Biomedical subjects

Nobutaka Shimizu

Publications and source records attributed to Nobutaka Shimizu.

16 recordsLinked to original sources

PubMed2006

Dose dependence of radiation damage for protein crystals studied at various X-ray energies.

Radiation damage to protein crystals is the most serious problem in obtaining accurate structures from protein crystallography. In order to examine the photon energy dependence of radiation damage, 12 to 15 data sets from each of nine tetragonal lysozyme crystals were collected at nine different X-ray energies (6.5, 7.1, 8.3, 9.9, 12.4, 16.5, 20.0, 24.8 and 33.0 keV) using beamline BL41XU at SPring-8. All results were compared on the basis of absorbed dose, expressed in Gray (Gy). Crystallographic statistics, such as the values of lattice constants, R(merge) and I/sigma(I), for each data set degraded at all nine energies as the exposure time for each crystal increased. In all data sets, radiation damage was observed after the absorbed dose exceeded 10(6) Gy. However, from the point of view of crystallographic statistics normalized to the absorbed dose, no clear dependence on photon energy was observed in these results. Structural refinement showed that the average B-factor for the last data set was larger than that for the first data set at all energies tested. However, no energy dependence of radiation damage on B-factor was found. Furthermore, disruption of disulfide bonds due to radiation damage was observed in electron density maps even at the highest photon energy (33 keV) used in this study. Therefore, these results suggest that radiation damage in the energy range investigated could be evaluated based on absorbed dose without energy dependence, and that it is important to minimize the absorbed dose in a crystal sample for obtaining an accurate protein structure.

Computer Simulation↗

PubMed2006

Bladder leiomyosarcoma in a boy.

Leiomyosarcoma of the urinary bladder in childhood is uncommon. We reported bladder leiomyosarcoma in a 10-year-old boy. He had underwent the extirpation of the tumor and performed adjuvant chemotherapy in combination with radiotherapy according to the Intergroup Rhabdomyosarcoma Study-4 regimens. He had no evidence of recurrence of disease after 3 years.

Child↗

PubMed2006

The crystal structure of the R52Q mutant demonstrates a role for R52 in chromophore pKa regulation in photoactive yellow protein.

Mutating arginine 52 to glutamine (R52Q) in photoactive yellow protein (PYP) increases the pK(a) of the chromophore by 1 pH unit. The structure of the R52Q PYP mutant was determined by X-ray crystallography and was compared to the structure of wild-type PYP to assess the role of R52 in pK(a) regulation. The essential differences between R52Q and the wild type were confined to the loop region containing the 52nd residue. While the hydrogen bonds involving the chromophore were unchanged by the mutation, removing the guanidino group generated a cavity near the chromophore; this cavity is occupied by two water molecules. In the wild type, R52 forms hydrogen bonds with T50 and Y98; these hydrogen bonds are lost in R52Q. Q52 is linked to Y98 by hydrogen bonding through the two water molecules. R52 acts as a lid on the chromophore binding pocket and controls the accessibility of the exterior solvent and the pK(a) of the chromophore. R52 is found to flip out during the formation of PYP(M). The result of this movement is quite similar to the altered structure of R52Q. Thus, we propose that conformational changes at R52 are partly responsible for pK(a) regulation during the photocycle.

Arginine↗

PubMed2006

Ligand-induced domain rearrangement of fatty acid beta-oxidation multienzyme complex.

The quaternary structure of a fatty acid beta-oxidation multienzyme complex, catalyzing three sequential reactions, was investigated by X-ray crystallographic and small-angle X-ray solution scattering analyses. X-ray crystallography revealed an intermediate structure of the complex among the previously reported structures. However, the theoretical scattering curves calculated from the crystal structures remarkably disagree with the experimental profiles. Instead, an ensemble of the atomic models, which were all calculated by rigid-body optimization, reasonably explained the experimental data. These structures significantly differ from those in the crystals, but they maintain the substrate binding pocket at the domain boundary. Comparisons among these structures indicated that binding of 3-hydroxyhexadecanoyl-CoA or nicotinamide adenine dinucleotide induces domain rearrangements in the complex. The conformational changes suggest the structural events occurring during the chain reaction catalyzed by the multienzyme complex.

Catalysis↗

PubMed2006

[Clinical study of acute urinary retention].

PURPOSE: Clinical study of acute urinary retention seen by this department. MATERIALS AND METHODS: Subjects were 206 cases seen during office hours and during after-hours emergency care by the department of Urology at the Kinki University Hospital for acute urinary retention for the 12-year-period from April 1993 to April 2005. RESULTS: By gender, the 206 cases of acute urinary retention included 175 men (85%) and 31 women (15%). The ratio of men to women was 5.6:1, with a markedly larger number of male cases. Ages of the 206 cases overall were distributed from 6 to 93 years old and the mean age was 66 years old. In male cases, the mean age was 69.6 years old while in female cases it was 46.3 years old. With regard to the cause, bladder outlet obstruction (BOO) accounted for 123 (70.3%) of the 175 male cases; benign prostatic hyperplasia (BPH) was noted in 92 cases and accounted for 52.6%of the total. Detrusor Weakness (DW) was noted in 35 cases (20%). DW was most prevalent in women, being noted in 20 cases (64.5%). With regard to treatment, in male cases surgery was performed for BOO in 69 (56%) of 123 cases; surgery was performed for BPH in 56 (60.8%) of 92 cases, drug therapy was used in 19 cases, and 3 cases were observed. In female cases, 10 cases were able to urinate on their own through treatment of the causative disorder. With regard to outcome, ultimately a total of 139 cases (67.5%), 125 men and 14 women, were able to urinate on their own. CONCLUSIONS: 1. 85% of acute urinary retention cases were men. Of these, 70% were caused by some form of BOO. DW due to a cause other than obstruction accounted for about 70% of the remaining 30%. 2. Overall, 70% of cases were able to urinate on their own after treatment while 30% required catheterization. 3. After the cause of BOO was eliminated, cases were likely to be able to urinate on their own; CIC (clean intermittent catheterization) was frequently used in treatment of causes other than BOO. 4.15% of acute urinary retention cases were women.

Acute Disease↗

PubMed2005

pH-dependent equilibrium between long lived near-UV intermediates of photoactive yellow protein.

The long lived intermediate (signaling state) of photoactive yellow protein (PYP(M)), which is formed in the photocycle, was characterized at various pHs. PYP(M) at neutral pH was in equilibrium between two spectroscopically distinct states. Absorption maxima of the acidic form (PYP(M)(acid)) and alkaline form (PYP(M)(alkali)) were located at 367 and 356 nm, respectively. Equilibrium was represented by the Henderson-Hasselbalch equation, in which apparent pK(a) was 6.4. Content of alpha- and/or beta-structure of PYP(M)(acid) was significantly greater than PYP(M)(alkali) as demonstrated by the molar ellipticity at 222 nm. In addition, changes in amide I and II modes of beta-structure in the difference Fourier transform infrared spectra for formation of PYP(M)(acid) was smaller than that of PYP(M)(alkali). The vibrational mode at 1747 cm(-1) of protonated Glu-46 was found as a small band for PYP(M)(acid) but not for PYP(M)(alkali), suggesting that Glu-46 remains partially protonated in PYP(M)(acid), whereas it is fully deprotonated in PYP(M)(alkali). Small angle x-ray scattering measurements demonstrated that the radius of gyration of PYP(M)(acid) was 15.7 Angstroms, whereas for PYP(M)(alkali) it was 16.2 Angstroms. These results indicate that PYP(M)(acid) assumes a more ordered and compact structure than PYP(M)(alkali). Binding of citrate shifts this equilibrium toward PYP(M)(alkali). UV-visible absorption spectra and difference infrared spectra of the long lived intermediate formed from E46Q mutant was consistent with those of PYP(M)(acid), indicating that the mutation shifts this equilibrium toward PYP(M)(acid). Alterations in the nature of PYP(M) by pH, citrate, and mutation of Glu-46 are consistently explained by the shift of the equilibrium between PYP(M)(acid) and PYP(M)(alkali).

Bacterial Proteins↗

PubMed2005

Medial prefrontal cortex lesions inhibit reflex micturition in anethetized rats.

The medial prefrontal cortex is thought to participate in the control of micturition and urinary continence, based on evidence from clinical reports, but its exact role is not fully understood. This study investigated whether ibotenic acid lesions of the medial prefrontal cortex would influence volume-evoked micturition in urethane-anesthetized rats. The incidence and amplitude of bladder contractions were recorded during continuous saline infusion (0.1 ml/min) immediately before and 1 week after ibotenic acid (0.5 microg) or vehicle (0.5 microl) was injected into the medial prefrontal cortex. Vehicle injection did not change the incidence or amplitude of bladder contractions compared to pre-injection values. Ibotenic acid lesions prolonged the time interval between bladder contractions significantly although it did not affect the amplitude of bladder contractions. Histological analysis revealed that ibotenic acid lesions were restricted primarily to the anterior cingulate and prelimbic cortices. Larger ibotenic acid lesions extending ventrally into the infralimbic cortex produced a variable response but did not change the overall incidence or amplitude of bladder contractions significantly. These data indicate that the medial prefrontal cortex influences the timing of bladder contractions but does not affect contraction amplitudes.

Anesthesia↗

PubMed2005

Edaravone protects against ischemia/reperfusion-induced functional and biochemical changes in rat urinary bladder.

OBJECTIVES: To investigate the effects of edaravone on ischemia/reperfusion (I/R) injury in the rat bladder. Increasing evidence has shown that I/R are major etiologic factors in the progression of bladder dysfunction induced by partial outlet obstruction, and that part of the damage is due to the generation of free radicals. Edaravone is a newly developed radical scavenging agent that has been used for protection against I/R injury in patients with cerebral infarction. METHODS: Thirty-five adult male rats were divided into five groups. Groups 1 to 4 underwent 1 hour of ischemia followed by 1 hour of reperfusion. Groups 1 to 3 were treated with edaravone at 1, 3, or 10 mg/kg body weight and group 4 with saline. Group 5 consisted of age-matched control rats. In vivo ischemia was created by clamping the vesical arteries for 1 hour. Reperfusion was accomplished by removing the clips and also lasted for 1 hour. Edaravone or saline was administered into the femoral artery after reperfusion for 30 minutes. After reperfusion, the bladder was excised and separated. The responses to electrical field stimulation, carbachol, and KCl were recorded. Other materials were analyzed for malondialdehyde as a measure of lipid peroxidation. RESULTS: Edaravone administration resulted in protection of the contractile responses to both field stimulation and carbachol, although the responses to KCl were not affected. I/R resulted in an increase in malondialdehyde, which was reduced to control levels by edaravone. CONCLUSIONS: These results suggest that edaravone has a potential protective effect against I/R-induced damage in the rat bladder.

Animals↗

PubMed2005

[Pharmacological evaluation of efficacy and safety of propiverine hydrochloride in patients of overactive bladder--relationship between urodynamic observation and propiverine pharmacokinetics--].

PURPOSE: The clinical benefit of propiverine hydrochloride against overactive bladder was evaluated, and the relationships between urinary voiding functions and the pharmacokinetics were investigated by means of clinical pharmacology with PK/PD approach. PATIENTS AND METHODS: Total 7 patients suffering urgency with urinary frequency and incontinence received propiverine hydrochloride in doses of 10 mg qd or 20 mg qd for 4 weeks, and then the doses were switched in cross-over manner to continue the treatment for further 4 weeks. The urodynamic measurements as well as pharmacokinetic samplings were done before the medication, 4 weeks and 8 weeks after the starting medication, to examine the dose-response and concentration-response relationships. RESULTS: The volume at first desire to void increased according to dose increased, and the volume at first involuntary contraction tended to increase according to both dose and drug concentration in plasma. However, no apparent dose-response relationships were observed for maximum urinary flow rate and the detrusor pressure at the maximum urinary flow rate. The PK/PD analysis using Emax model suggested that, approximately 75 ng/mL of the propiverine concentration in plasma allowed the increase in the volume at first involuntary contraction for 50%. The urinary residual volume increased in dose-dependent manner only in the patients with severe grade of lower urinary tract obstruction, but scarcely increased in the patients with moderate grade or below. CONCLUSION: Propiverine hydrochloride improved the urinary voiding functions with a tendency to depend on both dose and concentration in plasma. After the administration of propiverine hydrochloride, the concentration in plasma will immediately reach the level at which the drug can increase in the volume at first involuntary contraction for 50%, and then the concentration level will sustain the effect ranging from 10% to 50% increase in bladder volume. Furthermore, the lower urinary tract obstruction will be a predictor of increase in urinary residual volume.

Aged↗

PubMed2005

[Squamous cell carcinoma of bladder producing granulocyte colony-stimulating factor (G-CSF): a case report].

A case of bladder squamous cell cancer producing granulocyte colony-stimulating factor (G-CSF) is reported. A 55-year-old female presented with macroscopic hematuria and urinary frequency. Cystoscopy demonstrated a large bladder tumor. Computerized tomography and magnetic resonance imaging of abdomen and pelvis showed bladder muscular invasion, but did not reveal any metastasis. Laboratory examination showed leukocytosis of 23,600/mm3 and a high value of granulocyte colony-stimulating factor (G-CSF) 77 pg/ml in the peripheral blood. She was diagnosed with bladder tumor 52 months ago, but did not seek further medical attention. The leukocyte count at that point was 10,700/mm3. Radical cystectomy was performed. The histopathological diagnosis was well differentiated squamous cell carcinoma. Immunohistochemical examination was positive for G-CSF. After surgery the leukocyte count and value of G-CSF in the peripheral blood decreased to the normal range. There has been no recurrence of cancer for 10 months after surgery without any adjuvant therapy.

Biomarkers, Tumor↗

PubMed2005

[Small cell carcinoma of the prostate: an autopsy case report].

An 83-year-old man was admitted to our hospital with complaints of dysuria and backache. The patient had been treated with a luteinizing hormone-releasing hormone analogue for 2 years. An enlarged, hard and nodular prostate was palpable on digital rectal examination. Serum level of prostate specific antigen (PSA) was 1.7 ng/ml (0-4) and that of neuron-specific enolase (NSE) were 263.2 ng/ml (0-10). Multiple bone and liver metastases were seen on CT and MRI. After hospitalization, the liver function worsened markedly. The patient died on the 34th hospital day, and was diagnosed with primary small cell carcinoma of prostate on autopsy.

Aged↗

PubMed2004

Disturbed voiding associated with von Recklinghausen's neurofibroma.

Abstract The incidence of von Recklinghausen's disease is most frequent among neurocutaneous syndrome. It is an intractable neurological disease. About half of the disease is caused by an autosomal dominant inheritance and another half by mutation. It is characterized by neurofibroma which arises on various parts of the body and gradually increases with age. We recently encountered a case where a growth of neurofibroma within the foreskin had disturbed voiding.

Humans↗

PubMed2003

[Statistics on operations at the Department of Urology, Nippon Telegraph and Telephone West Osaka Hospital during a ten-year period from 1992 to 2001].

A 10-year clinical statistic survey was made on the operations performed at the department of urology, NTT West Osaka Hospital between 1992 and 2001. The total number of operations was 2,540, comprising 1,899 males and 641 females, and a total of 1,559 ESWL was performed. The number of operations per year was chronologically constant. Major operations were transurethral resection of prostate (299 cases). The number of operations for malignant tumors, for examples radical nephrectomy and enucleation for renal cancer, nephroureterectomy for upper urinary tract cancer and transurethral resection of bladder tumor, has increased since 1997 year after year. The number of ESWL has decreased since 1994. Renal transplantation was introduced in 1995, and laparoscopic adrenalectomy also in 1997.

Female↗

PubMed2002

Light-induced global conformational change of photoactive yellow protein in solution.

The light-induced global conformational change of photoactive yellow protein was directly observed by small-angle X-ray scattering (SAXS). The N-terminal 6, 15, or 23 amino acid residues were enzymatically truncated (T6, T15, or T23, respectively), and their near-UV intermediates were accumulated under continuous illumination for SAXS measurements. The Kratky plot demonstrated that illumination induced partial loss of globularity. The change in globularity was marked in T6 but very small in T15 and T23, suggesting that structural change in positions 7-15 mainly reduces the globularity. The radius of gyration (R(g)) estimated by Guinier plot was increased by 1.1 A for T6 and 0.7 A for T15 and T23 upon illumination. As T23 lacks most of the N-terminal loop, structural change in the main part composed of the PAS core, helical connector, and beta-scaffold caused an increase of R(g) by 0.7 A. The structural change of positions 7-15 caused an additional increase by 0.4 A. The decrease of R(g) upon truncation of positions 7-15 for dark state was 0.3 A, while that for the intermediate was 0.7 A, suggesting that this region moves outward on formation of the intermediate. These results indicate that a light-induced structural change of PYP takes place in the main part and N-terminal 15 amino acid residues. The former induces only dimensional increase, but the latter results in additional change in shape.

Bacterial Proteins↗

PubMed2002

Effect of organic anions on the photoreaction of photoactive yellow protein.

In order to clarify changes in the structure and surface properties of photoactive yellow protein (PYP) upon light absorption, the spectroscopic properties and solution structure of its photo-intermediate (PYP(M)) were examined in the presence of various anions. At identical ionic strengths, citrate slowed the decay rate of PYP(M) more than acetate. Although the absorption spectrum in the dark was not affected by organic anions, citrate induced a 5-nm blue shift of the absorption maximum for PYP(M). Solution X-ray scattering experiments indicated that the radius of gyration (Rg) and apparent molecular weight in the dark were constant in all buffer systems. However, the Rg of PYP(M) in citrate buffer at high concentration was 16.2 (+/-0.2) A, while the Rg of PYP(M) in acetate buffer was 15.6 (+/-0.2) A. The apparent molecular weight increased 7% upon PYP(M) formation in citrate buffer at high concentration compared to other conditions. These results suggest that citrate molecules specifically bind to PYP(M). A cluster of basic amino acid residues with a hydrogen bond donor would be exposed upon PYP(M) formation and responsible for the specific binding of citrate.

Acetates↗

PubMedDate not supplied

Preparation of large crystals of photoactive yellow protein for neutron diffraction and high resolution crystal structure analysis.

The exact positions of all the hydrogen atoms in photoactive yellow protein (PYP) is important for understanding the molecular mechanism of the photoreaction because the protonation/deprotonation of certain amino acid residues and rearrangements in the hydrogen bond network are involved in the conformational changes of PYP. Neutron crystallography is one of the most effective methods to determine the hydrogen positions. However, a large crystal is required for neutron crystallography because a neutron-incident flux is quite limited. In addition, the crystal should be grown from heavy water to reduce the incoherent background from hydrogen. We prepared a large crystal of PYP (dimensions: 1.5 x 0.7 x 0.7 mm3) for neutron crystallography using ammonium sulfate with sodium chloride. The obtained large crystal gave X-ray diffraction spots up to 0.84 angstroms. Although some of the hydrogen atoms could be observed in the high resolution X-ray crystal structure, functionally important hydrogen atoms were impossible to see, indicating the importance of neutron crystallography. Thus, we optimized the crystallization conditions with heavy water and successfully obtained neutron diffraction spots up to 2.1 angstroms with the crystal in D2O.

Bacterial Proteins↗