Biomedical subjects

S Shibata

Publications and source records attributed to S Shibata.

At least 91 records · Page 5Linked to original sources

PubMed2000

PCCX1, a novel DNA-binding protein with PHD finger and CXXC domain, is regulated by proteolysis.

We identified a novel gene PCCX1 that encoded a nuclear protein carrying a PHD finger, a CXXC domain, and an acidic region. The CXXC domain was found to be sufficient for binding to DNA. The acidic region exhibited a high transactivation ability, but the full-length protein was inactive due to regions which inhibited the acidic region, including the C-terminal region. We examined the expression of PCCX1 during cellular aging and immortalization of SV40-transformed human fibroblasts. PCCX1 mRNA was expressed constitutively through stages of cellular aging and immortalization, but at the protein level, a shorter form lacking the C-terminal region appeared as the cells approached crisis. These results suggested that PCCX1 was activated by proteolytic cleavage, which removed the C-terminal inhibitory region.

Amino Acid Sequence↗

PubMed2000

Involvement of glial fibrillary acidic protein (GFAP) expressed in astroglial cells in circadian rhythm under constant lighting conditions in mice.

To clarify the role of glial fibrillary acidic protein (GFAP)-expressed glial cells in the circadian clock, we examined GFAP expression in the suprachiasmatic nucleus (SCN) and the intergeniculate leaflet (IGL) under various lighting conditions in mice. We demonstrated that GFAP expression did not show daily change in the SCN under a light-dark cycle; however, long-term housing under constant lighting conditions led to dramatic changes in GFAP expression, i.e., a decrease in the SCN and an increase in the IGL. Furthermore, mice that had a targeted deletion in the GFAP gene (GFAP mutant mice) showed longer and more arrhythmic circadian activity rhythms in constant lighting conditions than wild-type mice, while GFAP mutant mice exhibited stable circadian rhythms both in a light-dark cycle and constant darkness, and showed normal entrainment to environmental light stimuli. These results suggest that the GFAP-expressed astroglial cells in the SCN and the IGL may have some role in circadian oscillation under constant lighting conditions.

Animals↗

PubMed2000

PLC beta 4-independent Ca2+ rise via muscarinic receptors in the mouse suprachiasmatic nucleus.

Cholinergic regulation of the suprachiasmatic nucleus (SCN) has been extensively studied although the intracellular signaling mechanisms are not well understood. We examined immunostaining for phospholipase C-beta (PLC-beta) families that couple to muscarinic acetylcholine receptors (mAChR) and demonstrated the expression of PLC-beta 1 and beta 4 in the mouse SCN. Ca2+ imaging analysis indicated that the MI-mAChR antagonist, pirenzepine blocked carbachol-induced Ca2+ elevation in the SCN and the response was equivalent between the wild type and the PLC-beta 4-knockout mice. In addition, the knockout mice displayed locomotor and temperature rhythms coupling to 24 h light/dark cycles. Therefore, it was proposed that PLC-beta 1 but not PLC-beta 4 was involved in the mAChR-mediated Ca2+ signaling in the SCN.

Animals↗

PubMed2000

Properties of the delayed rectifier potassium current in porcine sino-atrial node cells.

Whole-cell currents were recorded in single, spontaneously active cells dissociated from porcine sino-atrial node, and the conductance and gating properties of the delayed rectifier K+ current (IK) were investigated. The isolated cells exhibited spontaneous action potentials at a rate of 80.5 +/- 5.4 min-1 (mean +/- s.e.m., n = 11). Under Ca2+ current block, depolarization from -40 mV to various potentials activated a time-dependent outward current (IK). The activation curve of IK showed a half-activation potential (V½) of 20.5 +/- 2.1 mV and a slope factor (S) of 16.4 +/- 1.2 mV (n = 8). As the duration of the depolarizing pulse to either +10 or +60 mV was prolonged, the amplitude of the tail current increased in proportion to that of the activated outward current during depolarization. E4031 (2-5 µM), a selective blocker for the rapidly activating component of IK (IK,r), hardly affected IK, but chromanol 293B, a selective blocker for the slowly activating component (IK,s), inhibited IK with an IC50 of 8.79 µM. The reversal potential of IK was -75.2 +/- 2.3 mV with 5.4 mM external and 150 mM internal K+. The time courses of activation and deactivation of IK were fitted by the sum of two exponential functions at various potentials. The relationship between the time constants and membrane potential showed a bell-shaped curve with a peak at around -10 mV for both fast and slow components. The results indicate that in porcine sino-atrial node cells IK is largely derived from IK,s and that IK,s plays a functional role in the slow diastolic depolarization. IK,s may, in part, account for the relatively slower heart rate of pigs than that of rabbit in which IK,r is a functionally dominant component of IK.

Journal Article↗

PubMed2000

Properties of the delayed rectifier potassium current in porcine sino-atrial node cells.

1. Whole-cell currents were recorded in single, spontaneously active cells dissociated from porcine sino-atrial node, and the conductance and gating properties of the delayed rectifier K+ current (I(K)) were investigated. 2. The isolated cells exhibited spontaneous action potentials at a rate of 80.5 +/- 5.4 min(-1) (mean +/- S.E.M., n = 11). Under Ca2+ current block, (depolarization from -40 mV to various potentials activated a time-dependent outward current (I(K)). The activation curve of I(K) showed a half-activation potential (V1/2) of 20.5 +/- 2.1 mV and a slope factor (S) of 16.4 +/- 1.2 mV (n = 8). 3. As the duration of the depolarizing pulse to either +10 or +60 mV was prolonged, the amplitude of the tail current increased in proportion to that of the activated outward current during depolarization. 4. E4031 (2-5 microM), a selective blocker for the rapidly activating component of I(K) (I(K,r)), hardly affected I(K), but chromanol 293B, a selective blocker for the slowly activating component (I(K,s)), inhibited I(K) with an IC(50) of 8.79 microM. 5. The reversal potential of I(K) was -75.2 +/- 2.3 mV with 5.4 mM external and 150 mM internal K+. The time courses of activation and deactivation of I(K) were fitted by the sum of two exponential functions at various potentials. The relationship between the time constants and membrane potential showed a bell-shaped curve with a peak at around -10 mV for both fast and slow components. 6. The results indicate that in porcine sino-atrial node cells I(K) is largely derived from I(K,s) and that I(K,s) plays a functional role in in the slow diastolic depolarization. I(K,s) may, in part, account for the relatively slower heart rate of pigs than that of rabbit in which I(K,r) is a functionally dominant component of I(K).

Action Potentials↗

PubMed2000

Mutant mice lacking ryanodine receptor type 3 exhibit deficits of contextual fear conditioning and activation of calcium/calmodulin-dependent protein kinase II in the hippocampus.

As it is known that ryanodine receptor type 3 is expressed in the hippocampus, we examined the contribution of this receptor to contextual fear conditioning behavior and to the activation of Ca(2+)/calmodulin-dependent protein kinase II using mice lacking the receptor. Ryanodine receptor type 3-deficient mice exhibited impairments of performance in the contextual fear conditioning test, passive avoidance test, and Y-maze learning test. Both the activities of Ca(2+)/calmodulin-dependent protein kinase IIbeta and Ca(2+)/calmodulin-dependent protein kinase IIalpha were significantly increased in the experimental group compared to the control group in the hippocampus, but not in the cingulate cortex on the testing day 24 h after contextual fear training. However, the activities of Ca(2+)/calmodulin-dependent protein kinase IIbeta and alpha were almost the same in the experimental and control groups in the hippocampus on the training day. Ryanodine receptor type 3-deficient mice did not show the increment of Ca(2+)/calmodulin-dependent protein kinase IIbeta and alpha activities in the hippocampus on the testing day. In addition, these mutant mice showed the reduction of fear response in the elevated plus-maze test. The present results suggest that calcium-induced calcium release through the activation of ryanodine receptor type 3 in the hippocampus is important to the expression of the performance of contextual learning through the elevation of Ca(2+)/calmodulin-dependent protein kinase IIbeta and alpha activities.

Animals↗

PubMed2000

Identification of lysophospholipid receptors in human platelets: the relation of two agonists, lysophosphatidic acid and sphingosine 1-phosphate.

Lysophosphatidic acid (LPA) and sphingosine 1-phosphate (Sph-1-P) are known as structurally related bio-active lipids activating platelets through their respective receptors. Although the receptors for LPA and Sph-1-P have been recently identified in various cells, the identification and characterization of ones in platelets have been reported only preliminarily. In this report, we first investigated the distinct modes of LPA and Sph-1-P actions in platelet activation and found that LPA functioned as a much stronger agonist than Sph-1-P, and high concentrations of Sph-1-P specifically desensitized LPA-induced intracellular Ca(2+) mobilization. In order to identify the responsible receptors underlying these observations, we analyzed the LPA and Sph-1-P receptors which might be expressed in human platelets, by RT-PCR. We found for the first time that Edg2, 4, 6 and 7 mRNA are expressed in human platelets.

Blood Platelets↗

PubMed2000

Phase II trial of combination intraperitoneal cisplatin and 5-fluorouracil in previously treated patients with advanced ovarian cancer: long-term follow-up.

OBJECTIVES: This trial was performed to determine the response rate and progression-free and overall survivals of patients with advanced recurrent ovarian cancer who were treated with intraperitoneal cisplatin and 5-fluorouracil. METHODS: Twenty-four patients with ovarian cancer were entered on this trial and treated with intraperitoneal (ip) cisplatin (DDP) and ip 5-fluorouracil, every 3 weeks for eight cycles. Following iv hydration, the cisplatin and 5-fluorouracil were administered through an ip catheter in 2 liters of 0.9% normal saline with a 4-h dwell. RESULTS: All patients were evaluable for progression-free and overall survival and toxicity analysis, and 22 patients for response. The median age was 59 (range, 35-71); initial disease status included 9 patients with residual disease following chemotherapy prior to entry on this study; 5 patients had progressed, and 10 patients had recurrent disease more than 6 months following initial chemotherapy. Of the 9 patients with residual disease, 1 complete response and 3 partial responses were observed; of 10 patients with recurrent disease, 1 complete and 1 partial response were observed for an overall response rate of 27%. No objective responses were seen in the 7 patients who were platinum-refractory on protocol entry. The median progression-free and overall survivals are 7.0 (range, 0.5-137) and 15.5 (range, 3-147) months, respectively. Toxicity included hypomagnesemia, vomiting, abdominal pain, and mild anemia. Only one patient required a dosage adjustment of cisplatin for a serum creatinine elevation >2.0 mg/dl. CONCLUSIONS: We conclude that the combination of ip cisplatin and 5-FU is an effective regimen for patients with residual or relapsed epithelial ovarian cancer with survival durations, response rates, and toxicity profiles that compare favorably with those of other second-line ovarian cancer regimens. Patients who are primarily platinum-refractory are unlikely to benefit from these agents administered into the peritoneal cavity.

Adult↗

PubMed2000

Nondestructive and real-time evaluation of liver viability in brain dead donor for liver transplantation using near-infrared spectroscopy.

A reliable and less-invasive method is currently desired to assess the hemodynamic and functional alteration associated with brain death in the organs of donor candidates. Near-infrared spectroscopy (NIRs) was applied to rat liver in brain-dead donors for assessing tissue oxygenation and intracellular energy metabolism as a means of monitoring the liver viability in the brain-dead donor. Brain-dead rats were divided into 4 according to doses of epinephrine and vasopressin administered. Arterial ketone bodies ratio (AKBR), hyaluronic acid (HA), and NIRs monitoring of a liver graft were performed in the brain-dead phase before the grafts were transplanted into syngeneic rats. NIRs monitoring of oxygenated hemoglobin (Hb) and cytochrome aa3 oxidase (Cytaa3) redox state reflected changes in the hepatic microcirculation and intracellular oxygenation. The administration of high-dose epinephrine proved to be contraindicated due to catecholamine-induced hypoxic stress, while combined administration of adrenaline and vasopressin at an optimal dose rate was beneficial for preservation of the liver viability. The data obtained by NIRs were significantly correlated with the 7-day survival of recipients after liver transplantation. Thus, we conclude that NIRs is a sensitive and nondestructive method for monitoring alterations in the viability of brain-dead liver and can predict liver graft outcome.

Animals↗

PubMed2000

Efficacy of HSP72 induction in rat liver by orally administered geranylgeranylacetone.

It is well known that heat-shock proteins (HSPs) have a cytoprotective function as "molecular chaperones" when cells are exposed to several stress conditions. Geranylgeranylacetone (GGA) is an antiulcer drug that was developed in Japan and it has recently been reported to induce HSP72 in rat gastric mucosa. In this experiment, we investigated the induction of HSP72 in rat liver in response to oral administration of GGA and assessed its ability to induce tolerance to warm ischemic injury by this approach. We prepared donor rats by orally administering GGA to them and compared HSP72 expression in graft liver, survival rates, and serum TNF-alpha concentrations after liver transplantation with the findings in controls. The survival rates were significantly increased when the livers were obtained from donor rats given GGA. Western blotting revealed expression of HSP72 in graft livers given GGA, and the serum TNF-alpha levels were significantly suppressed in the rats given GGA. Oral administration of GGA induced HSP72 in graft livers, and they were better able to tolerate warm ischemic injury. Oral administration of GGA appears to provide a promising new strategy for preventing ischemia-reperfusion injury.

Administration, Oral↗

PubMed2000

Phase I study of AG2034, a targeted GARFT inhibitor, administered once every 3 weeks.

PURPOSE: To identify a recommended phase II dose for the second generation glycinamide ribonucleotide transformylase (GARFT) inhibitor, AG2034, administered by intravenous bolus every 3 weeks without folate supplementation and to describe AG2034 pharmacokinetics. METHODS: Adults with advanced malignancies were enrolled in cohorts of three per dose level with expansion to six upon observation of dose-limiting toxicity (DLT). The maximum tolerated dose (MTD) was defined as the dose at which two of up to six patients experienced DLT. Upon identification of an MTD and evidence of cumulative toxicity, a lower intermediate dose was explored as a candidate phase II dose. AG2034 plasma concentrations were measured using an ELISA assay. RESULTS AND CONCLUSIONS: The recommended phase II dose is 5.0 mg/m2. DLTs were anemia, thrombocytopenia, mucositis, diarrhea, hyperbilirubinemia, fatigue, and insomnia. Toxicities were modestly cumulative over three courses. Pharmacokinetic analysis showed a dose-AUC0-24 relationship and a progressive increase in AG2034 AUC0-24 over three courses. Both pharmacokinetic and pharmacodynamic factors may contribute to the modest cumulative toxicity observed with AG2034.

Adult↗

PubMed2000

Mandibular deformities in parathyroid hormone-related protein (PTHrP) deficient mice: possible involvement of masseter muscle.

Previous studies using parathyroid hormone-related protein (PTHrP) null mutant mice have indicated severe abnormalities in the endochondral ossification, suggesting that PTHrP affects chondrocyte differentiation. In this study, we found in newborn PTHrP-deficient mice some deformities in the mandible that is formed via intramembranous ossification. The mandibular ramus was bent downwards and a prominent bone crest to which the deep layer of masseter muscle was tendinously attached was observed in the mandibular body. Transmission electron microscopic studies showed that active bone formation was progressing along the tendon fibers of the masseter muscle. The examination of 3-D reconstruction models indicated that the mandibular ramus was bent at the site of muscle attachment, which was shifted in the direction of the muscle fibers. Muscle fiber type analysis using myosin ATPase staining showed that the masseter muscle in the newborn PTHrP-deficient mice contained numerous type 2B fibers, demonstrating premature maturation of this muscle. Based on these findings, we speculated that premature maturation of the masseter muscle leads, probably due to increased tensile forces, to accelerated bone crest formation and subsequent bending of the mandibular ramus. These results further suggest that PTHrP is involved in the regulation of muscle development in normal animals.

Animals↗

PubMed2000

Isolation of proteoglycan (versican) aggregate from rat dental pulp.

Versican is a large interstitial proteoglycan that is believed to be able to bind hyaluronan to form large aggregate structures, but no study has isolated native versican aggregates from any tissue. In this study, ternary aggregate structures consisting of versican, hyaluronan, and link protein were isolated from rat dental pulp by associative extractions followed by caesium sulphate rate zonal sedimentation centrifugation. Fractions from the centrifugation were analysed by dot blot and Western blot using monoclonal antibodies and hyaluronan-binding protein. About 60% of the hexuronic acid was extracted by associative extractions. Positive reactions for versican, hyaluronan and link protein were clearly detected in the bottom fractions from the centrifugation, but were barely detectable in the top fractions. These results suggest that the majority of the versican, hyaluronan, and link protein forms ternary aggregate structures in the rat dental pulp.

Animals↗

PubMed2000