Expression and biochemical characterization of iron regulatory proteins 1 and 2 in Saccharomyces cerevisiae

Iron-regulatory proteins (IRPs) 1 and 2 are cytosolic RNA-binding proteins that bind to specific stem-loop structures, termed iron-responsive elements (IREs) that are located in the untranslated regions of specific mRNAs encoding proteins involved in iron metabolism. The binding of IRPs to IREs regulates either translation or stabilization of mRNA. Although IRP1 and IRP2 are similar proteins in that they are ubiquitously expressed and are negatively regulated by iron, they are regulated by iron by different mechanisms. IRP1, the well-characterized IRP in cells, is a dual-function protein exhibiting either aconitase activity when cellular iron is abundant or RNA-binding activity when cellular iron is scarce. In contrast, IRP2 lacks detectable aconitase activity and functions exclusively as an RNA-binding protein. To study and compare the biochemical characteristics of IRP1 and IRP2, we expressed wild-type and mutant rat IRP1 and IRP2 in the yeast Saccharomyces cerevisiae. IRP1 and IRP2 expressed in yeast bind the IRE RNA with high affinity, resulting in the inhibition of translation of an IRE-reporter mRNA. Mutant IRP2s lacking a 73 amino acid domain unique to IRP2 and a mutant IRP1 containing an insertion of this domain bound RNA, but lacked detectable aconitase activity, suggesting that the presence of this domain prevents aconitase activity. Like IRP1, the RNA-binding activity of IRP2 was sensitive to inactivation by N-ethylmaleimide (NEM) or 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB), indicating IRP2 contains a cysteine(s) that is (are) necessary for RNA binding. However, unlike IRP1, where reconstitution of the 4Fe-4S cluster resulted in a loss in RNA-binding activity, the RNA-binding activity of IRP2 was unaffected using the same iron treatment. These data suggested that IRP2 does not contain a 4Fe-4S cluster similar to the cluster in IRP1, indicating that they sense iron by different mechanisms.     

Ionic conductivity, transference numbers, composition and mobility of ions in cross-linked lysozyme crystals

Basic fibroblast growth factor (bFGF) gene expression as well as its immunoreactivity were studied after partial unilateral hemitransection of the rat brain during a time course of 24 h, 72 h, 7 and 14 days. The mechanical injury resulted in a global increase of bFGF gene expression at the 24-h time interval. This global increase was seen at the ipsilateral site at the level of the lesion as well as rostral to the lesion in the ipsilateral hemisphere. The upregulation in bFGF gene expression was in most of the areas investigated due to an upregulation in glial cells as seen by means of nonradioactive in situ hybridization compared with immunocytochemistry for glial fibrillary acidic protein (GFAP). Basic FGF immunoreactivity (IR) was increased around the lesion in glial cell nuclei 7 days after the injury. This increase was also detected in GFAP positive glial cells surrounding small vessels in the lesioned area. Moreover, in the present paper we demonstrate increased tenascin immunoreactivity in the lesioned area 7 days after injury. The tenascin IR was increased at the edges of the lesion as well as in vessel like structures. The tenascin IR was partially codistributed with GFAP IR in the lesioned area. The lesion was also characterized by an increase in vimentin IR as well as in laminin IR. It is suggested that the observed changes in the expression of bFGF, matrix proteins (laminin, tenascin) and intermediate filaments (vimentin) are involved in (a) tissue repair, (b) protection of neuronal cells from excitotoxic influences and (c) formation of new vessels in the lesioned area.     

Compartment syndromes of the hand

We retrospectively reviewed the records of nineteen patients who had been managed with fasciotomy because of compartment syndrome of the hand. The patients were five months to sixty-seven years old and included ten adults and nine children. Seventeen patients were followed for an average of twenty-one months (range, one to fifty-eight months), one patient was lost to follow-up after discharge, and one patient died four days postoperatively. All of the patients had a tense, swollen hand and elevated pressure in at least one interosseous compartment. Eight patients also had a compartment syndrome of the forearm. The compartment syndromes developed after intravenous injections (eleven patients); after a gunshot wound, a crush injury, or a complication related to the use of an arterial line (two patients each); and after a complication related to an arthrodesis of the wrist or a crush injury due to prolonged pressure on the upper extremity secondary to a drug overdose (one patient each). Fifteen patients had an obtunded sensorium-either because of a serious illness or injury or secondary to prolonged anesthesia-when the compartment syndrome was recognized. In thirteen of these patients, including eight children and five adults, the compartment syndrome developed because of a complication related to the intravenous or intra-arterial administration of drugs. Carpal tunnel release and decompression of the involved compartments led to a satisfactory result for thirteen of the seventeen patients who were followed. The remaining four patients (including two children who had an amputation, one child who had impaired function of the hand secondary to brain damage, and one adult who had extensive involvement of the forearm and complete loss of function of the hand) had a poor result. All four of these patients had been obtunded when the compartment syndrome developed. The treating physician should maintain a high index of suspicion for a compartment syndrome of the hand when managing seriously ill, obtunded patients-particularly children-who are receiving multiple intravenous or intra-arterial injections.     

The efficacy of fluticasone propionate aqueous nasal spray for allergic rhinitis and its relationship to topical effects

Fluticasone propionate aqueous nasal spray is an intranasal corticosteroid for the treatment of patients with allergic rhinitis. This double-masked, double-dummy, parallel-group study was conducted to confirm that the efficacy of fluticasone propionate nasal spray is attributable to topical rather than systemic effects. A total of 304 patients with documented seasonal allergic rhinitis were randomly assigned to receive fluticasone propionate nasal spray 200 micrograms once daily (n = 77), oral fluticasone propionate 5 mg once daily (n = 73), oral fluticasone propionate 10 mg once daily (n = 77), or placebo (n = 77) for 14 days. Plasma fluticasone propionate concentrations were determined at baseline and after 14 days of treatment (day 15). Nasal symptoms were recorded daily by patients and assessed weekly by clinicians. On day 15, more patients in the oral fluticasone propionate 5-mg or 10-mg groups, compared with patients in the fluticasone propionate nasal spray group or the placebo group, had detectable plasma fluticasone propionate concentrations, and mean concentrations were higher in the oral fluticasone propionate groups. Both clinician- and patient-rated total and individual nasal symptom scores for obstruction, rhinorrhea, sneezing, and itching were significantly lower in the fluticasone propionate nasal spray group compared with either of the oral fluticasone propionate groups or the placebo group. With few exceptions, oral fluticasone propionate (5 mg or 10 mg) was not significantly different from placebo on any measures of efficacy. These findings indicate that the efficacy of fluticasone propionate nasal spray (200 micrograms once daily) in the treatment of allergic rhinitis results from direct topical effects rather than from indirect effects after systemic absorption.     

Invasive malignant fibrous histiocytoma in a cow

An invasive malignant fibrous histiocytoma associated with the left cornual process, and causing lysis of the frontal bone, was diagnosed in a cow. The mass compressed the left cerebral hemisphere focally and extended into the frontal sinus and ethmoid and nasal turbinates. It was composed of pleomorphic to spindle-shaped cells with ultra-structural evidence of fibroblastic, myofibroblastic, and fibrohistiocytic differentiation. Trauma and chronic inflammation may be predisposing factors for development of neoplasia in cattle.     

Modeling the interaction between propagating cardiac waves and monophasic and biphasic field stimuli: the importance of the induced spatial excitatory response

INTRODUCTION: Biphasic (BP) defibrillation waveforms have been shown to be significantly more efficacious than equivalent monophasic (MP) waveforms. However, when defibrillation fails, it tends to do so first in distal regions of the heart where induced field gradient magnitudes are lowest. We tested the hypothesis that the improved efficacy of BP waveforms results from their enhanced ability to prevent the initiation of new postshock activation fronts behind preexisting wavetails, rather than from any significantly improved ability to terminate preexisting wavefronts. METHODS AND RESULTS: An idealized computer model of a one-dimensional cardiac strand was used to investigate the spatial and temporal interactions between an underlying propagation front (or tail) and uniform MP or BP field stimuli of various intensities. Axial discontinuities from intercellular junctions induced sawtooth patterns of polarization during such field stimuli, enabling the shocks to interact directly with all cells. MP and BP diastolic thresholds were essentially equal. All suprathreshold MP and BP field stimuli successfully terminated preexisting wavefronts by directly depolarizing tissue ahead of those fronts, thus blocking their continued progression. However, the postshock response at the wavetail was significantly dependent on the shape and strength of the administered field. Low-strength MP stimuli induced an all-or-none excitation response across the wavetail, producing a sharp spatial transmembrane voltage gradient from which a new sustained anterogradely propagating wavefront was initiated. In contrast, low-strength BP field stimuli induced a spatially graded excitatory response whose voltage gradient was insufficient to initiate such a wavefront. Higher-strength MP and BP stimuli both produced graded excitatory responses with no subsequent propagation. CONCLUSIONS: Shock-induced spatial "all-or-none" excitatory responses facilitate, and graded excitatory responses prevent, the postshock initiation of new propagating wavefronts. Moreover, BP field stimuli can induce such graded excitatory responses at significantly lower stimulus strengths than otherwise equivalent MP stimuli. Therefore, these results support an alternative "graded excitatory response" mechanism for the improved efficacy of BP over MP field stimuli in low gradient regions.