Role of cardiac renin-angiotensin system in sarcoplasmic reticulum function and gene expression in the ischemic-reperfused heart

The aim of this study was to explore the possible participation of cardiac renin-angiotensin system (RAS) in the ischemia-reperfusion induced changes in heart function as well as Ca²⁺-handling activities and gene expression of cardiac sarcoplasmic reticulum (SR) proteins. The isolated rat hearts, treated for 10 min without and with 30 M captopril or 100 M losartan, were subjected to 30 min ischemia followed by reperfusion for 60 min and processed for the measurement of SR function and gene expression. Attenuated recovery of the left ventricular developed pressure (LVDP) upon reperfusion of the ischemic heart was accompanied by a marked reduction in SR Ca²⁺-pump ATPase, Ca²⁺-uptake and Ca²⁺-release activities. Northern blot analysis revealed that mRNA levels for SR Ca²⁺-handling proteins such as Ca²⁺-pump ATPase (SERCA2a), ryanodine receptor, calsequestrin and phospholamban were decreased in the ischemia-reperfused heart as compared with the non-ischemic control. Treatment with captopril improved the recovery of LVDP as well as SR Ca²⁺-pump ATPase and Ca²⁺-uptake activities in the postischemic hearts but had no effect on changes in Ca²⁺-release activity due to ischemic-reperfusion. Losartan neither affected the changes in contractile function nor modified alterations in SR Ca²⁺-handling activities. The ischemia-reperfusion induced decrease in mRNA levels for SR Ca²⁺-handling proteins were not affected by treatment with captopril or losartan. The results suggest that the improvement of cardiac function in the ischemic-reperfused heart by captopril is associated with the preservation of SR Ca²⁺-pump activities; however, it is unlikely that this action of captopril is mediated through the modification of cardiac RAS. Furthermore, cardiac RAS does not appear to contribute towards the ischemia-reperfusion induced changes in gene expression for SR Ca²⁺-handling proteins.     

Analysis of a muscular control system in human movements

In this work, we have studied a muscular control system under experimental conditions for analyzing the dynamic behavior of individual muscles and theoretical considerations for elucidating its control strategy. Movement of human limbs is achieved by joint torques and each torque is specified as the sum of torques generated by muscle forces. The behavior of individual muscles is controlled by the neural input which is estimated by means of an electromyogram (EMG). In this study, the EMGs for a flexor and an extensor are measured in elbow joint movements and the dynamic behavior of individual muscles is analyzed. As a result, it is verified that both a flexor and an extensor are activated throughout the entire movement and that the activation of muscles is controlled above a specific limit independent of the hand-held load. Subsequently, a system model for simulating elbow joint movements is developed which includes the muscle dynamic relationship between the neural input and the isometric force. The minimum limit of muscle activation that has been confirmed in experiments is provided as a constraint of the neural input and the criterion is defined by a derivative of the isometric force of individual muscles. The optimal trajectories formulated under these conditions are quantitatively compared with the experimentally observed trajectories, and the control strategy of a muscular control system is studied. Finally, a muscular control system in multi-joint arm movements is discussed with regard to the comparative analysis between observed and optimal trajectories. Received: 7 April 1999 / Accepted in revised form: 27 July 1999     

Phosphorylation-dependent activation of TAK1 mitogen-activated protein kinase kinase kinase by TAB1

TAK1 is a mitogen-activated protein kinase kinase kinase (MAP3K) that is involved in the c-Jun N-terminal kinase/p38 MAPKs and NF-kappaB signaling pathways. Here, we characterized the molecular mechanisms of TAK1 activation by its specific activator TAB1. Autophosphorylation of two threonine residues in the activation loop of TAK1 was necessary for TAK1 activation. Association with TAK1 and induction of TAK1 autophosphorylation required the C-terminal 24 amino acids of TAB1, but full TAK1 activation required additional C-terminal Ser/Thr rich sequences. These results demonstrated that the association between the kinase domain of TAK1 and the C-terminal TAB1 triggered the phosphorylation-dependent TAK1 activation mechanism.     

Genes Involved in Cell Wall Localization and Side Chain Formation of Rhamnose-Glucose Polysaccharide in Streptococcus mutans

We identified in Streptococcus mutans six new genes (rgpA through rgpF), whose disruption results in a loss of serotype-specific antigenicity, specified by the glucose side chains of rhamnose-glucose polysaccharide from the cell wall. Rhamnose and glucose content of the cell wall decreased drastically in all these disruption mutants, except that in the rgpE mutant only the glucose content decreased. RgpC and RgpD are homologous to ATP-binding cassette transporter components and may be involved in polysaccharide export, whereas RgpE may be a transferase of side chain glucose.     

Involvement of protein tyrosine phosphorylation in the effect of green tea polyphenols on Ehrlich ascites tumor cells in vitro

Green tea extract and its polyphenolic components have been found to possess anticarcinogenic, antimutagenic, antihypertensive and antihepatotoxic effects, and several mechanisms have been proposed for these effects. In this study, the effects of five tea polyphenols, (−)-epigallocatechin-3-gallate (EGCG), (−)-epigallocatechin (EGC), (−) epicatechin-3-gallate (ECG), (−) epicatechin (EC) and (+)-catechin (C), were examined on the viability of Ehrlich ascites tumor cells in vitro and a possible relationship with tyrosine phosphorylation was determined. Proteins extracted from the cells treated with the tea polyphenols were separated by SDS-PAGE, and tyrosine-phosphorylated proteins were detected by immunoblotting with anti-phosphotyrosine antibody and the extent of phosphorylation determined. EGC (100 μM) caused a significant decrease in cell viability to 4.1±0.2% of the control value, and this correlated with a stimulation of protein tyrosine kinase (PTK) activity. EGCG (100 μM) also caused a slight decrease in cell viability (70% of the control value) but this and the other polyphenols, which had no effect on cell viability likewise, had no effect on tyrosine phosphorylation. Tyrosine phosphorylations of 42 and 45 kDa proteins were also observed for EGC. Further evaluation of the effect of EGC showed that the activity of ornithine decarboxylase (ODC), a key enzyme in polyamine biosynthesis in cells, decreased significantly as well. A significant correlation has therefore been observed between a cellular event, namely, a reduction in the viability of Ehrlich ascites tumor cells and an association with a tyrosine phosphorylation of 42 and 45 kDa proteins by the polyphenol EGC.     

DIFFERENTIATION OF MITOCHONDRIAL DNA POLYMORPHISMS IN POPULATIONS OF FIVE JAPANESE ABIES SPECIES

Mitochondrial DNA polymorphism of 40 populations of five Abies species was investigated using PCR-amplified coxI and coxIII gene probes. Using four combinations of probe and restriction enzyme, we detected three major haplotypes and 15 total haplotypes. We also found varied levels of gene diversity for the different species: 0.741, 0.604, 0.039, 0.000, and 0.292 for A. firma, A. homolepis, A. veitchii, A. mariesii, and A. sachalinensis, respectively. The marginal and southern populations of A. firma and A. homolepis have unique haplotypes, especially the Kyushu, Shikoku, and Kii Peninsula populations, which inhabit areas coinciding with probable refugia of the last glacial period and possess high levels of mtDNA genetic diversity. The haplotypes in some populations suggested mtDNA capture also occurred between species through introgression/hybridization. The strong mtDNA population differentiation in Abies is most likely due to the maternal inheritance of mitochondria and restricted seed dispersal. A phenetic tree based on the genetic similarity of the mtDNA suggests that some species are polyphyletic. Based on mtDNA variation, the five Abies species could be divided roughly into three groups: (1) A. firma and A. homolepis, (2) A. veitchii and A. sachalinensis, and (3) A. mariesii. However, we found that all these Abies species, except A. mariesii, are genetically very closely related according to an analysis of their cpDNA sequences. This showed that the chloroplast rbcL gene differed by only one base substitutions among the four species. We believe that the mtDNA variation and cpDNA similarity clearly reflect relationships among, and the dissemination processes affecting these Abies species since the last glacial period.     

Characterization of a Maize Ca2+-Dependent Protein Kinase Phosphorylating Phosphoenolpyruvate Carboxylase

Phosphoenolpyruvate carboxylase (PEPC) [EC 4.1.1.31 [EC] ] of plantsundergoes regulatory phosphorylation in response to light ornutritional conditions. However, the nature of protein kinase(s)for this phosphorylation has not yet been fully elucidated.We separated a Ca²⁺-requiring protein kinase from Ca²⁺-independentone, both of which can phosphorylate maize leaf PEPC and characterizedthe former kinase after partial purification. Several linesof evidence indicated that the kinase is one of the characteristicCa²⁺-dependent but calmodulin-independent protein kinase (CDPK).Although the M_r, of native CDPK was estimated to be about 100kDa by gel permeation chromatography, in situ phosphorylationassay of CDPK in a SDS-polyacrylamide gel revealed that thesubunit has an M_r of about 50 kDa suggesting dimer formationor association with other protein(s). Several kinetic parameterswere also obtained using PEPC as a substrate. Although the CDPKshowed an ability of regulatory phosphorylation (Ser-15 in maizePEPC), no significant desensitization to feedback inhibitor,malate, could be observed presumably due to low extent of phosphorylation.The kinase was not specific to PEPC but phosphorylated a varietyof synthetic peptides. The possible physiological role of thiskinase was discussed. ¹Present address: NEOS Central Research Laboratory, 1-1 Ohike-machi,Kosei-cho, Shiga, 520-3213 Japan. ²Present address: Chugai Pharmaceutical Co., Ltd., 1-135 Komakado,Gotemba, 412-0038 Japan. ⁴N.O. and N.Y. contributed equally to this work.     

Cytotoxic cycloartane-type triterpenes from Combretum quadrangulare

Seven novel cycloartane-type triterpenes were isolated from Combretum quadrangulare, and their structures were elucidated on the basis of spectral analysis. All these compounds were tested for their cytotoxicity against murine colon 26-L5 carcinoma cells. Methyl quadrangularate B (2) and methyl quadrangularate D (4) exhibited potent cytotoxicity having ED₅₀ values 9.54 and 5.42 μM, respectively.     

Spicachlorantins A and B,new dimeric sesquiterpenes from the roots of Chloranthus spicatus

Two new lindenane sesquiterpene dimers, spicachlorantins A and B (1 and 2), were isolated from the roots of Chloranthus spicatus along with a known related compound, chloramultilide A (3). Their structures and the absolute stereostructures were established by 1D and 2D NMR as well as by CD spectroscopic analyses.