Members of the opioid receptor family of G-protein-coupled receptors (GPCRs) are found throughout the peripheral and central nervous system, where they have key roles in nociception and analgesia. Unlike the 'classical' opioid receptors, δ, κ and μ (δ-OR, κ-OR and μ-OR), which were delineated by pharmacological criteria in the 1970s and 1980s, the nociceptin/orphanin FQ (N/OFQ) peptide receptor (NOP, also known as ORL-1) was discovered relatively recently by molecular cloning and characterization of an orphan GPCR. Although it shares high sequence similarity with classical opioid GPCR subtypes (∼60%), NOP has a markedly distinct pharmacology, featuring activation by the endogenous peptide N/OFQ, and unique selectivity for exogenous ligands. Here we report the crystal structure of human NOP, solved in complex with the peptide mimetic antagonist compound-24 (C-24) (ref. 4), revealing atomic details of ligand-receptor recognition and selectivity. Compound-24 mimics the first four amino-terminal residues of the NOP-selective peptide antagonist UFP-101, a close derivative of N/OFQ, and provides important clues to the binding of these peptides. The X-ray structure also shows substantial conformational differences in the pocket regions between NOP and the classical opioid receptors κ (ref. 5) and μ (ref. 6), and these are probably due to a small number of residues that vary between these receptors. The NOP-compound-24 structure explains the divergent selectivity profile of NOP and provides a new structural template for the design of NOP ligands.

Naturally occurring and synthetic short arginine containing protein transduction domains (PTDs), including HIV1 TAT, poly-Arg and Antp, have been used to deliver a wide variety of macromolecular, biologically active therapeutic cargo into cells, including peptides, proteins, antisense oligonucleotides and liposomes, in vitro and to treat pre-clinical models of cancer and stroke. PTDs enter cells in a rapid, receptor-independent fashion. Recently, large TAT-fusion proteins (in excess of 30,000 Da) were shown to transduce into cells by fluid-phase macropinocytosis, a specialized form of endocytosis that is independent of caveolae, clathrin and dynamin. However, it remains controversial as to whether or not PTD peptides (1000-5000 Da) enter cells via macropinocytosis and/or through an unknown alternative mechanism. Due to strong ionic interactions with the cell surface, previous measurements of PTD peptide internalization were inaccurate. Cationic PTD peptides containing variable numbers of arginine residues and conditions entered cells exclusively through macropinocytosis. In addition, no PTD peptide was found to enter cells at 4 degrees C, a long held assumption of transduction. Taken together, these observations provide a solid scientific basis for the development of novel biologically active transducible anticancer PTD peptide therapeutics.

Apoptosis is held in check by prosurvival proteins of the Bcl-2 family. The distantly related BH3-only proteins bind to and antagonize them, thereby promoting apoptosis. Whereas binding of the BH3-only protein Noxa to prosurvival Mcl-1 induces Mcl-1 degradation by the proteasome, binding of another BH3-only ligand, Bim, elevates Mcl-1 protein levels. We compared the three-dimensional structures of the complexes formed between BH3 peptides of both Bim and Noxa, and we show that a discrete C-terminal sequence of the Noxa BH3 is necessary to instigate Mcl-1 degradation.

We have solved the high-resolution X-ray structure of 14-3-3 bound to two different phosphoserine peptides, representing alternative substrate-binding motifs. These structures reveal an evolutionarily conserved network of peptide-protein interactions within all 14-3-3 isotypes, explain both binding motifs, and identify a novel intrachain phosphorylation-mediated loop structure in one of the peptides. A 14-3-3 mutation disrupting Raf signaling alters the ligand-binding cleft, selecting a different phosphopeptide-binding motif and different substrates than the wild-type protein. Many 14-3-3: peptide contacts involve a C-terminal amphipathic alpha helix containing a putative nuclear export signal, implicating this segment in both ligand and Crm1 binding. Structural homology between the 14-3-3 NES structure and those within I kappa B alpha and p53 reveals a conserved topology recognized by the Crm1 nuclear export machinery.

Parathyroid hormone (PTH), a major regulator of mineral ion metabolism, and PTH-related peptide (PTHrP), which causes hypercalcemia in some cancer patients, stimulate multiple signals (cAMP, inositol phosphates, and calcium) probably by activating common receptors in bone and kidney. Using expression cloning, we have isolated a cDNA clone encoding rat bone PTH/PTHrP receptor from rat osteosarcoma (ROS 17/2.8) cells. The rat bone PTH/PTHrP receptor is 78% identical to the opossum kidney receptor; this identity indicates striking conservation of this receptor across distant mammalian species. Additionally, the rat bone PTH/PTHrP receptor has significant homology to the secretin and calcitonin receptors but not to any other G protein-linked receptor. When expressed in COS cells, a single cDNA clone, expressing either rat bone or opossum kidney PTH/PTHrP receptor, mediates PTH and PTHrP stimulation of both adenylate cyclase and phospholipase C. These properties could explain the diversity of PTH action without the need to postulate other receptor subtypes.

Complexes of five peptides (from HIV-1, influenza A virus, HTLV-1, and hepatitis B virus proteins) bound to the human class I MHC molecule HLA-A2 have been studied by X-ray crystallography. While the peptide termini and their second and C-terminal anchor side chains are bound similarly in all five cases, the main chain and side chain conformations of each peptide are strikingly different in the center of the binding site, and these differences are accessible to direct TCR recognition. Each of the central peptide residues is seen to point up for some bound peptides, but down or sideways for others. Thus, although fixed at its ends, the structure of an MHC-bound peptide appears to be a highly complex function of its entire sequence, potentially sensitive to even small sequence differences. In contrast, MHC structural variation is relatively limited. These results offer a structural framework for understanding the role of nonanchor peptide side chains in both peptide-MHC binding affinity and TCR recognition.

Glucagon-like peptide-1 (GLP-1) released from the gut functions as an incretin that stimulates insulin secretion. GLP-1 is also a brain neuropeptide that controls feeding and drinking behavior and gastric emptying and elicits neuroendocrine responses including development of conditioned taste aversion. Although GLP-1 receptor (GLP-1R) agonists are under development for the treatment of diabetes, GLP-1 administration may increase blood pressure and heart rate in vivo. We report here that centrally and peripherally administered GLP-1R agonists dose-dependently increased blood pressure and heart rate. GLP-1R activation induced c-fos expression in the adrenal medulla and neurons in autonomic control sites in the rat brain, including medullary catecholamine neurons providing input to sympathetic preganglionic neurons. Furthermore, GLP-1R agonists rapidly activated tyrosine hydroxylase transcription in brainstem catecholamine neurons. These findings suggest that the central GLP-1 system represents a regulator of sympathetic outflow leading to downstream activation of cardiovascular responses in vivo.

The seven-residue peptide N-acetyl-Lys-Leu-Val-Phe-Phe-Ala-Glu-NH(2), called A beta(16-22) and representing residues 16-22 of the full-length beta-amyloid peptide associated with Alzheimer's disease, is shown by electron microscopy to form highly ordered fibrils upon incubation of aqueous solutions. X-ray powder diffraction and optical birefringence measurements confirm that these are amyloid fibrils. The peptide conformation and supramolecular organization in A beta(16-22) fibrils are investigated by solid state (13)C NMR measurements. Two-dimensional magic-angle spinning (2D MAS) exchange and constant-time double-quantum-filtered dipolar recoupling (CTDQFD) measurements indicate a beta-strand conformation of the peptide backbone at the central phenylalanine. One-dimensional and two-dimensional spectra of selectively and uniformly labeled samples exhibit (13)C NMR line widths of <2 ppm, demonstrating that the peptide, including amino acid side chains, has a well-ordered conformation in the fibrils. Two-dimensional (13)C-(13)C chemical shift correlation spectroscopy permits a nearly complete assignment of backbone and side chain (13)C NMR signals and indicates that the beta-strand conformation extends across the entire hydrophobic segment from Leu17 through Ala21. (13)C multiple-quantum (MQ) NMR and (13)C/(15)N rotational echo double-resonance (REDOR) measurements indicate an antiparallel organization of beta-sheets in the A beta(16-22) fibrils. These results suggest that the degree of structural order at the molecular level in amyloid fibrils can approach that in peptide or protein crystals, suggest how the supramolecular organization of beta-sheets in amyloid fibrils can be dependent on the peptide sequence, and illustrate the utility of solid state NMR measurements as probes of the molecular structure of amyloid fibrils. A beta(16-22) is among the shortest fibril-forming fragments of full-length beta-amyloid reported to date, and hence serves as a useful model system for physical studies of amyloid fibril formation.

We have investigated the cell surface recognition mechanisms used by human monocyte-derived macrophages (M phi) in phagocytosis of intact aging human neutrophils (PMNs) undergoing apoptosis. This study shows that the adhesive protein thrombospondin (TSP) was present in the interaction, both associated with the M phi surface and in solution at a mean concentration of 0.59 micrograms/ml. The interaction was inhibited by treatment of M phi (but not aged PMN) with cycloheximide, but could be "rescued" by replenishment with exogenous TSP. Under control conditions, M phi recognition of aged PMNs was specifically potentiated by purified platelet TSP at 5 micrograms/ml, present either in the interaction or if preincubated with either cell type, suggesting that TSP might act as a "molecular bridge" between the two cell types. In support, both aged PMN and M phi were found to adhere to TSP, and phagocytosis of aged PMN was specifically inhibited by (a) excess soluble TSP; (b) antibodies to TSP that also inhibit TSP-mediated adhesion to aged PMN; and (c) down-regulation of M phi receptors for TSP by plating M phi on TSP-coated surfaces. Furthermore, inhibition with mAbs/Arg-Gly-Asp-Ser peptide of the candidate M phi receptors for TSP, CD36, and alpha v beta 3 exerted synergistic effects on both M phi recognition of aged PMN and M phi adhesion to TSP, indicating that "two point" adhesion of TSP to these M phi structures is involved in phagocytosis of aged PMN. Our findings indicate newly defined roles for TSP and CD36 in phagocytic clearance of senescent neutrophils, which may limit inflammatory tissue injury and promote resolution.

Phorbol esters (e.g., TPA) activate protein kinase C (PKC), increase connexin43 (Cx43) phosphorylation, and decrease cell-cell communication via gap junctions in many cell types. We asked whether PKC directly phosphorylates and regulates Cx43. Rat epithelial T51B cells metabolically labeled with (32)P(i) yielded two-dimensional phosphotryptic maps of Cx43 with several phosphopeptides that increased in intensity upon TPA treatment. One of these peptides comigrated with the major phosphopeptide observed after PKC phosphorylation of immunoaffinity-purified Cx43. Purification of this comigrating peptide and subsequent sequencing indicated that the phosphorylated serine was residue 368. To pursue the functional importance of phosphorylation at this site, fibroblasts from Cx43(-/-) mice were transfected with either wild-type (Cx43wt) or mutant Cx43 (Cx43-S368A). Intercellular dye transfer studies revealed different responses to TPA and were followed by single channel analyses. TPA stimulation of T51B cells or Cx43wt-transfected fibroblasts caused a large increase in the relative frequency of approximately 50-pS channel events and a concomitant loss of approximately 100-pS channel events. This change to approximately 50-pS events was absent when cells transfected with Cx43-S368A were treated with TPA. These data strongly suggest that PKC directly phosphorylates Cx43 on S368 in vivo, which results in a change in single channel behavior that contributes to a decrease in intercellular communication.

Conformational aspects of N-glycosylation have been investigated with a series of proline-containing peptides as molecular probes. The results demonstrate that, depending on the position of the imino acid in the peptide chain, dramatic alterations of glycosylation rates are produced, pointing to a critical contribution of the amino acids framing the 'marker sequence' triplet Asn-Xaa-Thr(Ser) on the formation of a potential sugar-attachment site. No glycosyl transfer at all was detectable to those peptides containing a proline residue either in position Xaa or in the next position beyond the threonine of the Asn-sequon on the C-terminal side, whereas the hexapeptide Pro-Asn-Gly-Thr-Ala-Val was glycosylated at a high rate. (Emboldened residues denote the 'marker sequence' that is identical in all the peptides; italicized residues distinguish the positions of proline in the various peptides.) Studies with space-filling models reveal that the lack of glycosyl-acceptor capabilities of Ala(Pro)-Asn-Gly-Thr-Pro-Val might be directly related to their inability to adopt and/or stabilize a turn or loop conformation which permits the catalytically essential interaction between the hydroxy amino acid and the asparagine residue within the 'marker sequence' [Bause & Legler (1981) Biochem. J. 195, 639-644]. This conclusion is supported by circular-dichroism spectroscopic data, which suggest structure-forming potentials in this type of non-acceptor peptides dominating over those that favour the induction of an appropriate sugar-attachment site in the acceptor peptides. The lack of acceptor properties of Tyr-Asn-Pro-Thr-Ser-Val indicates that even small modifications in the 'recognition' pattern are not tolerated by the N-glycosyltransferases.

Mammalian mitochondrial complex I is a multisubunit membrane-bound assembly with a molecular mass approaching 1 MDa. By comprehensive analyses of the bovine complex and its constituent subcomplexes, 45 different subunits have been characterized previously. The presence of a 46th subunit was suspected from the consistent detection of a molecular mass of 10,566 by electrospray ionization mass spectrometry of subunits fractionated by reverse-phase high pressure liquid chromatography. The component was found associated with both the intact complex and subcomplex Ibeta, which represents most of the membrane arm of the complex, and it could not be resolved chromatographically from subunit SGDH (the subunit of bovine complex I with the N-terminal sequence Ser-Gly-Asp-His). It has now been characterized by tandem mass spectrometry of intact protein ions and shown to be a C-terminal fragment of subunit SGDH arising from a specific peptide bond cleavage between Ile-55 and Pro-56 during the electrospray ionization process. Thus, the subunit composition of bovine complex I has been established. It is a complex of 45 different proteins plus non-covalently bound FMN and eight iron-sulfur clusters.

OBJECTIVE

Nonalcoholic steatohepatitis (NASH) is a leading cause of chronic liver disease for which there is limited therapy available. Insulin sensitizing, anti-inflammatory, and antifibrotic properties of thiazolidinediones support their use in treating NASH. We have evaluated pioglitazone in the treatment of nondiabetic patients with NASH.

METHODS

We randomized 74 nondiabetic patients (45 men; median age, 54 y) with histologically proven NASH to 12 months of standard diet, exercise, and either placebo or pioglitazone (30 mg/day). Sixty-one patients (30 placebo, 31 pioglitazone) had liver biopsies both at the beginning and the end of the study.

RESULTS

Compared with placebo, pioglitazone therapy was associated with an increase in weight (mean change, -0.55 vs +2.77 kg; P = .04) and a reduction in glucose (+0.4 vs -0.1 mmol/L; P = .02), HbA1c (+0.16% vs -0.18%; P = .006), insulin C peptide level (+42 vs -78 pmol/L; P = .02), alanine aminotransferase level (-10.9 vs -36.2 u/L; P = .009), gamma-glutamyltransferase level (-9.4 vs -41.2 u/L; P = .002), and ferritin (-11.3 vs -90.5 microg/L; P = .01). Histologic features including hepatocellular injury (P = .005), Mallory-Denk bodies (P = .004), and fibrosis (P = .05) were reduced in patients treated with pioglitazone compared with those in the placebo group.

CONCLUSIONS

Pioglitazone therapy over a 12-month period in nondiabetic subjects with NASH resulted in improvements in metabolic and histologic parameters, most notably liver injury and fibrosis. Larger extended trials are justified to assess the long-term efficacy of pioglitazone in this patient group.

Substrates enter the cylindrical 20S proteasome through a gated channel that is regulated by the ATPases in the 19S regulatory particle in eukaryotes or the homologous PAN ATPase complex in archaea. These ATPases contain a conserved C-terminal hydrophobic-tyrosine-X (HbYX) motif that triggers gate opening upon ATP binding. Using cryo-electron microscopy, we identified the sites in the archaeal 20S where PAN's C-terminal residues bind and determined the structures of the gate in its closed and open forms. Peptides containing the HbYX motif bind to 20S in the pockets between neighboring alpha subunits where they interact with conserved residues required for gate opening. This interaction induces a rotation in the alpha subunits and displacement of a reverse-turn loop that stabilizes the open-gate conformation. This mechanism differs from that of PA26/28, which lacks the HbYX motif and does not cause alpha subunit rotation. These findings demonstrated how the ATPases' C termini function to facilitate substrate entry.

BACKGROUND

Although insulin secretion is severely decreased in most patients with type 1 diabetes, levels of residual insulin secretion often vary early in the disease. The significance of residual insulin secretion with regard to metabolic control and to long-term complications and ways to preserve such secretion are not well understood.

OBJECTIVE

To compare the effects of intensive and conventional therapy on residual insulin secretion in Diabetes Control and Complications Trial (DCCT) participants.

METHODS

Multicenter, randomized, controlled clinical trial.

METHODS

29 DCCT clinical centers.

METHODS

855 of the 1441 DCCT participants had had type 1 diabetes for 1 to 5 years at baseline. Of these 855 patients, 303 were C-peptide responders (C-peptide level, 0.20 to 0.50 pmol/mL after ingestion of a standardized, mixed meal); 138 of these patients were randomly assigned to intensive therapy, and 165 were assigned to conventional therapy. Five hundred fifty-two patients were nonresponders (stimulated C-peptide level < 0.2 pmol/mL); 274 of these patients were assigned to intensive therapy, and 278 were assigned to conventional therapy.

METHODS

1) Intensive therapy with 3 or more insulin injections daily or continuous subcutaneous infusion of insulin, guided by 4 or more glucose tests per day or 2) conventional therapy with 1 or 2 insulin injections daily.

METHODS

Stimulated C-peptide level was measured annually in responders. Development of retinopathy and microalbuminuria was assessed annually, hemoglobin A1c levels were measured quarterly, and episodes of hypoglycemia were ascertained quarterly.

RESULTS

Responders receiving intensive therapy maintained a higher stimulated C-peptide level and a lower likelihood of becoming nonresponders than did responders receiving conventional therapy (risk reduction, 57% [95% CI, 39% to 71%]; P < 0.001). As in the entire DCCT cohort, intensively treated responders had a reduced risk for retinopathy progression and development of microalbuminuria and a higher risk for severe hypoglycemia compared with conventionally treated responders. Among intensively treated patients, responders had a lower hemoglobin A1c value (P < 0.01), a 50% (95% CI, 12% to 72%) reduced risk for retinopathy progression, and a lower risk for severe hypoglycemia (risk reduction, 65% [CI, 53% to 74%]; P < 0.001) compared with nonresponders.

CONCLUSIONS

Intensive therapy for type 1 diabetes helps sustain endogenous insulin secretion, which, in turn, is associated with better metabolic control and lower risk for hypoglycemia and chronic complications. These observations underscore the importance of initiating intensive diabetic management as early as safely possible after type 1 diabetes is diagnosed.

Temporally precise inhibition of distinct cell types in the intact nervous system has been enabled by the microbial halorhodopsin NpHR, a fast light-activated electrogenic Cl(-) pump. While neurons can be optically hyperpolarized and inhibited from firing action potentials at moderate NpHR expression levels, we have encountered challenges with pushing expression to extremely high levels, including apparent intracellular accumulations. We therefore sought to molecularly engineer NpHR to achieve strong expression without these cellular side effects. We found that high expression correlated with endoplasmic reticulum (ER) accumulation, and that under these conditions NpHR colocalized with ER proteins containing the KDEL ER retention sequence. We screened a number of different putative modulators of membrane trafficking and identified a combination of two motifs, an N-terminal signal peptide and a C-terminal ER export sequence, that markedly promoted membrane localization and ER export defined by confocal microscopy and whole-cell patch clamp. The modified NpHR displayed increased peak photocurrent in the absence of aggregations or toxicity, and potent optical inhibition was observed not only in vitro but also in vivo with thalamic single-unit recording. The new enhanced NpHR (eNpHR) allows safe, high-level expression in mammalian neurons, without toxicity and with augmented inhibitory function, in vitro and in vivo.

The antioxidant response element (ARE) and transcription factor Nrf2 regulate basal expression and antioxidant induction of NAD(P)H:quinone oxidoreductase-1 (NQO1) and other detoxifying genes. Under normal conditions, Nrf2 is targeted for proteasomal degradation by INrf2. Oxidative stress causes release of Nrf2 from INrf2. Nrf2 translocates to the nucleus, binds to the ARE, and activates gene expression. In this study, we demonstrate that protein kinase C (PKC) plays a significant role in the regulation of ARE-mediated NQO1 gene expression and induction in response to t-butylhydroquinone. Treatment of HepG2 cells with the PKC inhibitors staurosporine and calphostin C repressed ARE-mediated induction of a luciferase reporter as well as that of the endogenous NQO1 gene. Similar experiments with inhibitors of MEK/ERK, p38, phosphatidylinositol 3-kinase, and tyrosine kinases failed to repress ARE-mediated gene expression. The PKC inhibitor staurosporine blocked the nuclear translocation of Nrf2, suggesting that Nrf2 might be the target for PKC regulation. A Prosite search revealed the presence of seven putative PKC sites in mouse Nrf2. The PKC site at Ser40 is conserved among species and lies in the Neh2 domain, which interacts with INrf2. We demonstrate that phosphorylation of Ser40 is necessary for Nrf2 release from INrf2, but is not required for Nrf2 stabilization/accumulation in the nucleus and transcriptional activation of ARE-mediated NQO1 gene expression. A peptide that competes with endogenous Nrf2 for INrf2 binding was able to induce ARE activity more effectively than t-butylhydroquinone, and Nrf2 that accumulated in the nucleus as a result was not phosphorylated.

This study aims to determine the presence of the components of the metabolic syndrome in primary nonalcoholic steatohepatitis (NASH) and to assess the role of liver disease in the genesis of peripheral hyperinsulinemia. Nineteen patients (18 men and 1 woman; mean age, +/- SD, 38 +/- 10 years; body mass index [BMI], 26 +/- 2 kg/m(2)) with histologic evidence of NASH were enrolled; 19 age- and sex-matched normal subjects were investigated as controls. Plasma glucose, insulin, and C-peptide levels were measured during an oral glucose tolerance test, and a frequently sampled intravenous glucose tolerance test (FSIGT), analyzed by minimal modeling technique, was performed. Compared with controls, the NASH group had lower insulin sensitivity (3.84 +/- 2.44 vs. 7.48 +/- 3.01 10(-4) x min(-1)/microU/mL; P =.0003) and higher total insulin secretion (21 +/- 13 vs. 10 +/- 3 nmol/L in 240 minutes; P =.001). Hepatic insulin extraction was similar in both groups (69.8% +/- 16.1% vs. 70.2% +/- 18.3%; P =.854). According to the results of the oral glucose tolerance test, no patient was classified as diabetic, 5 were classified as glucose intolerant, and 1 was classified as having impaired fasting glycemia. Nine patients (47%) had at least the 2 minimum criteria required to define the metabolic syndrome according to the European Group for the Study of Insulin Resistance (EGIR). In conclusion, hyperinsulinemia and insulin resistance occur frequently in patients with NASH; these conditions do not stem from a reduced hepatic insulin extraction but from an enhanced pancreatic insulin secretion compensatory to reduced insulin sensitivity. The derangement of insulin regulation, often associated with the metabolic syndrome, may play a causal role in the pathogenesis of NASH.

The genetic structure of Mason-Pfizer monkey virus (MPMV), a D-type retrovirus, has been determined. In addition to the viral gag, pol, and env genes is an ORF overlapping both gag and pol and that encodes the viral protease. Surprisingly, the MPMV env protein is highly homologous to that of the avian C-type virus, reticuloendotheliosis associated virus REV-A. The env sequence encodes an immunosuppressive peptide, which suggests that MPMV, like REV-A, may transiently induce a T-suppressor cell population. The different phylogenies of the MPMV pol and env genes indicate a recombinatorial origin for the D-type viruses. Sequence comparisons show that SRV-1, an MPMV-like virus etiologically linked to simian AIDS (SAIDS), is in fact a variant of MPMV. While MPMV-like viruses cannot be used as direct models for the AIDS/SAIDS associated with lentiviruses, they provide an important system for studying the molecular basis of immunosuppressive diseases in primates.

Background: Cardiac injury and myocarditis have been described in adults with COVID-19. SARS-CoV-2 infection in children is typically minimally symptomatic. We report a series of febrile pediatric patients with acute heart failure potentially associated with SARS-CoV-2 infection and the multisystem inflammatory syndrome in children (MIS-C) as defined by the US Centers for Disease Control. Methods: Over a two-month period contemporary with the SARS-CoV-2 pandemic in France and Switzerland, we retrospectively collected clinical, biological, therapeutic, and early outcomes data in children who were admitted to pediatric intensive care units in 14 centers for cardiogenic shock, left ventricular dysfunction and severe inflammatory state. Results: Thirty-five children were identified and included in the study. Median age at admission was 10 years (range 2-16 years). Co-morbidities were present in 28% including asthma and overweight. Gastrointestinal symptoms were prominent. Left ventricular ejection fraction was <30% in one third; 80% required inotropic support with 28% treated with ECMO. Inflammation markers were suggestive of cytokine storm (interleukin 6 median 135 pg/mL) and macrophage activation (D-dimer median 5284 ng/mL). Mean brain natriuretic peptide was elevated (5743 pg/mL). Thirty-one/35 (88%) patients tested positive for SARS-CoV-2 infection by PCR of nasopharyngeal swab or serology. All patients received intravenous immune globulin, with adjunctive steroid therapy used in one third. Left ventricular function was restored in the 25/35 of those discharged from the intensive care unit. No patient died, and all patients treated with ECMO were successfully weaned. Conclusion: Children may experience an acute cardiac decompensation due to severe inflammatory state following SARS-CoV-2 infection (multisystem inflammatory syndrome in children - MIS-C). Treatment with immune globulin appears to be associated with recovery of left ventricular systolic function.

Keywords: COVID-19; SARS-CoV-2; myocardial stunning.

The genetic basis for the biosynthesis of large polypeptide antibiotics such as nisin has not been explained so far. We show here that the structural gene epiA encoding the antibiotic epidermin from Staphylococcus epidermidis is located on a 54-kilobase plasmid and codes for a 52-amino-acid prepeptide, which is processed to the tetracyclic 21-peptide amide antibiotic. The mature sequence of epidermin corresponds to the C-terminal 22-peptide segment of pre-epidermin and contains the precursor amino acids Ser, Thr and Cys, from which the unusual amino-acid constituents are derived. The more lipophilic epidermin is cleaved at a hydrophilic turn between Arg-1 and Ile+1 from the N-terminal segment-30 to -1, which probably assumes a partially amphiphilic alpha-helix conformation. We propose that the N-terminus (-30 to -1) plays a cooperative role during modification reactions and prevents toxicity of the mature epidermin to the producing strain before the antibiotic is cleaved off and secreted.

RNA interference is a naturally occurring endogenous regulatory process where short double-stranded RNA causes sequence-specific posttranscriptional gene silencing. Small interference RNA (siRNA) represents a promising therapeutic strategy. Clinical evaluations of siRNA therapeutics in locoregional treatment settings began in 2004. Systemic siRNA therapy is hampered by the barriers for siRNA to reach their intended targets in the cytoplasm and to exert their gene silencing activity. The three goals of this review were to provide an overview of (a) the barriers to siRNA delivery, from the perspectives of physicochemical properties of siRNA, pharmacokinetics and biodistribution, and intracellular trafficking; (b) the non-viral siRNA carriers including cell-penetrating peptides, polymers, dendrimers, siRNA bioconjugates, and lipid-based siRNA carriers; and (c) the current status of the clinical trials of siRNA therapeutics.

The docking of transport vesicles with their target membrane is thought to be mediated by p115. We show here that GM130, a cis-Golgi matrix protein, interacts specifically with p115 and so could provide a membrane docking site. Deletion analysis showed that the N-terminus binds to p115, whereas the C-terminus binds to Golgi membranes. Mitotic phosphorylation of GM130 or a peptide derived from the N-terminus prevented binding to p115. The peptide also inhibited the NSF- but not the p97-dependent reassembly of Golgi cisternae from mitotic fragments, unless it was mitotically phosphorylated. Together, these data provide a molecular explanation for the COPI-mediated fragmentation of the Golgi apparatus at the onset of mitosis.

OBJECTIVE

Chronic hepatitis C virus infection is associated with an increased prevalence of type 2 diabetes. We hypothesized that virus-induced insulin resistance may be a mechanism for fibrogenesis in chronic hepatitis C virus infection.

METHODS

In 260 hepatitis C virus-infected subjects, we examined the relationship between histological findings and anthropometric and biochemical data, including insulin resistance determined by the homeostasis model assessment (HOMA-IR). We also compared fasting serum insulin, C peptide, and HOMA-IR levels between the subset of 121 hepatitis C virus patients with stage 0 or 1 hepatic fibrosis and 137 healthy volunteers matched by sex, body mass index, and waist-hip ratio.

RESULTS

Hepatitis C virus-infected subjects with stage 0 or 1 hepatic fibrosis had higher levels of insulin, C peptide, and HOMA-IR (all P < or = 0.01) compared with matched healthy controls. In the 250 hepatitis C virus patients (fibrosis stage 0 to 4), viral genotype and portal, but not lobular, inflammation were univariate predictors of HOMA-IR. By multiple linear regression analysis, independent predictors of HOMA-IR included body mass index (P < 0.001), previous failed antiviral treatment (P < 0.001), portal inflammatory grade (P < 0.001), and genotype 3 status (P = 0.01). Genotype 3 had significantly lower HOMA-IR than other genotypes (which were comparable when adjusted for effects of the remaining independent predictors). HOMA-IR was an independent predictor for the degree of fibrosis (P < 0.001) and the rate of fibrosis progression (P = 0.03).

CONCLUSIONS

Hepatitis C virus may induce insulin resistance irrespective of the severity of liver disease, and this effect seems to be genotype specific. Further, our findings support the hypothesis that insulin resistance may contribute to fibrotic progression in chronic hepatitis C virus infection.