Biomedical subjects

K Kitamura

Publications and source records attributed to K Kitamura.

At least 73 records · Page 4Linked to original sources

PubMed2001

Adrenomedullin (11-26): a novel endogenous hypertensive peptide isolated from bovine adrenal medulla.

Adrenomedullin (AM) is a potent hypotensive peptide originally isolated from pheochromocytoma tissue. Both the ring structure and the C-terminal amide structure of AM are essential for its hypotensive activity. We have developed an RIA which recognizes the ring structure of human AM. Using this RIA, we have characterized the molecular form of AM in bovine adrenal medulla. Gel filtration chromatography revealed that three major peaks of immunoreactive AM existed in the adrenal medulla. The peptide corresponding to Mr 1500 Da was further purified to homogeneity. The peptide was determined to be AM (11-26) which has one intramolecular disulfide bond. Amino acid sequences of bovine AM and its precursor were deduced from the analyses of cDNA encoding bovine AM precursor. The synthetic AM (11-26) produced dose-dependent strong pressor responses in unanesthetized rats in vivo. The hypertensive activity lasted about one minute, and a dose dependent increase in heart rate was also observed. The present data indicate that AM (11-26) is a major component of immunoreactive AM in bovine adrenal medulla and shows pressor activity.

Adrenal Medulla↗

PubMed2001

A family affected by branchio-oto syndrome with EYA1 mutations.

Branchio-oto (BO) syndrome is complicated with congenital preauricular fistulae, branchial fistulae (cysts), and hearing loss (sensorineural, conductive or mixed). As well as branchio-oto-renal (BOR) syndrome. it is known to be an autosomal dominant hereditary disorder. Since mutations in the EYA1 gene have been identified in both BO and BOR syndromes, mutation screening of this gene has been drawing attention as a genetic test to diagnose BOR/BO syndromes. In this study, we genetically investigated the presence of EYA1 mutations in a BO syndrome family in which we observed congenital preauricular fistulae, branchial fistulae (cysts) and hearing loss in four generations. Whereas there was a variety of phenotype expressions in this family, all subjects tested had a nonsense mutation (R264X) in exon 8 of the EYA1 gene. The present report adds further examples to support the usefulness of molecular genetic testing for the diagnosis of patients with BO syndrome.

Branchio-Oto-Renal Syndrome↗

PubMed2001

Determination of partition coefficients of diazepam and flurazepam between phosphatidylcholine bilayer vesicles and water by second derivative spectrophotometric method.

Second derivative spectrophotometry allowed the establishment of a simple and accurate method for the determination of partition coefficients of benzodiazepine drugs in a liposome/water system. The absorption spectra of diazepam (DZ) and flurazepam (FZ) in phosphatidylcholine (egg yolk) bilayer vesicle suspensions showed small spectral changes depending on the concentration of phosphatidylcholine vesicles. However, the intense background signals caused by the light scattering of the phosphatidylcholine vesicles made it difficult to yield a correct base line, thus the quantitative spectral data could not be obtained. In the second derivative spectra, the spectral changes were enhanced and three derivative isosbestic points were observed for each drug indicating the entire elimination of the residual background signal effects. The derivative intensity change of each drug (DeltaD) induced by its interaction with phosphatidylcholine bilayers was measured at a specific wavelength. From the relationship between the DeltaD value and the lipid concentration, the molar partition coefficients (K(p)s) of DZ and FZ were calculated and obtained with a good precision of R.S.D below 10%. The fractions of the partitioned DZ and FZ calculated by using the obtained K(p) values agreed well with the experimental values. The results prove that the derivative method can be usefully and easily applied to the determination of partition coefficients of benzodiazepines in the liposomes/water system without any separation procedures.

Algorithms↗

PubMed2001

Phosphorylation of Mei2 and Ste11 by Pat1 kinase inhibits sexual differentiation via ubiquitin proteolysis and 14-3-3 protein in fission yeast.

Fission yeast Pat1 kinase inhibits sexual differentiation by phosphorylating the meiotic inducer Mei2 and the transcription factor Ste11. Here, we show how Pat1 downregulates these proteins. Mei2 is degraded via a ubiquitin-proteasome pathway in a phosphorylation-dependent fashion. The E2 Ubc2 and the E3 Ubr1 are required for this proteolysis. In addition, Pat1 negatively regulates Ste11 via Rad24/14-3-3, thereby repressing mei2+ transcription. The Pat1 phosphorylation sites of Ste11 match the consensus recognition sequence for 14-3-3. Rad24 binds preferentially to phosphorylated Ste11, and this binding results in inhibition of the transcriptional activation capacity of Ste11. Overall, therefore, these results show that Pat1 coordinates concerted molecular mechanisms that govern the sexual differentiation developmental decision.

14-3-3 Proteins↗

PubMed2001

Clinical significance of vascular endothelial growth factor-C (VEGF-C) in breast cancer.

Vascular endothelial growth factor-C (VEGF-C) is a specific ligand which induces lymphangiogenesis. We examined the expression of VEGF-C protein to determine its role in the progression of breast cancer. Immunohistochemical analysis revealed that VEGF-C was overexpressed in 39 of 98 breast cancer specimens (39.8%) but not in adjacent normal mammary glands. The expression of VEGF-C showed a significant correlation with lymphatic vessel invasion (p = 0.0004). It is noteworthy that the 5-year disease free survival rate of the VEGF-C positive group was significantly poorer than that of negative group (p = 0.0356). We suggest that as expression of VEGF-C is not implicated in lymphatic spread, it may prove to be a promising marker to predict the recurrence of breast cancer.

Adult↗

PubMed2001

Functional analysis for peripheral myelin protein PASII/PMP22: is it a member of claudin superfamily?

Two major glycoproteins, P0 and PASII/PMP22, are specifically expressed in peripheral myelin. Point mutations of these proteins and over or under expression of PASII/PMP22 cause various hereditary peripheral neuropathies. P0 is well characterized as a major adhesion molecule in PNS myelin, but the function of PASII/PMP22 is still unknown. Recently, an oligodendrocyte-specific protein (OSP) was identified as a member of the claudin family and as a component of tight junctions of central myelins. Since PASII/PMP22 shows similarity in structure to OSP, which is a tetraspan membrane protein, we speculated if PASII/PMP22 could be a member of claudin superfamily. The primary structure of PASII/PMP22 showed a significant homology of 48% and a 21% identity with the OSP sequence. Exogenous expression of PASII/PMP22 in C6 cells significantly inhibited BrdU incorporation to the cells. The C6 cells stably transfected with PASII/PMP22 cDNA showed no homophilic cell adhesive activity. When dorsal root ganglion (DRG) neurons were cocultured on PASII/PMP22 expressing cells, both neurite extension and branching of DRG neurons were significantly inhibited. These results indicate that PASII/PMP22 may play a role in a turning point of Schwann cell development from proliferation to differentiation. On the other hand, the cells expressing claudin family proteins are reported to show strong cell adhesive activity and an ability to form tight junctions with neighboring cells. For this reason, we currently do not have any functional data supporting that PASII/PMP22 is the member of claudin superfamily.

Animals↗

PubMed2001

Position effect of the excision frequency of the Antirrhinum transposon Tam3: implications for the degree of position-dependent methylation in the ends of the element.

We identified eight independent Tam3 copies residing in the same Antirrhinum majus genome. All the copies showed excision at 15 degrees C, but not at 25 degrees C. Under conditions promoting excision, each copy appeared to transpose in the leaves and flower lobes with a nearly constant frequency, whereas individual transposition abilities varied widely: the most active copy had an excision frequency more than 100-fold greater than that of the least active one. Despite the different transposition abilities, the structures of the eight Tam3 copies were almost identical. These results made it clear that the transpositional ability of Tam3 is regulated by chromosomal position, but they do not imply position-dependent transposase activity. The position effect of the Tam3 transposition was found to be correlated to the methylation state of the copy's end regions: DNA methylation in the Tam3 end regions tended to suppress the excision activity, and the degree of methylation was dependent on the chromosomal position. Our results also provide evidence of de novo methylation provoked by transposition of the endogenous element. We propose a mechanism of transpositional regulation of plant transposons that responds to the degree of methylation as determined by chromosomal position.

Blotting, Southern↗

PubMed2001

Multifocal micronodular pneumocyte hyperplasia and lymphangioleiomyomatosis in tuberous sclerosis with a TSC2 gene.

A 45-year-old woman with a long-standing diagnosis of tuberous sclerosis (TSC) is presented. She has multifocal micronodular pneumocyte hyperplasia (MMPH) and lymphangioleiomyomatosis (LAM) of the lung, together with the detection of TSC2 gene mutation. During surgery for spontaneous pneumothorax, an open-lung biopsy was performed. Micronodules were well defined, measuring approximately 4 mm in diameter. These MMPHs were histologically composed of papillary proliferation of Type II pneumocytes, with positive immunoreactivity of keratin and surfactant apoprotein. The cystlike spaces, with dilatation and destruction of air spaces, were diffusely formed, and the walls were composed of the spindle cells. Such LAM showed positive immunoreactivity for HMB-45 (a monoclonal antibody specific for human melanoma) and tuberin (the gene product of TSC2). On germline mutation analysis using leukocytes of the present patient, a TSC2 gene mutation was confirmed as a deletion of G (or g) on Exon 9 by polymerase chain reaction-single-strand conformational polymorphism. However, no mutation was detected in her son. With microdissection analysis using paraffin-embedding lung tissues, LOH of the TSC2 gene preliminarily was detected in a LAM lesion but not in MMPH. It is suggested that MMPH, in addition to LAM, could be another pulmonary lesion in TSC patients and that the detection of TSC2 and/or TSC1 gene could essentially be useful for the pathogenesis of MMPH and LAM in TSC patients.

Adult↗

PubMed2001

Improved diabetic syndrome in C57BL/KsJ-db/db mice by oral administration of the Na(+)-glucose cotransporter inhibitor T-1095.

1. The therapeutic effects of an orally active inhibitor of Na(+)-glucose cotransporter (SGLT), T-1095 (a derivative of phlorizin; 3-(benzo[b]furan-5-yl)-2',6'-dihydroxy-4'-methylpropiophenone 2'-O-(6-O-methoxycarbonyl-beta-D-glycopyranoside)) were examined in C57BL/KsJ-db/db (db/db) mice, a genetic animal model of obese type 2 diabetes. 2. The higher renal SGLT activity in db/db mice than normoglycaemic C57BL/KsJ-db/+m (db/+m) mice may support the rationale for using an SGLT inhibitor in the treatment regimen for type 2 diabetes. Both T-1095 and its metabolite, T-1095A, which had approximately 10 times more potency, effectively inhibited renal SGLT activity of these mice in vitro. 3. Single oral administration of T-1095 (10, 30, 100 mg kg(-1), p.o.) to db/db mice caused a dose-dependent reduction in blood glucose levels and a concomitant increase in glucose excretion into urine. In contrast, T-1095 only slightly affected blood glucose levels in db/+m mice. 4. Chronic administration of T-1095 (0.1% w w(-1) pellet chow, for 12 weeks) decreased blood glucose and haemoglobin A(1C) levels, and improved glucose intolerance in db/db mice. The age-related decrease in plasma insulin levels was markedly inhibited and there was a 2.5 fold increase of insulin content in the pancreas of T-1095-treated db/db mice. Food consumption was not changed, while impaired body weight gain was ameliorated by T-1095 treatment. 5. Both the development of albuminuria and the expansion of glomerular mesangial area in db/db mice were significantly suppressed by chronic T-1095 treatment, indicating the prevention of the progression of diabetic nephropathy. 6. These results demonstrate that the SGLT inhibitor T-1095 is able to improve the metabolic abnormalities and inhibit the development of diabetic complications in db/db mice. Thus, T-1095 can be used for therapy of type 2 diabetic patients.

Animals↗

PubMed2001

Gamma-rays enhance rAAV-mediated transgene expression and cytocidal effect of AAV-HSVtk/ganciclovir on cancer cells.

Adeno-associated virus (AAV) vector has several unique properties suited for gene therapy applications. However, relatively low efficiency of transgene expression, which is mainly due to a limited second-strand synthesis from the single-stranded AAV genome, can be a problem in some applications that require potent gene expression such as antitumor applications. Recently, gamma-ray irradiation has been reported to enhance the second-strand synthesis of the AAV genome, and consequently transgene expression. We demonstrate here that an AAV vector harboring the herpes simplex virus type-1 thymidine kinase (HSVtk) is able to kill cancer cells more efficiently when used in combination with gamma-ray irradiation. A human maxillary sinus cancer cell line, NKO-1, was efficiently killed in combination with HSVtk transduction and ganciclovir (GCV), as expected. More importantly, gamma-ray irradiation of practical dosages augmented the cytocidal effect of the HSVtk/GCV system. Southern analysis indicated that gamma-rays enhanced the double-strand synthesis of the rAAV genome in NKO-1 cells. These findings suggest that the combination of rAAVtk/GCV suicide gene therapy with radiotherapy has synergistic effects in the treatment of cancers and may lead to a reduction of the potential toxicity of both rAAVtk/GCV and gamma-ray irradiation.

Antiviral Agents↗

PubMed2001

Epidermal growth factor and 1alpha,25-dihydroxyvitamin D3 suppress lipogenesis in hamster sebaceous gland cells in vitro.

We have previously reported the establishment of a culture system of hamster auricular sebocytes. Although their morphologic and biochemical properties are very similar to those of human sebocytes, the regulation of lipogenesis is not clear. Therefore, we investigated the effect of epidermal growth factor, all-trans retinoic acid, 1alpha,25-dihydroxyvitamin D3, and androgens such as testosterone and 5alpha-dihydrotestosterone on lipogenesis in cultured hamster sebocytes. Intracellular lipid droplets detected with Oil-Red-O staining were observed in 5 d cultures and increased in a time-dependent manner; 40.7% +/- 1.11% of 2 wk cultured cells tested lipid-positive by flow cytometric analysis. When the hamster sebocytes were cultured in the presence of epidermal growth factor, all-trans retinoic acid, or 1alpha,25-dihydroxyvitamin D3, the intracellular lipid droplets were diminished by all-trans retinoic acid and epidermal growth factor, and slightly by 1alpha,25-dihydroxyvitamin D3. The intracellular lipid droplets consisted mainly of triglycerides (71.8%) and, as minor components, cholesterol (18.0%), wax esters (3.6%), and free fatty acids (6.6%). Epidermal growth factor and all-trans retinoic acid decreased the intracellular accumulation of triglycerides (92.6% and 96.1% inhibition, respectively) and free fatty acids (54.3% and 62.6% inhibition, respectively) in the sebocytes. In addition, 1alpha,25-dihydroxyvitamin D3 decreased the triglyceride level (34.3% inhibition), but augmented the accumulation of wax esters (30% increase). There was no difference in the level of cholesterol as a result of these treatments, however. In contrast, 5alpha-dihydrotestosterone augmented the formation of intracellular lipid droplets along with an increase in the accumulation of triglycerides in hamster sebocytes. Our findings that regulation of lipogenesis by all-trans retinoic acid and androgen in hamster sebocytes is identical to regulation in humans suggest that hamster sebocytes are useful for the elucidation of sebaceous function at the cellular level. Furthermore, this is the first evidence that epidermal growth factor and 1alpha,25-dihydroxyvitamin D3 may act as suppressors in the regulation of lipogenesis in hamster sebocytes in vitro.

Animals↗

PubMed2001

PAR-6 regulates aPKC activity in a novel way and mediates cell-cell contact-induced formation of the epithelial junctional complex.

BACKGROUND: PAR-6, aPKC and PAR-3 are polarity proteins that co-operate in the establishment of cell polarity in Caenorhabditis elegans and Drosophila embryos. We have recently shown that mammalian aPKC is required for the formation of the epithelia-specific cell-cell junctional structure. We have also revealed that a mammalian PAR-6 forms a ternary complex with aPKC and ASIP/PAR-3, and localizes at the most apical end of the junctional complex in epithelial cells. RESULTS: The ternary complex formation and junctional co-localization of PAR-6 with aPKC and ASIP/PAR-3 are observed during the early stage of epithelial cell polarization. In addition, over-expression of the PAR-6 mutant with CRIB/PDZ domain in MDCK cells disturbs the cell-cell contact-induced junctional localization of tight junction proteins, as well as inhibiting TER development. Furthermore, the binding of Cdc42:GTP to the CRIB/PDZ domain of PAR-6 enhances the kinase activity of PAR-6-bound aPKC. Detailed analyses suggest that the binding of PAR-6 to aPKC has the intrinsic potential to activate aPKC, which is only released when Cdc42:GTP binds to the CRIB/PDZ domain. CONCLUSION: The results indicate the involvement of PAR-6 in the aPKC function which is required for the cell-cell adhesion-induced formation of epithelial junctional structures, possibly through the cooperative regulation of aPKC activity with Cdc42.

Adaptor Proteins, Signal Transducing↗

PubMed2001

A vasoactive agent enhances the effect of ATP on cochlear blood flow.

Adenosine 5'-triphosphate (ATP) has a dual effect on cochlear blood flow (CBF). It induces on initial, transient decrease, which is followed by a long-lasting increase in CBF. Dilazep dihydrochloride (DZ) is a unique vasoactive agent, which increases CBF as well as decreases systemic blood pressure (BP). This agent is thought to keep the concentration of extracellular adenosine higher than normal by preventing the absorption of adenosine into the intracellular space. Thus, it may potentiate the effect of ATP on CBF. In this study, the effects are reported of ATP on CBF in the presence of DZ as measured with a laser Doppler flowmeter in the guinea pig cochlea. In the presence of DZ, the effect of ATP on CBF was amplified and prolonged. Specifically, the mean value of CBF initially decreased to 30.0% of the baseline, determined prior to administration of any agent, and increased gradually up to 287.8% of the baseline. This pharmaceutical effect of DZ might involve a longer activation of both P2x- and P2y-purinoceptors.

Adenosine Triphosphate↗

PubMed2001

Electronystagmographic findings in patients with cerebral degenerative disease.

Cerebral degenerative diseases produce a variety of abnormal neuro-otological findings on electronystamography (ENG). To assess their diagnostic value and determine which manifestations are helpful in making a diagnosis, ENG findings from 72 cases of confirmed cerebral degenerative disease were analyzed. We observed a high incidence of saccadic pursuit and upward ocular dysmetria, which is likely to be useful in diagnosing cerebral degenerative disease. In contrast, moderate incidences of horizontal ocular dysmetria, gaze-evoked and rebound nystagmus, vertical positioning nystagmus and impaired visual suppression appeared to reflect the degree of dysfunction, while optokinetic nystagmus appeared to reflect both the presence of disease and its severity. Cases of spinocerebellar ataxia 6 and spinocerebellar ataxia 3 tended to produce horizontal and vertical gaze-evoked nystagmus, whereas progressive supranuclear palsy produced a higher incidence of upward gaze-evoked nystagmus, and positioning nystagmus at the sagittal plane appeared frequently in cases of non-hereditary spinocerebellar degeneration.

Electronystagmography↗

PubMed2001

A family with liddle's syndrome caused by a mutation in the beta subunit of the epithelial sodium channel.

Liddle's syndrome is a rare form of autosomal-dominant salt-sensitive hypertension. Constitutive activation of the amiloride-sensitive distal renal epithelial sodium channel (ENaC) is essential for salt-sensitive hypertension. Recently, several DNA analysis studies have indicated that there is a mutation of C-terminus of either the beta or y subunit. We sequenced the C-termini of the beta and -gamma subunits of the ENaC in a Japanese family with hypertension and hypopotassemia without excess minerarocorticoids, clinically diagnosed as Liddle's syndrome. The mutation of the ENaC of this family was beta R564X. Since such case seem to be rare in the literature, detailed data are shown in this report.

Aldosterone↗

PubMed2001

Is preventive resection of the extrahepatic bile duct necessary in cases of pancreaticobiliary maljunction without dilatation of the bile duct?

BACKGROUND: No consensus has been reached on whether preventive resection of the extrahepatic bile duct is necessary in cases of pancreaticobiliary maljunction (PBM) without dilatation of the extrahepatic bile duct (undilated type). METHODS: Sixty-eight patients with PBM underwent corrective surgery and several clinical characteristics and pathological findings including K-ras point mutation were evaluated. RESULTS: Unlike dilated bile duct, none of the patients with undilated type duct had clinical symptoms in early childhood. In patients with either cystic or spindle type duct, amylase levels in the bile duct were >10(4) U/l, whereas those in patients with undilated type duct were <10(4) U/l. Postoperative scintigraphy of the biliary system of undilated type revealed no evidence of cholestasis. After surgery, eight patients with undilated type duct, in whom the bile duct had been preserved, had no further clinical symptoms and no evidence of malignancy. Bile duct tissue specimens revealed no hyperplasia, dysplasia or cancerous lesions and they had no K-ras mutation in undilated type. CONCLUSION: The results showed that there was little bile stasis, injury to the mucosa was mild and less genetic changes could be seen in patients with undilated type duct. Therefore, in patients without dilatation of bile duct and advanced cancer, cholecystectomy alone is sufficient.

Adult↗

PubMed2001

Plasma levels of mature form of adrenomedullin in patients with haemodialysis.

BACKGROUND: Adrenomedullin (AM) is a potent vasodilator and natriuretic peptide with hypotensive effects. Immunoreactive AM in human plasma consists of the biologically active mature form, AM (1-52)-CONH2 (mAM) and the intermediate form, AM-gly-COOH (iAM). However, the different effects of mAM and iAM in patients on haemodialysis (HD) have remained unclear. METHODS: Thirty-nine patients on HD and 10 controls were included in this study. We determined plasma levels of mAM and iAM using an immunoradiometric assay that recognizes total AM (tAM) and another that is specific for only mAM. RESULTS: The plasma concentrations of mAM and iAM in patients before HD were significantly higher than those in the controls (n=10) (4.76+/-0.28 vs 1.28+/-0.22 fmol/ml, P<0.001, 25.99+/-1.47 vs 8.52+/- 0.91 fmol/ml, P<0.001 respectively). The plasma levels of mAM and iAM before HD significantly and negatively correlated with systolic blood pressure (SBP) (r=-0.46, P<0.01, and r=-0.32, P<0.05 respectively) and diastolic blood pressure (DBP) (r=-0.32, P<0.05, and r=-0.35, P<0.05 respectively). After HD, plasma mAM and iAM levels as well as SBP and DBP were significantly lower than before HD. Plasma levels of mAM and iAM correlated significantly (r=0.73, P<0.001). CONCLUSIONS: These data suggest that mAM and/or iAM are involved in blood pressure regulation in patients undergoing HD, and further work is needed to understand the precise role of adrenomedullin in this regulation.

Adrenomedullin↗

PubMed2001

Roles of protein kinase C and Ca2+-dependent signaling in angiotensin II-induced adrenomedullin production in rat cardiac myocytes.

OBJECTIVES: We showed that angiotensin II stimulates adrenomedullin production in cultured neonatal rat cardiac myocytes, and that the secreted adrenomedullin inhibits hypertrophy of the myocytes, although the intracellular mechanisms of adrenomedullin production are still unknown. Since protein kinase C (PKC) and the Ca2+ signaling system are involved in cardiac hypertrophy, we examined the roles of these intracellular signaling systems in the production of adrenomedullin by myocytes. METHODS: Cultured neonatal rat cardiac myocytes were incubated with agonists or antagonists of PKC and Ca2+ signaling systems for 24 h. Adrenomedullin secreted into the medium and adrenomedullin mRNA expression were measured by radioimmunoassay and quantitative polymerase chain reaction, respectively. RESULTS: Both phorbol-12-myristate-13-acetate (PMA), a PKC activator and A23187, a calcium ionophore, significantly increased adrenomedullin mRNA expression and secretion from the myocytes. The induction of adrenomedullin secretion by PMA was abolished by H7, a PKC inhibitor, and by downregulation of PKC induced by pre-incubation with PMA. Similarly, the stimulation of adrenomedullin secretion by 10(-6) mol/l angiotensin II was significantly reduced following the inhibition or downregulation of PKC activity in the myocytes. Blockade of the L-type Ca2+ channel and chelation of intracellular Ca2+ both resulted in a significant reduction of the stimulation of adrenomedullin secretion by angiotensin II. In addition, the secretion was significantly attenuated by inhibitors of calmodulin (W-7) and calmodulin kinase II (KN-62), and slightly attenuated by FK506, a calcineurin inhibitor. CONCLUSIONS: These results suggest that PKC and the Ca2+/calmodulin signaling systems are involved in angiotensin II-induced adrenomedullin secretion from rat cardiac myocytes.

Adrenomedullin↗