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Biomedical subjects

Jun Hasegawa

Publications and source records attributed to Jun Hasegawa.

At least 19 recordsLinked to original sources
PubMed2006

Structure of cytochrome c552 from a moderate thermophilic bacterium, Hydrogenophilus thermoluteolus: comparative study on the thermostability of cytochrome c.

We have studied the structure-thermostability relationship using cytochromes c from mesophilic and thermophilic bacteria; Pseudomonas aeruginosa (PAc(551)) growing at 37 degrees C and Hydrogenobacter thermophilus (HTc(552)) at 72 degrees C and showed that only five residues primarily differentiate their stabilities. For a more comprehensive study, we found Hydrogenophilus thermoluteolus (Pseudomonas hydrogenothermophila) growing at 52 degrees C and showed the moderate stability of the cytochrome c from this bacterium (PHc(552)). To explore the stabilization mechanisms, the crystal structure of PHc(552) was determined by X-ray analysis. The solution structure of HTc(552) elucidated previously by NMR was refined using distributed computational implementation. Furthermore, the recently reported crystal structure of HTc(552) has become available [Travaglini-Allocatelli, C. et al. (2005) J. Biol. Chem. 280, 25729-25734]. When the structures of these three cytochromes c were combined, this revealed that the five residues, corresponding to those mentioned above, determine the difference of stabilities among them as well. These facts suggested the stabilization mechanisms as follows: (1) improved van der Waals interactions by packing optimization at the N-terminal helix, (2) attractive electrostatic interactions with the heme propionate group, and (3) favorable van der Waals interaction with the heme. This comparative study, by supplementing the structural information of PHc(552) with its complementary feature, demonstrates that just a small number of amino acid residues determine the overall molecular stability by means of additivity of the effects of their substitutions. It is interesting that, in naturally occurring proteins, these adaptation strategies are accommodated by these bacteria to survive in the wide range of thermal conditions.

Amino Acid Sequence↗
PubMed2006

High-density presynaptic transporters are required for glutamate removal from the first visual synapse.

Reliable synaptic transmission depends not only on the release machinery and the postsynaptic response mechanism but also on removal or degradation of transmitter from the synaptic cleft. Accumulating evidence indicates that postsynaptic and glial excitatory amino acid transporters (EAATs) contribute to glutamate removal. However, the role of presynaptic EAATs is unclear. Here, we show in the mouse retina that glutamate is removed from the synaptic cleft at the rod to rod bipolar cell (RBC) synapse by presynaptic EAATs rather than by postsynaptic or glial EAATs. The RBC currents evoked by electrical stimulation of rods decayed slowly after pharmacological blockade of EAATs. Recordings of the evoked RBC currents from EAAT subtype-deficient mice and the EAAT-coupled anion current reveal that functional EAATs are localized to rod terminals. Model simulations suggest that rod EAATs are densely packed near the release site and that rods are equipped with an almost self-sufficient glutamate recollecting system.

Animals↗
PubMed2006

Falls of the elderly are associated with burden of caregivers in the community.

BACKGROUND: Little attention has been paid to the impact on caregivers who provide care to a family member who has falls. The purpose of the current study was to determine whether falls of care recipients are associated with caregivers' burden. METHODS: A cross-sectional study of 1874 community-dwelling care recipients and 1478 caregivers was conducted. We examined the characteristics of care recipients and caregivers, including demographic characteristics, depressive mood as assessed by the Geriatric Depression Scale (GDS-15), the basic activities of daily living (bADL), fall history in the past 6 months, and physician-diagnosed chronic diseases to determine whether there was an association with caregivers' burden as assessed by the Zarit Burden Interview (ZBI). RESULTS: A total of 567 care recipients (30.3%) had a history of falls in the past 6 months. The mean ZBI score of caregivers with falls was significantly higher than that of caregivers without falls. There were negative correlations between the ZBI score and recipient bADL score and positive correlations between the ZBI score and GDS-15 scores of the recipient and caregiver, the level of severity of dementia, and the Charlson comorbidity index. Male recipient, fall history, behavioral disturbance, and dementia had significantly higher ZBI scores than those of controls. The stepwise multiple regression analyses found that the GDS-15 score of caregivers and recipients, level of severity of dementia, bADL score, and fall history were independently associated with the ZBI score. CONCLUSION: Among the community-dwelling frail elderly, falls are associated with caregiver burden even when controlling for various possible confounding factors.

Accidental Falls↗
PubMed2006

Effects of hearing level on habitual sniffing in patients with cholesteatoma.

CONCLUSIONS: Quality of hearing is one of the key conditions promoting habitual sniffing associated with closing failure of the Eustachian tube and sniffing may stop with deterioration of hearing due to progression of the disease. OBJECTIVE: To examine habitual sniffing associated with closing failure of the Eustachian tube and hearing level in patients with cholesteatoma. PATIENTS AND METHODS: A total of 171 patients with middle ear cholesteatoma treated in Tohoku University Hospital (106 sides in 101 patients, 63 males and 38 females, mean age 43.3 years), and Sendai Ear Surgi Center (70 sides in 70 patients, 37 males and 33 females, mean age 42.7 years) were included. The patients were divided into two groups based on the presence or absence of habitual sniffing, and pre- and postoperative hearing was compared between the two groups. RESULTS: Habitual sniffing was not found in patients with air conduction hearing level worse than 60 dB or air-bone gap larger than 40 dB. Habitual sniffing disappeared in all patients with expanded air-bone gaps in lower frequency regions after surgery.

Adult↗
PubMed2006

Audiometry with nasally presented masking noise: novel diagnostic method for patulous eustachian tube.

OBJECTIVE: Nasal-noise masking audiometry was developed to assess the acoustic transfer function from the nasopharyngeal cavity to the middle ear via patulous eustachian tube (ET). STUDY DESIGN: Prospective. SETTING: Tertiary referral center. PATIENTS: Twenty-seven ears of 18 patients with patulous ET and 20 ears of 10 healthy subjects with no history of ear disease or complaints of aural symptoms. MAIN OUTCOME MEASURES: Audiometric measurement was conducted with and without masking noise presented in the nasal cavity. RESULTS: The masking effect of nasally presented noise caused elevation of the threshold for the tone presented in the external auditory canal. This threshold elevation was significantly greater, particularly in the lower-frequency region, in ears with patulous ET and was decreased to the normal range after obstructive treatment of the patulous ET. CONCLUSION: Nasal-noise masking audiometry is a simple and effective way to identify patulous ET.

Acoustic Stimulation↗
PubMed2006

Underuse of medications for chronic diseases in the oldest of community-dwelling older frail Japanese.

OBJECTIVES: To test the following hypotheses: (1) the rate of polypharmacy, defined as six or more prescribing medications, is lower in the oldest old (> or =85) than in younger older people (65-84); (2) beneficial medication use is lower in the oldest old; (3) the underuse of these medications in the oldest old is associated with physical or cognitive impairment or comorbid conditions. DESIGN: A cross-sectional study of the baseline data from the Nagoya Longitudinal Study for Frail Elderly. SETTING: Community-based. PARTICIPANTS: One thousand eight hundred seventy-five community-dwelling older people (632 men, 1,243 women). MEASUREMENTS: The data, which were collected at the patients' homes or from care-managing center records, included the clients' demographic characteristics, depression status as assessed using the short version of the Geriatric Depression Scale, a rating for basic activities of daily living (ADLs), prescribed medications, and physician-diagnosed chronic diseases. RESULTS: The oldest old had less polypharmacy even after controlling for ADLs and comorbid conditions. The underuse of beneficial medications for the oldest old was observed after adjusting for ADLs, cognitive impairment, comorbid conditions, antithrombotic agents for subjects with a history of cardiovascular diseases, acetylcholinesterase inhibitors for those with dementia, and antidepressants for those with depression. However, being aged 85 and older was not associated with the underuse of hypoglycemic and antihypertensive agents by those with diabetes mellitus and hypertension, respectively. CONCLUSION: Among community-dwelling frail older people, the rate of polypharmacy is lower in the oldest members than in the younger ones. The underuse of prescribed medications for chronic diseases/conditions of frail older people is common but not for all conditions.

Activities of Daily Living↗
PubMed2006

Day care service use is associated with lower mortality in community-dwelling frail older people.

OBJECTIVES: To clarify the association between day care service use and 21-month mortality in community-dwelling frail older people. DESIGN: Prospective cohort study (the Nagoya Longitudinal Study for Frail Elderly). SETTING: Community-based. PARTICIPANTS: One thousand six hundred seventy-three community-dwelling older people (540 men, 1,133 women). MEASUREMENTS: Data included the clients' demographic characteristics; depression as assessed using the short version of the Geriatric Depression Scale; a rating for basic activities of daily living (ADLs); comorbidity; number of prescribed medications and physician-diagnosed chronic diseases; use of home-care services, including day care, visiting nurse, and home-help services; and number of regular medical checkups. Survival analysis of 21-month mortality was conducted using Kaplan-Meier curves and multivariate Cox proportional hazards models. RESULTS: Of the 1,673 participants, 726 were day care service users at baseline, and 268 (94 day care service users, 174 nonusers) died during the 21-month follow-up. Multivariate Cox regression models adjusting for potential confounders showed that day care service use was associated with reduction in mortality. Subgroup analysis demonstrated that day care service use was associated with less risk of mortality in subjects who were female; were in the youngest age group (65-74); had higher ADL scores, lower comorbidity, depression, no dementia; and used a visiting nurse service. Participants using day care service two and three times or more a week had 63% or 44% lower relative hazard ratios, respectively, than participants not using the service. CONCLUSION: Among community-dwelling frail older people, day care service use two or more times per week was associated with 44% to 63% lower 21-month mortality.

Aged↗
PubMed2006

[High prevalence rate of depression among community-dwelling frail elderly Japanese].

AIM: We examined the prevalence of depression and identified the factors associated with depression among community-dwelling Japanese frail elderly eligible for long-term care insurance. METHODS: A cross-sectional study of the baseline data of 1,409 elderly from the Nagoya Longitudinal Study for Frail Elderly was conducted. The data, which were collected at the patients' homes or from care-managing center records, included the clients' demographic characteristics, care levels, depression as assessed by the short version of the Geriatric Depression Scale (GDS-15), a rating for basic activities of daily living (ADL), prescribing medications, and physician-diagnosed chronic diseases. The participants were considered to be in depression or severe depression if their GDS-15 score was 6 < or = or 10 < or =, respectively. RESULTS: Although 57.2% or 23.1% of the participants had a GDS-15 score of 6 or higher, or 10 or higher, only 4.2% of sever depressive participants were taking an antidepressant. The higher prevalence of depression was associated with a higher care level in the long-term care insurance program. The depression was associated with 3 or more eligible care level, poor nutritional status, 3 or more prescribed medications, and nonuse of day-care/service. CONCLUSION: We observed a high prevalence of depression among community-dwelling Japanese frail elderly eligible for long-term care insurance. However, very few depressed elderly were taking an antidepressant.

Aged↗
PubMed2005

Mechanisms of the inhibitory effect of epigallocatechin-3-gallate on cultured human vascular smooth muscle cell invasion.

OBJECTIVE: Although we recently showed that the administration of catechins reduced the neointimal formation in a rat balloon-injury model, the precise molecular mechanisms are largely unknown. In the present study, we tried to determine these mechanisms using an in vitro SMC invasion system. METHODS AND RESULTS: Boyden chamber assay was used to examine the effect of catechins on the invasive behavior of SMCs. The invasive activity of SMCs through collagen gel was restrained by EGCG in a concentration-dependent manner. The data from gelatin and collagen zymography and Western blot revealed that EGCG blocks the activation of pro-matrix metalloproteinase (MMP)-2 during an invasion assay and in the conditioned medium of cultured SMCs as well as the activities of MMP-2 and membrane type 1-MMP (MT1-MMP) even at 0.1 to 0.3 micromol/L of EGCG. EGCG was found to restrain MT1-MMPcat-dependent pro-MMP-2 activation. EGCG upregulated the expression of tissue inhibitor of MMP-2 (TIMP-2) protein. Reverse zymography showed that the increased TIMP-2 to expression was validated by an increased activity. The data from decreased TIMP-2 activity using its siRNA suggested that upregulation of TIMP-2 expression may be one of the major mechanisms for inhibition of SMC invasion by EGCG. CONCLUSIONS: These results indicate that EGCG targets multiple MMP-mediated SMC cellular events and provides a new major mechanism for the SMC invasion through upregulation of TIMP-2 expression to modulate MMP activity.

Antioxidants↗
PubMed2005

Control of the redox potential of Pseudomonas aeruginosa cytochrome c551 through the Fe-Met coordination bond strength and pKa of a buried heme propionic acid side chain.

Pseudomonas aeruginosa cytochrome c(551) and a series of its mutants exhibiting various thermostabilities have been studied by paramagnetic (1)H NMR and cyclic voltammetry in an effort to elucidate the molecular mechanisms responsible for control of the redox potentials (E degrees ') of the proteins. The study revealed that the E degrees ' value of the protein is regulated by two molecular mechanisms operating independently of each other. One is based on the Fe-Met coordination bond strength in the protein, which is determined by the amino acid side chain packing in the protein, and the other on the pK(a) of the heme 17-propionic acid side chain, which is affected by the electrostatic environment. The former mechanism alters the magnitude of the E degrees ' value throughout the entire pH range, and the latter regulates the pK values reflected by the pH profile of the E degrees ' value. These findings provide novel insights into functional regulation of the protein, which could be utilized for tuning the E degrees ' value of the protein by means of protein engineering.

Bacterial Proteins↗
PubMed2005

Cloning, expression, crystallization and preliminary X-ray characterization of cytochrome c552 from a moderate thermophilic bacterium, Hydrogenophilus thermoluteolus.

The amino-acid sequence of cytochrome c552 (PH c552) from a moderately thermophilic bacterium, Hydrogenophilus thermoluteolus, was more than 50% identical to that of cytochrome c from an extreme thermophile, Hydrogenobacter thermophilus (HT c552), and from a mesophile, Pseudomonas aeruginosa (PA c551). The PH c552 gene was overexpressed as a correctly processed holoprotein in the Escherichia coli periplasm. The overexpressed PH c552 has been crystallized by vapour diffusion from polyethylene glycol 4000 pH 6.5. The crystals belong to space group C222(1), with unit-cell parameters a = 48.98, b = 57.99, c = 56.20 A. The crystals diffract X-rays to around 2.1 A resolution.

Cloning, Molecular↗
PubMed2005

Motor and electrogastrographic activity of the gastric tube formed after esophagectomy.

In order to characterize the motor activity of a surgically constructed gastric tube, several hours of ambulatory intraluminal pressure recordings were performed in 6 patients following esophagectomy and gastric tube construction. Whole pressure waves were spectrally analyzed by Fast Fourier Transform (FFT). Simultaneous abdominal and thoracic electrogastrograms (EGGs) were recorded for about 20 min both before and after meals during ambulatory pressure recording. The pressure waves and EGGs for each 20 min recording were analyzed by the maximal entropy method (MEM). While the motility index of the pressure waves decreased after a meal, the 3 cpm component of these waves (2.4-3.7 cpm) increased significantly (n=6, P<0.05). Both bradygastria (0-2.4 cpm) and the duodeno-respiratory component (10-15 cpm) decreased, while the tachygastria component (3.7-10 cpm) increased, although these differences were not significant. The peak power of the gastric tube abdominal EGGs was significantly larger than that of control abdominal or thoracic EGGs in each of the 1 cpm (0-2.4 cpm), 6 cpm (5.0-7.4) and 8 cpm components (7.5-9.9). The thoracic EGG consisted mainly of the 3 cpm component, while the spectral amplitudes of the 1, 6, 8 and 10 cpm components were below 6% of the 3 cpm component. The peak spectral frequency both of the intraluminal pressure waves by FFT and of the thoracic EGGs by MEM occurred within the 3 cpm component. A cross correlation of about 0.2-0.3 occurred between the thoracic EGGs and the intraluminal pressure waves. Thus the gastric tube seems to preserve most of the original gastric motor characteristics and to contribute as a substitute for the original esophagus and stomach.

Abdomen↗
PubMed2005

Generation of knock-in mice carrying third cones with spectral sensitivity different from S and L cones.

Red-green color vision in primates is unique in the sense that it is mediated by two photoreceptor cells that are indistinguishable in all aspects except for their visual pigments. In order to generate an animal model for investigation of the interaction between red-green inputs at the molecular level, we applied knock-in technology and X-chromosome inactivation machinery to make a mouse model with cone cells possessing visual pigments with different spectral sensitivities. We introduced a S308A point mutation into the Green opsin gene allele on the X-chromosome. This manipulation generated a 24 nm red-shift of absorption maximum in the cone pigment with negligible functional differences in other molecular properties. Amplitudes of responses in ERG and ganglion cell recordings of homozygotes were similar to those of wild-types, although the spectral sensitivities differed. Heterozygotes showed variable spectral sensitivities of ganglion cell responses due to the different integration of the native and the S308A cone inputs on the dendritic fields. In situ hybridization experiments showed that cone cells with respective pigments formed patch-like clusters of specific L cone-types, approximately 30 mum in diameter, which were randomly distributed in the dorsal region of the retinas. Since the patch-like clustering was arranged by X-inactivation, such clustering could be present in the peripheral retinas of New World monkeys with polymorphic L pigments, indicating that our mice would be a suitable model to study evolution of the mammalian color vision system.

Animals↗
PubMed2005

Hydroxylated polychlorinated biphenyls (OH-PCBs): recent advances in wildlife contamination study.

The exposure of wildlife and human population to environmental hazardous contaminants has been of global concern for over several decades. More recently, focus has been directed towards potential toxicological effects such as the endocrine disrupting function of xenobiotics. The hydroxylated metabolites of PCBs (OH-PCBs) should be pointed out as these compounds. OH-PCBs have emerged as important classes of environmental contaminants in wildlife and humans because of their ability to bind with the thyroxin transport protein, transthyretin (TTR), and their interaction with thyroid hormone receptors. However, data on their occurrence in wildlife and their behavior in the matrices of environment are limited. Topics include the formation of OH-PCBs, their physicochemical properties (octanol-water partition coefficient, Kow), analytical procedures and contamination status in wildlife. The guidance for improving the study of OH-PCB contamination is also briefly mentioned.

Animals↗
PubMed2004

Five amino acid residues responsible for the high stability of Hydrogenobacter thermophilus cytochrome c552: reciprocal mutation analysis.

Five amino acid residues responsible for extreme stability have been identified in cytochrome c(552) (HT c(552)) from a thermophilic bacterium, Hydrogenobacter thermophilus. The five residues, which are spatially distributed in three regions of HT c(552), were replaced with the corresponding residues in the homologous but less stable cytochrome c(551) (PA c(551)) from Pseudomonas aeruginosa. The quintuple HT c(552) variant (A7F/M13V/Y34F/Y43E/I78V) showed the same stability against guanidine hydrochloride denaturation as that of PA c(551), suggesting that the five residues in HT c(552) necessarily and sufficiently contribute to the overall stability. In the three HT c(552) variants carrying mutations in each of the three regions, the Y34F/Y43E mutations resulted in the greatest destabilization, by -13.3 kJ mol(-1), followed by A7F/M13V (-3.3 kJ mol(-1)) and then I78V (-1.5 kJ mol(-1)). The order of destabilization in HT c(552) was the same as that of stabilization in PA c(551) with reverse mutations such as F34Y/E43Y, F7A/V13M, and V78I (13.4, 10.3, and 0.3 kJ mol(-1), respectively). The results of guanidine hydrochloride denaturation were consistent with those of thermal denaturation for the same variants. The present study established a method for reciprocal mutation analysis. The effects of side-chain contacts were experimentally evaluated by swapping the residues between the two homologous proteins that differ in stability. A comparative study of the two proteins was a useful tool for assessing the amino acid contribution to the overall stability.

Amino Acids↗
PubMed2004

Complete thermal-unfolding profiles of oxidized and reduced cytochromes C.

The complete thermal-unfolding profiles of both oxidized and reduced forms of cytochrome c551 (PA) from mesophilic Pseudomonas aeruginosa and cytochrome c552 (HT) from thermophilic Hydrogenobacter thermophilus were obtained by the newly developed pressure-proof cell compartment installed in a circular dichroic spectrometer, which facilitates protein thermal-unfolding experiments up to 180 degrees C. The thermodynamic cycle, which relates protein stability and redox function, indicated that the redox potentials of PA and HT in the native state are regulated by the stability of the oxidized proteins rather than by that of the reduced ones.

Circular Dichroism↗
PubMed2004

Effects of axial methionine coordination on the in-plane asymmetry of the heme electronic structure of cytochrome c.

The paramagnetic susceptibility ( chi) tensors of the oxidized forms of thermophile Hydrogenobacter thermophilus cytochrome c(552) (Ht cyt c(552)) and a quintuple mutant (F7A/V13 M/F34Y/E43Y/V78I; qm) of mesophile Pseudomonas aeruginosa cytochrome c(551) (Pa cyt c(551)) have been determined on the basis of the redox-dependent (1)H NMR shift changes of the main-chain NH and C(alpha)H proton resonances of non-coordinated amino acid residues and the NMR structures of the reduced forms of the corresponding proteins (J. Hasegawa, T. Yoshida, T. Yamazaki, Y. Sambongi, Y. Yu, Y. Igarashi, T. Kodama, K. Yamazaki, Y. Kyogoku, Y. Kobayashi (1998) Biochemistry 37:9641-9649; J. Hasegawa, S. Uchiyama, Y. Tanimoto, M. Mizutani, Y. Kobayashi, Y. Sambongi,Y. Igarashi (2000) J Biol Chem 275:37824-37828). From the chi tensors determined, we obtained the contact shifts for heme methyl proton resonances, which provided the heme electronic structures of the oxidized forms of Ht cyt c(552) and qm. We also characterized the heme electronic structure of the cyanide adducts of the proteins, where the axial Met was replaced by an exogenous cyanide ion, through the analysis of (1)H NMR spectra. The results indicated that the heme electronic structures of both the proteins in their oxidized forms with axial His and Met coordination are largely different to each other, while those in their cyanide adducts are similar to each other. These results demonstrated that the orientation of the axial Met sulfur lone pair, with respect to heme, predominantly contributes to the spin delocalization into the porphyrin-pi system of heme in the oxidized proteins with axial His and Met coordination.

Cytochromes c↗