The Usefulness of T1-Weighted Magnetic Resonance Images for Diagnosis of Acute Leukemia Manifesting Musculoskeletal Symptoms prior to Appearance of Peripheral Blood Abnormalities.

The patients with acute leukemia occasionally present with musculoskeletal symptoms initially, including bone pain, joint pain, muscular pain, and functional impairment. Without abnormal findings of peripheral blood cell counts or smear, the correct diagnosis tends to be delayed. Magnetic resonance imaging is often performed to examine musculoskeletal abnormalities; it can simultaneously reveal the bone marrow composition with high anatomical resolution and excellent soft tissue contrast. We present 4 pediatric patients who were initially diagnosed with acute pyogenic osteomyelitis or arthritis, based on the elevated white blood cell counts and/or C-reactive protein in addition to the localized high signal intensity on T2-weighted magnetic resonance images. Finally, they were diagnosed with B-cell precursor acute lymphoblastic leukemia by bone marrow examination. The period between the onset of musculoskeletal symptoms and the diagnosis of leukemia ranged from 20 days to 6 months. In all cases, the T1-weighted magnetic resonance images taken prior to detection of peripheral blood abnormality revealed diffuse low signal intensity of the bone marrow in regions adjacent or contralateral to localized musculoskeletal symptoms. These findings should raise the suspicion of leukemia even without abnormalities in peripheral blood.

Epilepsy in children with trisomy 18.

Although the reported incidence of epilepsy associated with trisomy 18 is 25-50%, there have been no detailed descriptions of the characteristics of trisomy 18-related epilepsy. We investigated the characteristics of epilepsy in children with trisomy 18 who remained alive for over 1 year by sending questionnaires to pediatric neurologists belonging to the Kyoto Multi-institutional Study Group of Pediatric Neurology. Eleven patients with trisomy 18 were enrolled (age at the study, from 15 to 134 months; median, 43 months), of whom seven (64%) had epilepsy. The age at seizure onset ranged from 1 to 42 months (median: 11 months). Among the seven patients with epilepsy, two had focal epilepsy, four had generalized epilepsy including infantile spasms in three, and the remaining one had an unclassified type. Seizure seminology included complex partial seizures in both the patients with focal epilepsy. At the time of the investigation, three children with generalized epilepsy still had daily seizures, while the remaining four were seizure-free. In conclusion, the characteristics of epilepsy in patients with trisomy 18 were as follows: over half of the children developed epilepsy during infancy or early childhood; infantile spasms might be one of the common epileptic syndromes; the epilepsy was intractable in half of the children, especially in those with generalized epilepsy.

Characterization of chronic idiopathic thrombocytopenic purpura in Japanese children: a retrospective multi-center study.

The objectives of this study are to clarify (1) the difference in demographic and clinical variables at initial presentation between acute and chronic idiopathic thrombocytopenic purpura (ITP), and (2) the prognostic factors of patients with chronic ITP. We conducted a retrospective analysis of 247 children with newly diagnosed ITP between April 1991 and March 2006 who visited one of the 12 hospitals belonging to the Kyoto University Pediatric Hematologic Study Group. 180 and 67 cases were classified as the acute type and as the chronic type, respectively. Older age, higher initial platelet count, positive medical history or concomitant medical diagnosis, the absence of preceding infection or vaccination, and the absence of an increase in immunoglobulin were risk factors for the chronicity. The prognostic factors in chronic ITP were evaluated in 53 patients after excluding patients receiving splenectomy or having insufficient follow-up data. The overall time required for 50% resolution in patients with chronic ITP was approximately 5.6 years. Age at presentation of less than 3 years and higher platelet counts at the time of chronic ITP diagnosis were good prognostic factors. On the other hand, gender, initial platelet counts, and preceding infection or vaccination were not associated with the prognosis.

Uncovering the physical origin of the difference between aliphatic chain and aromatic ring in the "hydrophobic" effects on partial molar volume.

The partial molar volume changes in the transfer of several hydrophobic molecules, which are composed of aromatic rings and an aliphatic chain of different lengths, from carbon tetrachloride to water (DeltaV(hyd)) are calculated using the three-dimensional interaction site model theory of molecular solvation. The theory reproduces recent experimental observations: the addition of a methyl group decreases DeltaV(hyd); in contrast, the addition of an aromatic ring increases DeltaV(hyd). The discrepancy is found to originate from the difference between chain and ring structures rather than that between aliphaticity and aromaticity. Furthermore, a general rule of the variation in DeltaV(hyd) due to the addition of a hydrocarbon is found through the theoretical analysis. An outward addition at the trans position, which is to form chain structure, decreases DeltaV(hyd), while an inward addition at the cis position, which is to form ring structure, increases DeltaV(hyd). This is explained in terms of solvent packing rather than the so-called hydrophobic hydration. The present findings argue against the traditional idea that the hydrophobic hydration can be represented by the observed values of DeltaV(hyd).

[New theories in molecular mechanisms of general anesthesia].

The controversy of the specific and non-specific theories of general anesthesia continues. Recently, non-specific theory began to have the persuasive power again. Molecular dynamics of target protein of the anesthesia and hydrophobic dehydration due to anesthetic binding to the protein have been drawing researcher's attention. Clarification of relation between the action site and the molecular dynamics modification of target protein has been an increasingly important problem. With these contexts, I believe that the research in anesthetic mechanisms will shift to a new paradigm.

Effects of pressure and solvents on the infrared absorption intensities of C-I stretching modes of methyl and ethyl iodides in solutions.

We have investigated effects of pressure and solvents on infrared intensities of methyl and ethyl iodides in solutions using a hydrostatic high-pressure cell with synthetic diamond windows. We focused on the absolute intensity of the C-I stretching mode, which was measured in carbon disulfide solvent up to 300MPa and at 293K, and in n-hexane solvent at 298K. For comparison, we investigated the effect of solvents on the absorption intensity. Effects of pressure and solvents on the infrared intensity were analyzed using two electrostatic models, which assume the shape of solute cavity as sphere or spheroid. The latter model is approximately in agreement with both effects on the intensity, particularly, for the pressure effect. This paper demonstrated that the electrostatic model taking the shape of the cavity into account is useful to explain the medium effect on the infrared intensity and also suggests that more improved models could provide information of the solvation structure from the medium effect on the infrared intensity.

A case of Hirayama disease.

We report a male patient with Hirayama disease aged 13. The disease was insidiously progressive and he had severe disability of the right hand at presentation. He had muscular atrophy in the intrinsic muscles of the right hand and in the distal muscles of the right forearm. The atrophy was pronounced on the ulnar side. Cold paresis was also noticed. There was no sensory disturbance. On Electromyography, neurogenic changes were recorded in several atrophic muscles. Motor and sensory nerve conduction was normal. MR images of the spinal cord were normal when it was performed with a conventional method (i.e., without neck flexion). However, characteristic MR findings were obtained when the patient lay with maximum neck flexion. The posterior wall of the cervical dural canal was shifted anteriorly at the C3-7 vertebral level, which caused cord compression at the lower cervical spinal canal. The epidural space was crescent-shaped and showed high signal intensity on T2-weighted imaging. These clinical features are typical of Hirayama disease. Pediatrician should be aware of this disease and treat it as soon as possible in order to prevent progression of the atrophy.

Promotion of insulin aggregation by protein disulfide isomerase.

We examined the aggregation of insulin as a result of reduction of disulfide bonds catalyzed by protein disulfide isomerase (PDI) using various techniques. We demonstrated the kinetic correlation between PDI-catalyzed insulin reduction and the aggregate formation, the relationship between aggregation and amyloid formation, and the structural information on the secondary structure of the aggregates. The initial rate of PDI-catalyzed reduction of insulin, apparent rate constants of aggregate growth for sigmoidal features, and lag times were obtained by changing the PDI concentration, temperature, and insulin concentration. In situ kinetics were studied using the dyes; thioflavin T (ThT) and Congo red (CR) in addition to turbidimetry with the insulin reduction by PDI. The ThT and CR binding assay revealed sigmoidal kinetics, and the spectrum of binding CR showed a red shift against time, suggesting an orderly formation of insulin aggregates. The secondary structure of the PDI-promoted insulin aggregates showed antiparallel beta-sheet conformation by FT-IR measurement.

Pressure effects on the structure and function of human thioredoxin.

Thioredoxin is one of the major proteins that catalyze disulfide reduction and defines the thioredoxin superfamily bearing the CXXC structural motif. Human thioredoxin contains only 1 Trp residue proximal to the active site (WCGPC). We are interested in thioredoxin structure-function relationships, in particular, active site hydration and flexibility. Hence, in this study, we used hydrostatic pressure as a perturbation and monitored the conformational changes around the active site of thioredoxin by analyzing Trp fluorescence. The structure of thioredoxin was drastically altered by increasing pressure and did not completely refold after pressure release. The conformation in the active site vicinity was modified at low pressure (less than 100 MPa) and the Trp residue was completely exposed to aqueous medium at pressures above 350 MPa. Upon pressure release, thioredoxin showed no activity, although it folded 80% of the alpha-helical content relative to the native state. According to these results, pressure denaturation induces critical damage for the activity of thioredoxin, indicating extreme fragility of the active site with respect to pressure. This result is in contrast to the pressure effect on protein disulfide isomerase (PDI) which is organized by four thioredoxin-like domains including two WCGHC motifs.

Pressure effects on the heat-induced aggregation of equine serum albumin by FT-IR spectroscopic study: Secondary structure, kinetic and thermodynamic properties.

Pressure can restrain the heat-induced aggregation and dissociate the heat-induced aggregates. We investigated the aggregation-preventing pressure effect and the aggregates-dissociating pressure effect to characterize the heat-induced aggregation of equine serum albumin (ESA) by Fourier transform infrared spectroscopy. The results suggest that the alpha-helical structure collapses at the beginning of heat-induced aggregation, then the rearrangement of structure from partially unfolded structure to the intermolecular beta-sheet takes place through the activated state. We determined the activation volume for the heat-induced aggregation (DeltaV( not equal)=+92+/-8 ml mol(-1)) and the partial molar volume difference between native state and heat-induced aggregates (DeltaV(N-->HA)=+32 ml mol(-1)). This positive partial molar volume difference suggests that the heat-induced aggregates have larger internal voids than the native structure. Moreover, the positive volume change implies that the formation of the intermolecular beta-sheet is unfavorable under high pressure. We also determined the free energy profile of ESA. This energy profile explains the restriction of the formation of heat-induced aggregates by pressure. These results explain the structural differences between heat-induced aggregates with intermolecular beta-sheet and pressure-induced aggregates without intermolecular beta-sheet.

Pressure stability of the alpha-helix structure in a de novo designed protein (alpha-l-alpha)(2) studied by FTIR spectroscopy.

The pressure-induced structural changes of a de novo designed four-helix bundle protein, (alpha-l-alpha)(2), in aqueous solution have been investigated by FTIR spectroscopy. Changes in the amide I' band intensity show that pressure induces disruption of tertiary interactions and stabilizes the solvated alpha-helical form. This may suggest that the exposure of the hydrophobic core to the solvent by pressure is not a sufficient condition for pressure-induced unfolding of the alpha-helices of proteins.

The secondary structure of pressure- and temperature-induced aggregates of equine serum albumin studied by FT-IR spectroscopy.

The protein aggregation is divided into amyloid fibrils and amorphous aggregates. Amyloid fibrils are composed of the 3-dimensional ordered structure and are bound to thioflavin T and Congo red dyes. The amorphous aggregates with the disordered structure do not bind to these dyes. We have investigated the pressure- and heat-induced aggregates of equine serum albumin (ESA) from the secondary structural viewpoint using FT-IR spectroscopy. We show the secondary structural differences between heat- and pressure-induced aggregates of ESA. The heat-induced irreversible aggregates of ESA are composed of the intermolecular beta-sheet structure without binding thioflavie T and Congo red to be amorphous form. On the other hand, the pressure-induced reversible aggregates are composed of the random structure to be also amorphous form. From the comparison of pressure effects on ESA in native and reducing conditions of disulfide bridges, we demonstrate that the restriction of structural flexibility by disulfide bridges is an important factor for the reversibility of the pressure-induced aggregation.

Cold denaturation of ubiquitin at high pressure.

Cold-induced conformational transition of ubiquitin was studied at pH 4.5 under a constant pressure of 2 kbar using variable pressure one-dimensional 1H and two-dimensional 15N/1H NMR spectroscopy as well as IR spectroscopy. Although a tendency for preferential stabilization of a peculiar locally disordered and partially hydrated conformer I, identical with that previously found with variable-pressure NMR at 0 degrees C, is recognized, the transition of the folded conformer N to the unfolded conformer U occurs largely cooperatively with decreasing temperature, reaching near completion at - 21 degrees C. NMR spectral features as well as the analysis of NMR relaxation parameters indicate that the polypeptide chain is almost fully unfolded, fairly well-hydrated and floppy at - 21 degrees C, whereas the IR spectrum shows a substantial decrease of the beta-sheet. The Gibbs energy change from the folded state (a mixture of N and I) to the unfolded state at 2 kbar obtained from the 1H NMR data is fitted well with a single DeltaCp value of 2.43 +/- 0.13 (kJ/K mol) for the entire temperature range between - 21 and 90 degrees C, covering both the cold denaturation and heat denaturation, showing that the two denatured states actually belong to a single thermodynamic phase of the protein. The DeltaCp value determined at 2 kbar is substantially smaller than the DeltaCp determined at 1 bar (3.8-5.8 (kJ/Kmol), which is consistent with the fact that the denaturation takes place from a mixture of N and I at 2 kbar rather than from pure N at 1 bar.

The pressure effect on the structure and functions of protein disulfide isomerase.

Studying on the pressure effects of the structure and functions of the multidomain protein, protein disulfide isomerase (PDI), the intrinsic Trp fluorescence spectra of PDI were measured under high pressure. PDI has 5 Trp residues and the two of all Trp residues are located at the neighborhood of the active site (WCGHC) for isomerase activity. On the basis of the red shift of center of spectral mass (CSM) of the intrinsic Trp fluorescence and the decrease in its fluorescence intensity, the changes in tertiary structure of PDI were observed above 100 MPa. These structural changes were completed at 400 MPa. The CSM of 400 MPa denatured PDI was comparable to that of 6.0 M GdnHCl denatured one. All of the Trp residues included in PDI are completely exposed to aqueous medium at 400 MPa. However, there is the significant difference between the pressure and GdnHCl-denatured PDI. The Trp fluorescence intensity was decreased with increasing pressure, but increased with the increase of the GdnHCl concentration. It is implied that the pressure-denatured state of PDI might remain compact not to be extensively unfolded. In the point of view about the reversibility of pressure-treated PDI, the tertiary structure was completely recovered after released to ambient pressure. The disulfide reduction and chaperone activity of 400 MPa-treated PDI were also recovered to be comparable to those of native one. Despite of a multidomain protein, the excellence in both structural and functional recovery of pressure-denatured PDI is quite remarkable. These unique properties of PDI against high pressure provide the insights into understanding the pressure-induced denaturation of PDI.

Understanding high pressure stability of helical conformation of oligopeptides and helix bundle protein high pressure FT-IR and RISM theoretical studies.

The pressure effect on conformational equilibria of simple organic compounds and the pressure denaturation of proteins have been well investigated by using vibrational spectroscopy. However, there was no systematic investigation of the pressure effect on conformational equilibria of oligopeptides, which are located between the simple organic compounds and proteins. Here, we review the recent vibrational spectroscopic and theoretical studies of the pressure effect on conformational equilibria of model oligopeptides and helix bundle protein in aqueous solution.

Theoretical study of volume changes associated with the helix-coil transition of an alanine-rich peptide in aqueous solution.

The changes in the partial molar volume (PMV) associated with the conformational transition of an alanine-rich peptide AK16 from the alpha-helix structure to various random coil structures are calculated by the three-dimensional interaction site model (3D-RISM) theory coupled with the Kirkwood-Buff theory. The volume change is analyzed by decomposing it into contributions from geometry and hydration: the changes in the van der Waals, void, thermal, and interaction volume. The total change in the PMV is positive. This is primarily due to the growth of void space within the peptide, which is canceled in part by the volume reduction resulting from the increase in the electrostatic interaction between the peptide and water molecules. The changes in the void and thermal volume of the coil structures are widely distributed and tend to compensate each other. Additionally, the relations between the hydration volume components and the surface properties are investigated. We categorize coil structures into extended coils with the PMV smaller than helix and general coils with the PMV larger than helix. The pressure therefore can both stabilize and destabilize the coil structures. The latter seems to be a more proper model of random coil structures of the peptide.

Pressure-tuning FT-IR spectroscopic study on the helix-coil transition of Ala-rich oligopeptide in aqueous solution.

We investigated the effect of pressure on the helix-coil transition of an Ala-rich peptide (AK16: YGAAKAAAAKAAAAKA-NH(2)) in aqueous solution by FT-IR spectroscopy. The spectra of the amide I' region of AK16 in aqueous solution was decomposed into some component bands using a curve fitting method. The peak at around 1635 cm(-1) corresponding to the solvent exposed alpha-helix conformer increases with increasing pressures, while the peak at around 1655 cm(-1) corresponding to the random coil conformer decreases. From the pressure dependence of the band intensities, we determined the volume change from the alpha-helix to random coil conformers of AK16 to be +10.5+/-0.3 cm(3)/mol. The positive volume change is different from the negative volume change generally observed in the pressure denaturation of proteins.

Basement membrane fragility underlies embryonic lethality in fukutin-null mice.

Fukuyama-type congenital muscular dystrophy (FCMD), associated with brain malformation due to defects in neuronal migration, is caused by mutations in fukutin. Several lines of evidence suggest that the fukutin protein plays a pivotal role in synthesis of O-mannosyl sugar moieties of alpha-dystroglycan, a cell surface laminin receptor. Here, through targeted disruption of the orthologous mouse fukutin gene, we show that the fukutin protein is essential, as homozygous-null embryos die by E9.5 of gestation. Fukutin-null embryos show phenotypic diversity, features of which include growth retardation, folding of the egg cylinder, leakage of maternal red blood cells into the yolk sac cavity, and an increased number of apoptotic cells in the ectoderm. Loss of immunoreactivity against sugar moieties in alpha-dystroglycan suggests a reduced laminin-binding capacity. Ultrastructural analysis shows thin and breached basement membranes (BMs). BM fragility may underlie all of these abnormal phenotypes, and maintenance of BM function may require fukutin-mediated glycosylation of alpha-dystroglycan early in embryonic development.

Effects of pressure and solvents on the infrared absorption intensity of the O-H stretching mode of phenol in solutions.

Effects of pressure and solvents on the infrared spectrum of phenol in solutions have been investigated using a hydrostatic high-pressure cell with synthetic diamond windows. For the first time, we performed a quantitative investigation of the effect of pressure on the absolute intensity of O-H stretching mode up to 150 MPa (in CCl4) and 200 MPa (in CS2). For comparison, we measured the effect of solvents on the absorption intensity. The Polo-Wilson theory, which is the most traditional theory for medium effects on the intensity, was tested for present results. The pressure dependence was in sufficient agreement with their formula, while the solvent dependence is unsatisfactory. This suggests that the traditional intensity correction by Polo-Wilson's formula is practically valid for pressure-tuning infrared experiments.

Chain-length dependence of alpha-helix to beta-sheet transition in polylysine: model of protein aggregation studied by temperature-tuned FTIR spectroscopy.

The chain-length dependence of the alpha-helix to beta-sheet transition in poly(L-lysine) is studied by temperature-tuned FTIR spectroscopy. This study shows that heterogeneous samples of poly(L-lysine), comprising polypeptide chains with various lengths, undergo the alpha-beta transition at an intermediate temperature compared to homogeneous ingredients. This holds true as long as each individual fraction of the polypeptide is capable of adopting an antiparallel beta-sheet structure. The tendency is that the longer chain is, the lower the alpha-beta transition temperature is, which has been linked to the presence of distorted or solvated helices with turns or beta sheets in elongating chains of poly(L-lysine). As such helical structures are apparently conducive to the alpha-beta transition, this draws a comparison to the hypothesis of metastable protein conformational states being a common stage in amyloid-formation pathways. The antiparallel architecture of the beta sheet is likely to reflect the pretransition interhelical interactions in poly(L-lysine). Namely, the chains are arranged in an antiparallel manner because of energetically favored antiparallel pre-assembly of dipolar alpha helices.