with thrombospondin type I repeats-13 (ADAMTS13) activity is decreased in both acute and chronic inflammation. We hypothesized that by cleaving hyperactive ultralarge VWF (ULVWF) multimers, ADAMTS13 down-regulates both thrombosis and inflammation. Using intravital microscopy, we show that ADAMTS13 deficiency results in increased leukocyte rolling on unstimulated veins and increased leukocyte adhesion in inflamed veins.