Challenges and potential solutions to enrollment in a clinical trial of arteriovenous fistula vs arteriovenous graft vascular access strategy.

This article presents the rationale, challenges, and adaptive strategies employed during the initiation and execution of the arteriovenous (AV) access trial-a multicenter randomized controlled trial (RCT) comparing AV fistulas and AV grafts for hemodialysis in older adults with major comorbidities. Motivated by shifts in epidemiologic landscapes and evolving guidelines moving away from a fistula-first approach and to more patient-centric approaches, the objective of this randomized controlled trial was to fill critical knowledge gaps in determining the optimal vascular access for this complex patient population. We outline the challenges encountered in patient recruitment along with measures employed to overcome these obstacles in recruitment.

Inositol phosphates dynamically enhance stability, solubility, and catalytic activity of mTOR.

Mechanistic target of rapamycin (mTOR) binds the small metabolite inositol hexakisphosphate (IP) as shown in structures of mTOR; however, it remains unclear if IP, or any other inositol phosphate species, function as an integral structural element(s) or catalytic regulator(s) of mTOR. Here, we show that multiple, exogenously added inositol phosphate species can enhance the ability of mTOR and mechanistic target of rapmycin complex 1 (mTORC1) to phosphorylate itself and peptide substrates in in vitro kinase reactions, with the higher order phosphorylated species being more potent (IP = IP > IP >> IP). IP increased the V and decreased the apparent K of mTOR for ATP.

Multiple inositol phosphate species enhance stability of active mTOR.

Mechanistic Target of Rapamycin (mTOR) binds the small metabolite inositol hexakisphosphate (IP) as shown in structures of mTOR, however it remains unclear if IP, or any other inositol phosphate species, can activate mTOR kinase activity. Here, we show that multiple, exogenously added inositol phosphate species (IP, IP, IP and IP) can all enhance the ability of mTOR and mTORC1 to auto-phosphorylate and incorporate radiolabeled phosphate into peptide substrates in kinase reactions. Although IP did not affect the apparent K of mTORC1 for ATP, monitoring kinase activity over longer reaction times showed increased product formation, suggesting inositol phosphates stabilize an active form of mTORC1 .

Coordinated inositide lipid-phosphatase activities of synaptojanin modulates actin cytoskeleton organization.

Synaptojanin proteins are evolutionarily conserved regulators of vesicle transport and membrane homeostasis. Disruption of synaptojanin function has been implicated in a wide range of neurological disorders. Synaptojanins act as dual-functional lipid phosphatases capable of hydrolyzing a variety of phosphoinositides (PIPs) through autonomous SAC1-like PIP 4-phosphatase and PIP 5-phosphatase domains.

Abiotrophia Causing Prosthetic Joint Septic Arthritis.

A 71-year-old Caucasian male with a past medical history of Charcot-Marie-Tooth disease type 2 presented to our rural hospital for left knee pain, swelling, and difficulty walking. The patient had prior bilateral total knee replacements with a subsequent left knee revision due to infection. Joint aspiration was culture-positive and 16S recombinant DNA (rDNA) sequence positive for .

Inhibition of p53 Sulfoconjugation Prevents Oxidative Hepatotoxicity and Acute Liver Failure.

Background & Aims: Sulfoconjugation of small molecules or protein peptides is a key mechanism to ensure biochemical and functional homeostasis in mammals. The PAPS synthase 2 (PAPSS2) is the primary enzyme to synthesize the universal sulfonate donor 3'-phosphoadenosine 5'-phosphosulfate (PAPS). Acetaminophen (APAP) overdose is the leading cause of acute liver failure (ALF), in which oxidative stress is a key pathogenic event, whereas sulfation of APAP contributes to its detoxification.

Bisphosphate nucleotidase 2 (BPNT2), a molecular target of lithium, regulates chondroitin sulfation patterns in the cerebral cortex and hippocampus.

Bisphosphate nucleotidase 2 (BPNT2) is a member of a family of phosphatases that are directly inhibited by lithium, the first-line medication for bipolar disorder. BPNT2 is localized to the Golgi, where it metabolizes the by-products of glycosaminoglycan sulfation reactions. BPNT2-knockout mice exhibit impairments in total-body chondroitin-4-sulfation which lead to abnormal skeletal development (chondrodysplasia).

Sulfation of glycosaminoglycans depends on the catalytic activity of lithium-inhibited phosphatase BPNT2 in vitro.

Golgi-resident bisphosphate nucleotidase 2 (BPNT2) is a member of a family of magnesium-dependent, lithium-inhibited phosphatases that share a three-dimensional structural motif that directly coordinates metal binding to effect phosphate hydrolysis. BPNT2 catalyzes the breakdown of 3'-phosphoadenosine-5'-phosphate, a by-product of glycosaminoglycan (GAG) sulfation. KO of BPNT2 in mice leads to skeletal abnormalities because of impaired GAG sulfation, especially chondroitin-4-sulfation, which is critical for proper extracellular matrix development.

Intestinal Sulfation Is Essential to Protect Against Colitis and Colonic Carcinogenesis.

Background & Aims: Sulfation is a conjugation reaction essential for numerous biochemical and cellular functions in mammals. The 3'-phosphoadenosine 5'-phosphosulfate (PAPS) synthase 2 (PAPSS2) is the key enzyme to generate PAPS, which is the universal sulfonate donor for all sulfation reactions. The goal of this study was to determine whether and how PAPSS2 plays a role in colitis and colonic carcinogenesis.

A structural basis for lithium and substrate binding of an inositide phosphatase.

Inositol polyphosphate 1-phosphatase (INPP1) is a prototype member of metal-dependent/lithium-inhibited phosphomonoesterase protein family defined by a conserved three-dimensional core structure. Enzymes within this family function in distinct pathways including inositide signaling, gluconeogenesis, and sulfur assimilation. Using structural and biochemical studies, we report the effect of substrate and lithium on a network of metal binding sites within the catalytic center of INPP1.

Vip1 is a kinase and pyrophosphatase switch that regulates inositol diphosphate signaling.

Inositol diphosphates (PP-IPs), also known as inositol pyrophosphates, are high-energy cellular signaling codes involved in nutrient and regulatory responses. We report that the evolutionarily conserved gene product, Vip1, possesses autonomous kinase and pyrophosphatase domains capable of synthesis and destruction of D-1 PP-IPs. Our studies provide atomic-resolution structures of the PP-IP products and unequivocally define that the Vip1 gene product is a highly selective 1-kinase and 1-pyrophosphatase enzyme whose activities arise through distinct active sites.

Modulation of sulfur assimilation metabolic toxicity overcomes anemia and hemochromatosis in mice.

Sulfur assimilation is an essential metabolic pathway that regulates sulfation, amino acid metabolism, nucleotide hydrolysis, and organismal homeostasis. We recently reported that mice lacking bisphosphate 3'-nucleotidase (BPNT1), a key regulator of sulfur assimilation, develop iron-deficiency anemia (IDA) and anasarca. Here we demonstrate two approaches that successfully reduce metabolic toxicity caused by loss of BPNT1: 1) dietary methionine restriction and 2) overproduction of a key transcriptional regulator hypoxia inducible factor 2α (Hif-2a).

Metabolic Labeling of Inositol Phosphates and Phosphatidylinositols in Yeast and Mammalian Cells.

Inositol phosphate (IP) and phosphatidylinositol (PI) signaling are critical signal transduction pathways responsible for generating numerous receptor-mediated cellular responses. Biochemical and genetic studies have revealed diverse roles of IP and PI signaling in eukaryotic signaling, but detailed characterization of unique IP and PI signaling profiles in response to different agonists and among cell types remains largely unexplored. Here, we outline steady-state inositol metabolic-labeling techniques that can be leveraged to assess the IP and PI signaling state in eukaryotic cells.

A synthetic biological approach to reconstitution of inositide signaling pathways in bacteria.

Inositide lipid (PIP) and soluble (IP) signaling pathways produce essential cellular codes conserved in eukaryotes. In many cases, deconvoluting metabolic and functional aspects of individual pathways are confounded by promiscuity and multiplicity of PIP and IP kinases and phosphatases. We report a molecular genetic approach that reconstitutes eukaryotic inositide lipid and soluble pathways in a prokaryotic cell which inherently lack inositide kinases and phosphatases in their genome.

Direct Activation of Human MLKL by a Select Repertoire of Inositol Phosphate Metabolites.

Necroptosis is an inflammatory form of programmed cell death executed through plasma membrane rupture by the pseudokinase mixed lineage kinase domain-like (MLKL). We previously showed that MLKL activation requires metabolites of the inositol phosphate (IP) pathway. Here we reveal that I(1,3,4,6)P, I(1,3,4,5,6)P, and IP promote membrane permeabilization by MLKL through directly binding the N-terminal executioner domain (NED) and dissociating its auto-inhibitory region.

MLKL Requires the Inositol Phosphate Code to Execute Necroptosis.

Necroptosis is an important form of lytic cell death triggered by injury and infection, but whether mixed lineage kinase domain-like (MLKL) is sufficient to execute this pathway is unknown. In a genetic selection for human cell mutants defective for MLKL-dependent necroptosis, we identified mutations in IPMK and ITPK1, which encode inositol phosphate (IP) kinases that regulate the IP code of soluble molecules. We show that IP kinases are essential for necroptosis triggered by death receptor activation, herpesvirus infection, or a pro-necrotic MLKL mutant.

Modulation of intestinal sulfur assimilation metabolism regulates iron homeostasis.

Sulfur assimilation is an evolutionarily conserved pathway that plays an essential role in cellular and metabolic processes, including sulfation, amino acid biosynthesis, and organismal development. We report that loss of a key enzymatic component of the pathway, bisphosphate 3'-nucleotidase (Bpnt1), in mice, both whole animal and intestine-specific, leads to iron-deficiency anemia. Analysis of mutant enterocytes demonstrates that modulation of their substrate 3'-phosphoadenosine 5'-phosphate (PAP) influences levels of key iron homeostasis factors involved in dietary iron reduction, import and transport, that in part mimic those reported for the loss of hypoxic-induced transcription factor, HIF-2α.

Inositol phosphate multikinase dependent transcriptional control.

Production of lipid-derived inositol phosphates including IP and IP is an evolutionarily conserved process essential for cellular adaptive responses that is dependent on both phospholipase C and the inositol phosphate multikinase Ipk2 (also known as Arg82 and IPMK). Studies of Ipk2, along with Arg82 prior to demonstrating its IP kinase activity, have provided an important link between control of gene expression and IP metabolism as both kinase dependent and independent functions are required for proper transcriptional complex function that enables cellular adaptation in response to extracellular queues such as nutrient availability. Here we define a promoter sequence cis-element, 5'-CCCTAAAAGG-3', that mediates both kinase-dependent and independent functions of Ipk2.

Structural analyses of sterol 14α-demethylase complexed with azole drugs address the molecular basis of azole-mediated inhibition of fungal sterol biosynthesis.

With some advances in modern medicine (such as cancer chemotherapy, broad exposure to antibiotics, and immunosuppression), the incidence of opportunistic fungal pathogens such as has increased. Cases of drug resistance among these pathogens have become more frequent, requiring the development of new drugs and a better understanding of the targeted enzymes. Sterol 14α-demethylase (CYP51) is a cytochrome P450 enzyme required for biosynthesis of sterols in eukaryotic cells and is the major target of clinical drugs for managing fungal pathogens, but some of the CYP51 key features important for rational drug design have remained obscure.

Balloon-Occluded Retrograde Transvenous Obliteration of a Gastric Vascular Malformation: An Innovative Approach to Treatment of a Rare Condition.

Arteriovenous malformations (AVMs) are a high-flow form of a vascular malformation, which can be found anywhere in the body. While historically treated surgically, a multidisciplinary approach utilizing multiple specialties and treatment modalities is now commonly employed. In order to effectively treat an AVM, the nidus must be targeted and eradicated, which can be done via multiple approaches.

Determining the Impact of Ligand and Alkene Substituents on Bonding in Gold(I)-Alkene Complexes Supported by N-Heterocyclic Carbenes: A Computational Study.

The nature of the gold(I)-alkene bond in [(NHC)Au(alkene)](+) complexes (where NHC is the N-heterocyclic carbene 1,3-bis(2,6-dimethylphenyl)imidazole-2-ylidine and its derivatives) has been studied using density functional theory. By utilization of a series of electron-withdrawing and electron-donating substituents ranging from -NO2 to -NH2, an examination of substituent effects has been undertaken with 4-substituted NHC ligands, monosubstituted ethylene derivatives, and 4-substituted styrene derivatives. Natural population, natural bond orbital (NBO), molecular orbital, and bond energy decomposition analysis (EDA) methods have been used to quantify a number of important parameters, including the charge of the coordinated alkenes and the magnitude of alkene→[(NHC)Au](+) and [(NHC)Au](+)→alkene electron donation.

Blast Injuries: From Improvised Explosive Device Blasts to the Boston Marathon Bombing.

Although most trauma centers have experience with the imaging and management of gunshot wounds, in most regions blast wounds such as the ones encountered in terrorist attacks with the use of improvised explosive devices (IEDs) are infrequently encountered outside the battlefield. As global terrorism becomes a greater concern, it is important that radiologists, particularly those working in urban trauma centers, be aware of the mechanisms of injury and the spectrum of primary, secondary, tertiary, and quaternary blast injury patterns. Primary blast injuries are caused by barotrauma from the initial increased pressure of the explosive detonation and the rarefaction of the atmosphere immediately afterward.

Inositol phosphate kinase 2 is required for imaginal disc development in Drosophila.

Inositol phosphate kinase 2 (Ipk2), also known as IP multikinase IPMK, is an evolutionarily conserved protein that initiates production of inositol phosphate intracellular messengers (IPs), which are critical for regulating nuclear and cytoplasmic processes. Here we report that Ipk2 kinase activity is required for the development of the adult fruit fly epidermis. Ipk2 mutants show impaired development of their imaginal discs, the primordial tissues that form the adult epidermis.