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Y Tsunoda

Publications and source records attributed to Y Tsunoda.

At least 73 records · Page 4Linked to original sources
PubMed1997

Beneficial effect of carboxy-PTIO on hemodynamic and blood gas changes in septic shock dogs.

BACKGROUND: Nitric oxide (NO) production following bacterial infection may play a physiological role in the host defense mechanism due to its antimicrobial activity. However, excess production of NO in severe infection such as sepsis has been implicated in the pathogenesis of septic shock. To determine whether a nitronyl nitroxide NO scavenger compound could prevent the hemodynamic and blood gas alterations in sepsis, bacterial lipopolysaccharide (LPS: 250ng/kg/min) was administered for 2 h in anesthetized dogs with or without infusion of carboxy-2-phenyl-4, 4, 5, 5-tetramethylimidazoline-1-oxyl-3-oxide (carboxy-PTIO: 0.1 mg/kg/min) for 1 h. Control animals received isotonic saline instead of LPS with or without carboxy-PTIO. RESULTS: Infusion of LPS caused a marked decrease in mean arterial pressure (MAP), metabolic acidosis, and hypoxia. These effects were reversed by co-administration of carboxy-PTIO, without affecting other hemodynamic parameters. In control animals, neither hemodynamic nor blood gas parameters changed with or without carboxy-PTIO. CONCLUSION: These results indicate that carboxy-PTIO attenuates LPS-induced hypotension, metabolic acidosis, and hypoxia by scavenging excess NO from the circulation without affecting NO synthase (NOS) activity. An NO scavenger, carboxy-PTIO, may be preferable to non-selective NOS inhibitors for the treatment of human septic shock.

Journal Article↗
PubMed1997

Adenosine 3',5'-cyclic monophosphate-stimulated Ca++ efflux and acetylcholine release in ileal myenteric plexus are mediated by N-type Ca++ channels: inhibition by the kappa opioid receptor agonist.

Adenosine 3',5'-cyclic monophosphate (cAMP) is an important second messenger involved in cholinergic transmission. The aims of this study were to characterize the calcium channels associated with cyclic AMP-mediated acetylcholine release and Ca++ efflux in ileal myenteric plexus. We also examined if this process can be inhibited by agents such as opioids that inhibit N-type calcium channels via a pertussis toxin-sensitive G protein. Application of a cell permeant analogue, 8-bromoadenosine cyclic AMP (8Br-cAMP) (1 mM), and an activator of the adenylyl cyclase system, forskolin (0.1 mM), in a superfusion system resulted in both Ca++ efflux and 3H-acetylcholine (ACh) release from the dispersed myenteric ganglia. A preferential N-type Ca++ channel blocker, omega-Conotoxin GVIA (omega-CgTx, 10-100 nM), significantly inhibited 3H-ACh release stimulated by 8Br-cAMP. 10 nM omega-CgTx also totally inhibited 8Br-cAMP-induced Ca++ efflux, whereas the L-type Ca++ channel blocker, nifedipine (1 microM), and the T-type Ca++ channel blocker, nickel (100 microM), both had no effects on the action of 8Br-cAMP. 3H-ACh release during 0.1 mM forskolin stimulation was inhibited by pretreatment with a kappa receptor agonist, U50488H at 1 to 100 nM. In addition, U50488H significantly inhibited 3H-Ach release and Ca++ efflux elicited by 8Br-cAMP. Inhibition of 3H-ACh release by U50488H was reversed by 3 hr pretreatment with 300 ng/ml pertussis toxin. These results suggest that, in the myenteric plexus, cyclic AMP-stimulated Ca++ efflux and Ach release were mediated by N-type calcium channels. This process may be inhibited by activation of the kappa opioid receptor through pertussis toxin-sensitive G protein(s).

3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cycloh↗
PubMed1996

Src kinase pathways in extracellular Ca(2+)-dependent pancreatic enzyme secretion.

We investigated the functional roles of Src kinase pathways in rat pancreatic acinar cells. CCK-8 dose-dependently increased Src kinase activities by a mechanism that was sensitive to herbimycin A. CCK-8 enhanced protein tyrosine kinase (PTK) activities when the synthetic Src peptide was used as a substrate. Increased PTK activities, sustained Ca2+ entry, and amylase secretion, all stimulated by CCK-8, were abolished by eliminating extracellular Ca2+ ([Ca2+]o). CCK-8 caused tyrosine phosphorylation of three major proteins: 60 KDa, 85 KDa, and 105 KDa. The elimination of [Ca2+]o prevented the tyrosine phosphorylations of p60 and p105, but not p85. Therefore, the p60 Src kinase and the p105 kinase, referred to as "Src kinase kinase', may be involved in [Ca2+]o-dependent pancreatic exocytosis.

Amylases↗
PubMed1996

Intracellular control of IP3-independent Ca2+ oscillations in pancreatic acini.

In pancreatic acini, the high affinity cholecystokinin (CCK) receptor agonist, CCK-OPE which utilizes the phospholipase A2 (PLA2)/arachidonic acid (AA) pathway, dose-dependently increased intracellular Ca2+ spike frequency and amplitude. An uncoupler of proton gradients, FCCP, abolished Ca2+ oscillations and amylase secretion induced by CCK-OPE. Furthermore, FCCP or decreasing extravesicular pH inhibited ATP-dependent 45Ca2+ uptake into the endoplasmic reticulum (ER) fraction. On the other hand, cytosolic acidification induced by Na(+)-free medium led to Ca2+ oscillations. Depletion of intracellular ATP by antimycin resulted in an abolition of the response to CCK-OPE. Administration of the K+ ionophore, valinomycin, abolished the action of CCK-OPE. Decreasing K+ concentrations outside the ER vesicles inhibited ATP-dependent 45Ca2+ uptake and AA-induced 45Ca2+ release. Caffeine inhibited the actions of CCK-OPE, whereas ryanodine did not have any effects. These data suggest that IP3-independent Ca2+ oscillations mediated by the PLA2 cascades involve the release of intracellular Ca2+ by AA and reuptake of Ca2+ by an ATP-dependent Ca2+/H+ antiport. Furthermore, the presence of K+ membrane potential gradient across the ER membrane is required for normal Ca2+ oscillations.

Adenosine Triphosphate↗
PubMed1996

Low temperature preservation of mouse fetal germ cells at 4 degrees C.

Mouse fetal germ cells(FGCs) isolated or within genital ridges from male fetuses at 15.5 d post coitum were preserved at 4 degrees C for 3-15 d with TCM-199 medium supplemented with 10, 50 or 100% fetal bovine serum (FCS) and 0.1, 0.6 or 1.0 M sucrose. The viability of FGCs was assessed by the dye exclusion test with trypan blue and nuclear transfer into enucleated oocytes and then into enucleated 2-cell embryos. Forty-one to 45 % of FGCs survived for 5 d after preservation in the medium containing 50% FCS and 0.1 M sucrose according to the results of the dye exclusion test. But significant good effect of sucrose was not observed regardless of its concentration. When genital ridges were preserved, 15-22% of FGCs survived even 10 d after preservation. The efficiency of the in vitro development of FGCs to blastocysts was similar when they were fused with enucleated oocytes and 2-cells blastomeres after 2 to 4 d storage(27-33%) compared with that of control FGCs(56%).

Journal Article↗
PubMed1996

Infrequent mutations of the TP53 gene and no amplification of the MDM2 gene in hepatoblastomas.

We have investigated the mutation of the TP53 gene in hepatoblastomas (HBLs) by using polymerase chain reaction-single strand conformation polymorphism and direct sequencing in 38 HBL tumor samples and in two HBL cell lines. We detected the TP53 gene mutation in an anaplastic hepatoblastoma cell line, but no aberration of the TP53 gene (exons 5-9) was found in tumor samples and in the other HBL cell line. The mutation of the cell line was a missense mutation from GAC (asparagine) to CAC (histidine) at codon 281, which was different from the G-to-T transversion of codon 249 that is frequently found in adult hepatocellular carcinomas (HCCs). In addition, we performed Southern blot analysis of the MDM2 gene, but we did not find MDM2 gene amplification in 19 cases tested. Our results suggest that, in contrast to the findings in HCCs in adults, TP53 gene aberrations are not involved in the development or progression of HBLs in children.

Adult↗
PubMed1996

Impaired intracellular signal transduction in gastric smooth muscle of diabetic BB/W rats.

The pathophysiological mechanisms responsible for diabetic gastroparesis remain unclear. Diabetes mellitus occurs spontaneously in 90% of a partially inbred colony of BB/W rats. This animal model resembles human insulin-dependent diabetes and is suitable for investigating the mechanism of diabetic gastroparesis. Diabetic BB/W rats were killed 6 mo after the onset of diabetes. Muscle contraction experiments and [3H]acetylcholine release studies were performed with muscle strips of the gastric body. Biochemical measurements of inositol trisphosphate (IP3) and protein kinase C (PKC) in gastric muscle were performed to characterize abnormalities of the intracellular signal transduction system in gastric myocytes. Circular muscle contractions in response to direct myogenic stimulants, carbachol (10(-7) - 10 (-3)M) or substance P (10(-7) - 10(-5)M), were significantly impaired in diabetic BB/W rats compared with controls. Similarly, muscle contractions in response to NaF (10 mM), a direct stimulant of G proteins, were also impaired in diabetic BB/W rats. In contrast, muscle contractions in response to KCl (25-75 mM) were similar between control and diabetic BB/W rats, indicating normal voltage-dependent Ca2+ entry in muscle strips obtained from diabetics BB/W rats. [3H]acetylcholine release from gastric myenteric plexus in response to electrical transmural stimulation remained intact in diabetic BB/W rats. In separate studies, we demonstrated that carbachol (10(-6) - 10(-4)M) -induced IP3 responses were significantly reduced in diabetic rats compared with control. In addition, there was also impairment of translocation of PKC in diabetic BB/W rats. These observations indicate that myogenic impairment occurred in diabetic BB/W rats. This resulted from altered intracellular signal transduction involving abnormal IP3 production and PKC translocation.

Acetylcholine↗
PubMed1996

Structural requirements of CCK analogues to differentiate second messengers and pancreatic secretion.

We previously demonstrated that, in rat pancreatic acinar cells, the high-affinity cholecystokinin (CCK) receptor agonist JMV-180 utilizes the phospholipase A2 (PLA2) cascade to mediate Ca2+ oscillations and amylase secretion. In contrast, the low-affinity CCK receptor utilizes the phospholipase C beta 1 (PLC beta 1) pathway. We have investigated structural requirements of CCK analogues to activate different intracellular pathways. CCK analogues such as CCK-8 [Met28,31; half-maximal effective concentration (EC50) = 0.4 pM], CCK-7 (Met28,31; EC50 = 0.7 pM), and NONA (Thr28/Nle31; EC50 = 5 pM) caused a biphasic amylase secretion. Reduction of secretion occurred with high doses of these peptides (> 100 pM). In contrast, CCK-5 (Met31; EC50 = 20,000pM), JMV-180 (Nle28,31; 1,500 pM), and OPE (Nle28,31; 200 pM) caused a monophasic secretion. CCK-8, but not JMV-180, increased protein kinase C (PKC) activities. The PKC activator phorbol ester inhibited an increase in myo-inositol 1,4,5-trisphosphate levels induced by CCK-8 and abolished monophasic amylase secretion induced by OPE. CCK-8, CCK-7, and NONA caused Ca2+ oscillations (< 100 pM) or large Ca2+ transients (> 100 pM). In contrast, JMV-180 and OPE evoked Ca2+ oscillations, even in high doses. Ca(2+)-signaling modes induced by CCK-5 were intermediate types between CCK-8 and JMV-180. CCK-8- and CCK-7-stimulated Ca2+ spikes were inhibited by the PLC inhibitor U-73122, but not by the PLA2 inhibitor ONO-RS-082. The action of CCK-5 was only partially sensitive to the PLC inhibitor. In contrast, JMV-180- and OPE-stimulated Ca2+ oscillations were inhibited by the PLA2, but not by the PLC, inhibitor. NONA was sensitive to PLC and PLA2 inhibitors. Although JMV-180 differs from CCK-8 by having an Asp-2 phenylethylester, rather than an Asp-phenylalanine amide, it is unlikely that these differences in the carboxyl terminus are important in determining which second-messenger systems will be activated. This is because CCK-5 (Phe33-CONH2) causes monophasic amylase secretion and Ca2+ oscillation in a manner similar to those induced by JMV-180 (2-phenylethylester). Meanwhile NONA (Phe33-CONH2) appeared to activate PLC and PLA2 pathways. The actions of all CCK analogues were abolished by L-364,718, indicating mediation by CCK-A receptors. Therefore, depending on the agonists used, CCK-A receptor activation in pancreatic acini may result in differential involvement of second-messenger systems, Ca2+ signal transduction, and amylase secretion. On the basis of the amino acid sequence of the carboxy terminus of CCK analogues, it appears that key amino acids for this differentiation are Met28 (or Thr28) for PLC pathways and Nle28 for PLA2 pathways.

Amino Acid Sequence↗
PubMed1996

Increased circulating adrenomedullin, a novel vasodilatory peptide, in sepsis.

Human adrenomedullin (hAM), a potent vasodilatory peptide originally identified in pheochromocytoma, has been shown to be present in various human tissues and circulate in human plasma. We measured plasma concentrations of immunoreactive hAM in patients with sepsis who had been admitted to intensive care unit (ICU). Plasma hAM concentrations in 12 septic patients upon entering the ICU were extremely elevated (107 +/- 139 fmol/ml: mean +/- SD) compared to those of 16 age-matched normal subjects (7.9 +/- 3 fmol/mL). Among 10 patients with normal renal function, plasma hAM levels either decreased or increased during the hospital course; the former group survived and the latter group succumbed. Two patients with acute renal failure had markedly elevated plasma hAM levels during the early course, which declined rapidly during the recovery course. High performance liquid chromatography of plasma extracts from one patient with acute renal failure revealed a single major component of immunoreactive hAM coeluting with authentic hAM (1-52) during acute and recovery phase. Plasma hAM concentration showed positive correlations with heart rate, right atrial pressure, and serum creatinine concentration, but not with other hemodynamic variables. These data suggest that a marked increase in circulating hAM in sepsis may be caused by its decreased clearance and/or its enhanced synthesis by multiple organ dysfunction, and that increased endogenous hAM may be involved in the mechanism of cardiovascular abnormalities associated with sepsis.

Adrenomedullin↗
PubMed1996

[Dual biochemical modulation therapy using 5-FU, leucovorin and cisplatin on human rectal carcinoma xenografts in nude mouse].

This study examined a combined treatment for colorectal carcinoma, the dual biochemical modulation therapy, consisting of 5-FU, Leucovorin (LV) and Cisplatin (CDDP). We compared its anti-tumor effects with other treatments: 5-FU alone, CDDP alone and 5-FU with LV. Primary diffuse infiltrated colorectal carcinoma is well known for its biological malignancy and its lack of response to chemotherapy. We used SRM cells from a cell line of carcinoma of the rectum, and subcutaneously injected them into nude mice. The anti-tumor effects were estimated from the growth rate, inhibition rate and thymidylate synthetase inhibition rates in the tumor tissue. Results indicated that even if the concentration of 5-FU and LV were reduced by half, these combined with CDDP were more effective than other therapies. Dual biochemical modulation therapy is particularly promising because the reduction of the dosages would reduce the side effects while still serving as an excellent anti-tumor therapy.

Animals↗
PubMed1995

The regulatory site of functional GTP binding protein coupled to the high affinity cholecystokinin receptor and phospholipase A2 pathway is on the G beta subunit of Gq protein in pancreatic acini.

A non-hydrolysable guanosine nucleotide analog, GTP[S] at 200 microM, stimulated amylase secretion which was inhibited by an anti-phospholipase A2 (PLA2) antibody in permeabilized pancreatic acini, indicating that the PLA2 pathway is linked to the GTP binding protein. A high affinity cholecystokinin (CCK) receptor agonist, CCK-OPE (10 microM), and a low affinity receptor agonist, CCK-8 (0.1 microM), both caused amylase secretion in permeabilized cells. The action of CCK-OPE was abolished by the G beta antibody but not by the G alpha-q,11 antibody, whereas the opposite was true of the CCK-8 response. Biscoclaurine alkaloid isotetrandrine (10 microM), a specific inhibitor of PLA2-coupled G proteins, abolished Ca2+ oscillations and amylase secretion induced by CCK-OPE (0.1-100 nM), but not by CCK-8 (10 pM) in intact acini. Gp antagonist-2A (10 microM), which inhibits the activation of Gq, also inhibited the actions of CCK-OPE (10 pM-1 microM) in intact acini. These observations indicate that the functional unit of the heterotrimeric G protein coupled to the high affinity CCK receptor appears to be different from that linked to the low affinity CCK receptor/Gq-alpha pathway. The regulatory site of this G protein coupled to the high affinity CCK receptor is on the beta subunit of Gq protein which elicits Ca2+ oscillations and monophasic amylase secretion via the PLA2 pathway.

Alkaloids↗
PubMed1995

Acute renal failure associated with Candida albicans infection.

A 16-year-old male with a long history of steroid-responsive nephrotic syndrome developed fever, abdominal pain, thrombocytopenia and acute renal failure. The clinical course and renal histology were similar to, but not typical of, haemolytic uremic syndrome. Positive cultures (throat, oesophagus, stool), an elevation in serum levels of specific antibody and fungal polysaccharide (1,3) beta-D-glucan and response to the antifungal therapy indicated an association between this syndrome and infection with Candida albicans.

Acute Kidney Injury↗
PubMed1995

Macrophages in childhood IgA nephropathy.

The role of glomerular macrophages in IgA nephropathy in children was investigated using a new monoclonal antibody (KP1) as a probe. The average number of glomerular macrophages per patient (ANM/P) was closely correlated with the degree of hematuria (P < 0.01) as well as with the degree of leukocyturia (P < 0.01) in the absence of any correlation with proteinuria, serum IgA levels or the interval between the detection of urine abnormalities and renal biopsy. ANM/P was significantly higher in patients diagnosed pathologically as having focal and diffuse proliferative glomerulonephritis than in patients with minor glomerular abnormalities or advanced sclerosis (P < 0.05). Among various types of glomerular morphology in individual patients, macrophages predominantly infiltrated glomeruli with cell-proliferative lesions despite an absence of any increase in glomeruli with minor abnormalities or with sclerosis. Macrophages were mainly localized within the capillary lumen in association with endocapillary proliferative lesions (tuft necrosis), they accumulated in areas of mesangial proliferation, and they were attached to Bowman's capsule in segmental lesions. Macrophages were less evident in sclerosis. Furthermore, ultrastructural analysis revealed macrophages in the paramesangial areas in close proximity to lytic changes in the glomerular basement membrane and effacement of epithelial foot processes. In addition, some cases in repeat biopsy shows prolonged or increased values of ANM/P after several years of interval in association with progression of proliferative lesions. These results suggest that macrophages infiltrate glomeruli during acute glomerular inflammation, and that they are involved in mesangial proliferation or the development of extracapillary lesions in the absence of apparent clinical symptoms. Furthermore, recurrence or prolonged infiltration may promote progression of IgA nephropathy.

Antibodies, Monoclonal↗
PubMed1995

High-affinity CCK receptors are coupled to phospholipase A2 pathways to mediate pancreatic amylase secretion.

It is well recognized that JMV-180, a cholecystokinin (CCK) analogue, acts as an agonist on the high-affinity CCK receptor in pancreatic acinar cells. It caused Ca2+ oscillations and amylase secretion in a manner independent of the phospholipase C-inositol 1,4,5-trisphosphate (IP3) pathway. We investigated the mechanism by which the high-affinity CCK receptor utilizes IP3-independent Ca2+ signal transduction to mediate amylase secretion. JMV-180 (1-1,000 nM)-stimulated Ca2+ oscillations and amylase secretion were significantly inhibited by the phospholipase A2 (PLA2) inhibitor, ONO-RS-082 (10 microM). Using streptolysin O-permeabilized cells, we showed that a porcine pancreatic anti-PLA2 antibody from rabbit serum (250 ng/ml) inhibited JMV-180-stimulated amylase secretion. In contrast to CCK octapeptide, JMV-180 (1 nM-10 microM) had no effect on intracellular IP3 levels. These concentrations of JMV-180 did, however, increase intracellular levels of arachidonic acid (AA) metabolite by 2.5-fold in a biphasic manner. Application of exogenous AA (10 microM) released 60% of ATP-incorporated 45Ca2+ from permeabilized pancreatic acini within 3 min in a transient manner. We also showed that active phorbol ester (100 nM) inhibited Ca2+ oscillations and amylase secretion stimulated by JMV-180 (10 nM) or CCK-OPE (100 nM). Application of Mn2+ (2 mM) to superfused acini resulted in a rapid quench of fura 2 fluorescence during 10 nM JMV-180 stimulation, suggesting an involvement of extracellular Ca2+ influx. However, the major source of Ca2+ utilized for oscillations during high-affinity CCK receptor activation was intracellular. In conclusion, we have demonstrated that the high-affinity CCK receptors are coupled to PLA2 pathways to produce AA, which mediates cytosolic Ca2+ oscillation and monophasic amylase secretion, in rat pancreatic acinar cells.

Amylases↗
PubMed1995

Effects of nitric oxide synthase inhibitor on hemodynamic change and O2 delivery in septic dogs.

To elucidate the role of nitric oxide (NO) in septic shock, we measured hemodynamic and pulmonary gas changes in anesthetized dogs after intravenous administration of bacterial lipopolysaccharide (LPS) with or without NO synthase inhibitor, NG-nitro-L-arginine (L-NNA). Infusion of LPS (250 ng.kg-1.min-1) for 2 h decreased mean arterial pressure over 1-4 h. Although L-NNA (10 mg/kg) blocked LPS-induced hypotension, it decreased cardiac index, oxygen delivery index, arterial pH, and arterial PO2 and increased systemic vascular resistance index in the presence or absence of LPS. Administration of NG-nitro-D-arginine (D-NNA, 10 mg/kg) alone caused fewer hemodynamic effects (increased systemic vascular resistance index and decreased cardiac index) than L-NNA alone. Our study provides evidence that L-NNA prevents endotoxin-induced hypotension but decreases cardiac output and oxygen delivery, effects that may, in part, be due to a nonspecific NO synthase-independent event. Thus clinical use of NO synthase inhibitors for the treatment of septic shock should be cautiously considered.

Amino Acid Oxidoreductases↗
PubMed1995

Germ cell nuclei of male fetal mice can support development of chimeras to midgestation following serial transplantation.

Chimeric embryos between fertilized eggs from F1 (C57BL x CBA) and 15.5-16.5 days post coitum (dpc) male fetal germ cells (FGCs) from CD-1 strain (glucose phosphate isomerase, Gpi-1a/a) mice were produced by nuclear transfer. Briefly, a single FGC was fused with enucleated oocytes and activated, and the reconstituted oocytes were cultured to the 2-cell stage. The nucleus from the reconstituted 2-cell embryos was then transferred into an enucleated blastomere of the same stage embryos derived from F1 mice to produce chimeric embryos. The reconstituted 2-cell embryos, which synchronously divided to the 4-cell stage after treatment with nocodazole, were further cultured in vitro. Compacted morula and blastocysts were transferred to the uteri of pseudopregnant female mice. Some recipients were allowed to develop to term and the others were killed at mid gestation to analyze the contribution of donor FGC-derived cells. Survival to term was low with no chimeric animals. Glucose phosphate isomerase (GPI) analysis at midgestation revealed that some conceptuses had chimerism in the fetuses, trophoblast and yolk sac at day 10.5 of pregnancy. The contribution of donor cells was 37-47%, 19-65% and 12-63%, respectively. It was concluded that the nucleus from 15.5-16.5 dpc male fetal germ cells had the potency to develop into fetus, trophoblast and yolk sac after serial nuclear transfer with oocytes and fertilized embryos. The reason for the low viability of chimeric embryos is discussed.

Animals↗