Single point mutations in the zinc finger motifs of the human immunodeficiency virus type 1 nucleocapsid alter RNA binding specificities of the gag protein and enhance packaging and infectivity

A specific interaction between the nucleocapsid (NC) domain of the Gag polyprotein and the RNA encapsidation signal (Psi) is required for preferential incorporation of the retroviral genomic RNA into the assembled virion. Using the yeast three-hybrid system, we developed a genetic screen to detect human immunodeficiency virus type 1 (HIV-1) Gag mutants with altered RNA binding specificities. Specifically, we randomly mutated full-length HIV-1 Gag or its NC portion and screened the mutants for an increase in affinity for the Harvey murine sarcoma virus encapsidation signal. These screens identified several NC zinc finger mutants with altered RNA binding specificities. Furthermore, additional zinc finger mutants that also demonstrated this phenotype were made by site-directed mutagenesis. The majority of these mutants were able to produce normal virion-like particles; however, when tested in a single-cycle infection assay, some of the mutants demonstrated higher transduction efficiencies than that of wild-type Gag. In particular, the N17K mutant showed a seven- to ninefold increase in transduction, which correlated with enhanced vector RNA packaging. This mutant also packaged larger amounts of foreign RNA. Our results emphasize the importance of the NC zinc fingers, and not other Gag sequences, in achieving specificity in the genome encapsidation process. In addition, the described mutations may contribute to our understanding of HIV diversity resulting from recombination events between copackaged viral genomes and foreign RNA.     

Short-Term Synaptic Plasticity Orchestrates the Response of Pyramidal Cells and Interneurons to Population Bursts

The synaptic drive from neuronal populations varies considerably over short time scales. Such changes in the pre-synaptic rate trigger many temporal processes absent under steady-state conditions. This paper examines the differential impact of pyramidal cell population bursts on post-synaptic pyramidal cells receiving depressing synapses, and on a class of interneuron that receives facilitating synapses. In experiment a significant shift of the order of one hundred milliseconds is seen between the response of these two cell classes to the same population burst. It is demonstrated here that such a temporal differentiation of the response can be explained by the synaptic and membrane properties without recourse to elaborate cortical wiring schemes. Experimental data is first used to construct models of the two types of dynamic synaptic response. A population-based approach is then followed to examine analytically the temporal synaptic filtering effects of the population burst for the two post-synaptic targets. The peak-to-peak delays seen in experiment can be captured by the model for experimentally realistic parameter ranges. It is further shown that the temporal separation of the response is communicated in the outgoing action potentials of the two post-synaptic cells: pyramidal cells fire at the beginning of the burst and the class of interneuron receiving facilitating synapses fires at the end of the burst. The functional role of such delays in the temporal organisation of activity in the cortical microcircuit is discussed.     

Signet-ring follicular adenoma of the thyroid diagnosed by fine needle aspiration. Report of a case with cytologic description

BACKGROUND: Follicular cells of the thyroid may undergo squamous, oncocytic or clear cell metaplastic changes. Of these, the clear cell change with signet-ring formation is the most unusual, and follicular adenoma of the thyroid with signet-ring formation is extremely rare. We describe for the first time the cytologic features of a clear cell adenoma of the thyroid obtained by fine needle aspiration (FNA). CASE: A 48-year-old woman in a euthyroid state presented with a 2-cm, right-sided thyroid nodule. Smears obtained by FNA of the nodule revealed clusters of large signet-ring cells in a bloody background. The signet-ring cells were round to oval, with large cytoplasmic vacuoles and hyperchromatic, eccentric nuclei. Colloid in the background was very scanty. Histologic examination of the right hemithyroidectomy specimen revealed a signet-ring follicular adenoma. CONCLUSION: Lack of familiarity with signet-ring cell adenoma of the thyroid could lead to an erroneous diagnosis of metastatic signet-ring cell carcinoma.     

Autoantibody reaction to myelin basic protein by plasma parvovirus B19 IgG in MS patients

The etiology of multiple sclerosis (MS) remains unclear. To determine if autoantibodies to myelin basic protein (MBP) are produced during parvovirus B19 infection, a competitive ELISA was performed using plasma from MS patients exhibiting high IgG titers for parvovirus. Our results showed the addition of MBP decreased the binding of IgG to B19 antigen in a dose dependent fashion suggesting a possible link between parvovirus B19 and a subset of patients with clinical MS.     

Deleterious mutations in LRBA are associated with a syndrome of immune deficiency and autoimmunity

Most autosomal genetic causes of childhood-onset hypogammaglobulinemia are currently not well understood. Most affected individuals are simplex cases, but both autosomal-dominant and autosomal-recessive inheritance have been described. We performed genetic linkage analysis in consanguineous families affected by hypogammaglobulinemia. Four consanguineous families with childhood-onset humoral immune deficiency and features of autoimmunity shared genotype evidence for a linkage interval on chromosome 4q. Sequencing of positional candidate genes revealed that in each family, affected individuals had a distinct homozygous mutation in LRBA (lipopolysaccharide responsive beige-like anchor protein). All LRBA mutations segregated with the disease because homozygous individuals showed hypogammaglobulinemia and autoimmunity, whereas heterozygous individuals were healthy. These mutations were absent in healthy controls. Individuals with homozygous LRBA mutations had no LRBA, had disturbed B cell development, defective in vitro B cell activation, plasmablast formation, and immunoglobulin secretion, and had low proliferative responses. We conclude that mutations in LRBA cause an immune deficiency characterized by defects in B cell activation and autophagy and by susceptibility to apoptosis, all of which are associated with a clinical phenotype of hypogammaglobulinemia and autoimmunity.     

Therapeutic effect of mbp immunodominant peptides encapsulated in nanovehicles in the development of experimental autoimmune encephalomyelitis in DA rats

Multiple sclerosis (MS) is a severe autoimmune neurodegenerative disease. It attacks mainly young people. The development of new approaches to MS treatment is a challenge to modern immunology and pharmacology. In the present study, a high therapeutic efficacy of immunodominant peptides of myelin basic protein (MBP) incorporated into unilamellar mannosylated liposomes in the development of experimental autoimmune encephalomyelitis (EAE) is demonstrated in DA rats. MBP is a component of the oligodendrocyte membrane, which forms the axonal sheath. This protein is among the major autoantigens in MS. We have analyzed the binding pattern of anti-MBP autoantibodies from MS patients using a previously designed MBP epitope library. Utilizing the same approach, we have investigated the pool of anti-MBP antibodies from SJL/J and C57BL/6mice and DA rats with EAE. According to the autoantibody binding patterns, the rodent model most closely mimicking MS is EAE in DA rats. We have chosen three immunodominant MBP fragments encapsulated in unilamellar mannosylated liposomes for the treatment of the verified DA rodent model. MBP fragment 46?C62 is the most efficient in mitigating the first EAE attack, whereas MBP 124?C139 and 147?C160 inhibit the development of pathology at the regression stage. Simultaneous administration of these peptides in liposomes significantly reduces the level of antibodies against MBP. The synergistic therapeutic effect of MBP fragments reduces the integral disease score by inhibiting the first EAE attack and mitigating the subsequent relapse. Thus, our findings offer new opportunities for the efficient treatment of multiple sclerosis.     

Fluoxetine induces vasodilatation of cerebral arterioles by co‐modulating NO/muscarinic signalling

Ischaemic stroke patients treated with Selective Serotonin Reuptake Inhibitors (SSRI) show improved motor, cognitive and executive functions, but the underlying mechanism(s) are incompletely understood. Here, we report that cerebral arterioles in the rat brain superfused with therapeutically effective doses of the SSRI fluoxetine showed consistent, dose‐dependent vasodilatation (by 1.2 to 1.6‐fold), suppressible by muscarinic and nitric oxide synthase (NOS) antagonists [atropine, NG‐nitro‐l ‐arginine methyl ester (l ‐NAME)] but resistant to nicotinic and serotoninergic antagonists (mecamylamine, methylsergide). Fluoxetine administered 10–30 min. following experimental vascular photo‐thrombosis increased arterial diameter (1.3–1.6), inducing partial, but lasting reperfusion of the ischaemic brain. In brain endothelial b.End.3 cells, fluoxetine induced rapid muscarinic receptor‐dependent increases in intracellular [Ca²⁺] and promoted albumin‐ and eNOS‐dependent nitric oxide (NO) production and HSP90 interaction. In vitro, fluoxetine suppressed recombinant human acetylcholinesterase (rhAChE) activity only in the presence of albumin. That fluoxetine induces vasodilatation of cerebral arterioles suggests co‐promotion of endothelial muscarinic and nitric oxide signalling, facilitated by albumin‐dependent inhibition of serum AChE.     

Slow oscillations in neural networks with facilitating synapses

The synchronous oscillatory activity characterizing many neurons in a network is often considered to be a mechanism for representing, binding, conveying, and organizing information. A number of models have been proposed to explain high-frequency oscillations, but the mechanisms that underlie slow oscillations are still unclear. Here, we show by means of analytical solutions and simulations that facilitating excitatory (E _f) synapses onto interneurons in a neural network play a fundamental role, not only in shaping the frequency of slow oscillations, but also in determining the form of the up and down states observed in electrophysiological measurements. Short time constants and strong E _f synapse-connectivity were found to induce rapid alternations between up and down states, whereas long time constants and weak E _f synapse connectivity prolonged the time between up states and increased the up state duration. These results suggest a novel role for facilitating excitatory synapses onto interneurons in controlling the form and frequency of slow oscillations in neuronal circuits.