The WASP-WAVE protein network: connecting the membrane to the cytoskeleton

Wiskott-Aldrich syndrome protein (WASP) and WASP-family verprolin-homologous protein (WAVE) family proteins are scaffolds that link upstream signals to the activation of the ARP2/3 complex, leading to a burst of actin polymerization. ARP2/3-complex-mediated actin polymerization is crucial for the reorganization of the actin cytoskeleton at the cell cortex for processes such as cell movement, vesicular trafficking and pathogen infection. Large families of membrane-binding proteins were recently found to interact with WASP and WAVE family proteins, therefore providing a new layer of membrane-dependent regulation of actin polymerization.     

Degranulation of mast cells located in median eminence in response to compound 48/80 evokes adrenocortical secretion via histamine and CRF in dogs

The effect of intracerebroventricular infusion of compound 48/80 (C48/80), a mast cell secretagogue, on adrenal cortisol secretion was investigated in dogs under pentobarbital sodium anesthesia. A marked increase in adrenal cortisol secretion was elicited by C48/80 along with a concomitant increase in the plasma levels of cortisol and immunoreactive ACTH, but neither arterial blood pressure and heart rate nor the plasma histamine level altered significantly. Pretreatment with either anti-CRF antiserum or pyrilamine maleate (H(1) histamine-receptor antagonist) significantly attenuated the C48/80-evoked increase in cortisol secretion, but pretreatment with metiamide (H(2)-receptor antagonist) significantly potentiated it. Significant attenuation of the C48/80-evoked increase in cortisol also occurred in dogs given ketotifen, a mast cell stabilizing drug, before pharmacologic challenge. In the pars tuberalis and median eminence (ME), mast cells were highly concentrated in close association with the primary plexus of the hypophysial portal system. Degranulated mast cells were extensively found in the ME of C48/80-treated animals. These results suggest that mast cells located in these regions liberated histamine within the brain as a result of degranulation induced by C48/80 and that this led to activation of the hypothalamic-pituitary-adrenocortical axis.     

Molecular mechanisms of invadopodium formation: the role of the N-WASP-Arp2/3 complex pathway and cofilin

Invadopodia are actin-rich membrane protrusions with a matrix degradation activity formed by invasive cancer cells. We have studied the molecular mechanisms of invadopodium formation in metastatic carcinoma cells. Epidermal growth factor (EGF) receptor kinase inhibitors blocked invadopodium formation in the presence of serum, and EGF stimulation of serum-starved cells induced invadopodium formation. RNA interference and dominant-negative mutant expression analyses revealed that neural WASP (N-WASP), Arp2/3 complex, and their upstream regulators, Nck1, Cdc42, and WIP, are necessary for invadopodium formation. Time-lapse analysis revealed that invadopodia are formed de novo at the cell periphery and their lifetime varies from minutes to several hours. Invadopodia with short lifetimes are motile, whereas long-lived invadopodia tend to be stationary. Interestingly, suppression of cofilin expression by RNA interference inhibited the formation of long-lived invadopodia, resulting in formation of only short-lived invadopodia with less matrix degradation activity. These results indicate that EGF receptor signaling regulates invadopodium formation through the N-WASP-Arp2/3 pathway and cofilin is necessary for the stabilization and maturation of invadopodia.     

Stability of 5-aminolevulinic acid on dried urine filter paper for a diagnostic marker of tyrosinemia type I

The chemical diagnosis of tyrosinemia type I generally involves the detection of succinylacetone (SA) in patient urine. However, 5-aminolevulinate (5ALA), which accumulates due to succinylacetone's inhibition of porphyrin synthesis, can also be used as diagnostic metabolites. Here we examined the stabilities of these markers on dried urine filter paper. After two weeks at room temperature, the succinylacetone was 10% of its original level, but over 80% of 5-aminolevulinate remained. Thus, although insufficient succinylacetone was recovered from dried urine filter paper to diagnose tyrosinemia type I, 5-aminolevulinate was readily detected, permitting the diagnosis.     

Histochemical differences of the lectin affinities of backbone polylactosamine structures carrying the ABO blood group antigens in papillary carcinoma and other types of thyroid neoplasm

Summary Endo--galactosidase from Escherichia freundii cleaves linear polylactosamine structure as follows: R-GlcNAc-1-3Gal-1-4GlcNAc-1-R + H₂O R-GlcNAc-1-3Gal + GlcNAc-1-R. Staining with Griffonia simplicifolia agglutinin II (GSA-II) following enzyme digestion reveals the distribution of R-GlcNac-1-3Gal-1-4GlcNAc-1-R structures in tissue sections. In this study, the procedure was applied to formalin-fixed, paraffin-embedded tissue sections from 26 cases of papillary carcinomas including 2 follicular variants, 8 follicular carcinomas, 7 adenomas, 1 anaplastic carcinoma and 1 medullary carcinoma in order to investigate whether different types of polyactosamine-containing structure are produced in these thyroid neoplasms. Simultaneously, the susceptibility of the ABH antigens expressed in these neoplastic cells to endo--galactosidase digestion was examined. Most of the papillary carcinoma cells from all the individuals examined were strongly stained by GSA-II following enzyme digestion. Without enzyme digestion, little or no reactivity with GSA-II was observed. Among other types of neoplasms, only one case of follicular carcinoma exhibited reactivity with GSA-II following enzyme digestion. ABH antigens were expressed in 22 cases of papillary carcinomas, 2 adenomas, 5 follicular carcinomas and 1 anaplastic carcinoma, and their expression was dependent on the ABO blood group of the patients. Endo--galactosidase digestion resulted in the elimination of these antigens not only in papillary carcinomas but also in other neoplasms. These results suggested that the polylactosamine-containing structures produced in papillary carcinomas are quite different from those in other neoplasms, and demonstrated that the procedure is a useful diagnostic means for distinguishing papillary carcinoma and other types of thyroid neoplasms.     

Urine screening of five-day-old newborns: metabolic profiling of neonatal galactosuria

We determined urinary galactose and 4-hydroxyphenyllactic acid (4HPLA) in 4338 of 5-day-old newborns using a newly developed GC–MS screening method. Fifty-two infants were chemically diagnosed as having transient galactosuria based upon elevated urinary galactose levels (4.78–30.53 mg/mg creatinine, control 1.10±0.89 mg/mg creatinine). These infants did not excrete galactitol or galactonic acid into the urine, which is typical of hereditary galactosemia. Nearly 40% of the transient galactosuria was associated with immature infants (low birth weight or borne before 37 gestational weeks). Immature hepatic function is one explanation for neonatal transient galactosuria, but heterozygotes or the carriers of galactose degradation enzyme deficiencies were also suspected in some of the newborns, judging from the comparisons of urinary galactose and 4HPLA excretion between neonates and patients with galactosemia.     

Rat intestinal epithelial cells modify the reactivity of bovine milk lactoferrin to concanavalin A

Bovine milk lactoferrin suppressed proliferation of concanavalin A-stimulated rat spleen lymphocytes by absorbing mitogenic lectin activity. Culture media, conditioned by incubating allogeneic intestinal epithelial villus and crypt cells with or without lactoferrin, also suppressed the proliferation. Villus cells absorbed lactoferrin during preparation of conditioned medium and the medium lost a lactoferrin-dependent lymphocyte proliferation-suppressing activity. Although crypt cells did not absorb lactoferrin, its conditioned medium lost the activity. These conditioned media did not alter proliferation of lymphocytes stimulated with 12-O-tetradecanoylphorbol-13-acetate plus ionomycin. Serum proteins, albumin and transferrin, did not substitute for milk lactoferrin. Thus, intestinal epithelial cells modified the reactivity of milk lactoferrin to concanavalin A.     

Effects of blood flow to the prefrontal cortex on high-intensity exercise combined with high-decibel music

We studied the effects of high-intensity exercise (70-75% of VO₂ max) combined with high-decibel music (100 dB) on cognitive function (measured by the Stroop test) and related blood flow changes to the prefrontal cortex (measured by Oxy-hemoglobin (Hb), Deoxy-Hb, tissue oxygen index (TOI), and normalized tissue hemoglobin index (nTHI)). The subjects of the study were 28 healthy female university students in their early 20s. Subjects were categorized into control group (CG), music group (MG), exercise group (Ex), and music and exercise group (MnEx). A crossover design was implemented so that all subjects participated in all test groups. We found no significant difference in reaction time between CG and MG for the neutral and incongruent tasks of Stroop test. However, there were significant improvements in the neutral and incongruent tasks for both the Ex (p < 0.01) and MnEx (p < 0.01) groups. Oxy-Hb measurements in the prefrontal cortex of the brain supported the Stroop test data. We found no difference between Ex and MnEx in the TOI; however, there was a significant decrease (p < 0.05) in MnEx compared to Ex. In addition, Ex resulted in a significant increase (p < 0.05) in nTHI as compared to CG. These results indicate that high decibel music could negatively affect prefrontal cortex activation of the brain during exercise.     

Phosphoinositide-binding domains: Functional units for temporal and spatial regulation of intracellular signalling

Inositol phospholipid (phosphoinositide) is a versatile lipid characterized by its isomer-specific localization, as well as its molecular diversity attributable to phosphorylation events. Phosphoinositides act as signal mediators in a spatially and temporally controlled manner. Information about the timing and location of their production is received by phosphoinositide-binding proteins and transmitted to multiple lines of intracellular events such as signal transduction, cytoskeletal rearrangement, and membrane trafficking. Among those proteins, a significant portion possess globular structural units, called domains, which are specialized for phosphoinositide binding. The pleckstrin homology (PH) domain was the first phosphoinositide-binding domain identified. It contains the largest number of members and is associated with the formation of signalling complexes on the plasma membrane. Recent studies identified other novel phosphoinositide-binding domains (Fab1p, YOTB, Vps27p, EEA1 (FYVE), Phox homology (PX), and epsin N-terminal homology (ENTH)), thus extending our knowledge of how the functional versatility of phosphoinositides is achieved.