Discrimination of ATP,ADP, and AMPPNP by chaperonin GroEL: hexokinase treatment revealed the exclusive role of ATP

The double ring chaperonin GroEL binds unfolded protein, ATP, and GroES to the same ring, generating the cis ternary complex in which folding occurs within the cavity capped by GroES (cis folding). The functional role of ATP, however, remains unclear since several reports have indicated that ADP and AMPPNP (5'-adenylyl-beta,gamma-imidodiphosphate) are also able to support the formation of the cis ternary complex and the cis folding. To minimize the effect of contaminated ATP and adenylate kinase, we have included hexokinase plus glucose in the reaction mixtures and obtained new results. In ADP and AMPPNP, GroES bound quickly to GroEL but bound very slowly to the GroEL loaded with unfolded rhodanese or malate dehydrogenase. ADP was unable to support the formation of cis ternary complex and cis folding. AMPPNP supported cis folding of malate dehydrogenase to some extent but not cis folding of rhodanese. In the absence of hexokinase, apparent cis folding of rhodanese and malate dehydrogenase was observed in ADP and AMPPNP. Thus, the exclusive role of ATP in generation of the cis ternary complex is now evident.     

Numerical simulation for electrochemical cultivation of iron oxidizing bacteria

A numerical simulation model was constructed for electrochemical cultivation of iron oxidizing bacterium, Thiobacillus ferrooxidans, based on Monod's dual limitation equation. In this model, two limiting factors were examined, low supply of Fe(II) ion and dissolved oxygen, from empirical viewpoints. The simulation model was constructed taking into consideration the energy balance based on the amount of the electronic flow from the electrode to bacteria via an iron ion, and then to oxygen. The model consisted of a logarithmic bacterial growth phase during the first three days, followed by a plateau and growth limitation thereafter. The predicted results were in agreement with the actual growth under electrochemical cultivation. It was predicted the growth limiting factor would be changed from insufficient supply of Fe(II) ions to that of oxygen by decreasing the value of oxygen transfer constant K, which correlated with the aeration rate. The optimum aeration rate was determined for the ideal electrochemical cultivation. The algorithm described here can be used in any electrochemical cultivation by modifying the parameters for each system.     

Role of bradykinin in acid-induced mesenteric hyperemia and duodenal villous damage

Intestinal mucosal capsaicin-sensitive afferent nerves mediate, in part, the mesenteric hyperemia after intraduodenal acidification. The hyperemia plays a role in protecting the duodenal mucosa against acid damage. We tested the hypothesis that bradykinin contributes to this protective hyperemia. A specific antagonist of bradykinin will attenuate the hyperemia and exacerbate duodenal villous damage induced by acid. Study 1: Intravenous vehicle, or the specific bradykinin B2 receptor antagonist (HOE 140) was administered to anesthetized rats. This was followed by intraduodenal bolus administration of 160 microM capsaicin or 0.1 N HCl, and then intravenous bradykinin. Study 2: Intravenous administration of vehicle or HOE 140 was followed by duodenal perfusion with 0.1 N HCl. Superior mesenteric artery blood flow (pulsed Doppler flowmetry) (Study 1) and duodenal villous damage (histology) (Study 2) were recorded. HOE 140 significantly reduced the hyperemia induced by bradykinin and intraduodenal capsaicin or acid. Deep villous injury was significantly increased after treatment with HOE 140. These findings support the hypothesis that acid-induced and afferent nerve-mediated mesenteric hyperemia is modulated by a mechanism that involves bradykinin B2 receptor. Antagonism of bradykinin B2 receptor also increased acid-induced deep duodenal villous damage. Thus, maintenance of bradykinin-mediated mesenteric hyperemia, is a previous unrecognized mechanism associated with protection of the rat duodenal mucosa against acid-induced damage.     

A role for inducible costimulator protein in the CD28- independent mechanism of resistance to Toxoplasma gondii

Long-term resistance to Toxoplasma gondii is dependent on the development of parasite-specific T cells that produce IFN-gamma. CD28 is a costimulatory molecule important for optimal activation of T cells, but CD28(-/-) mice are resistant to T. gondii, demonstrating that CD28-independent mechanisms regulate T cell responses during toxoplasmosis. The identification of the B7-related protein 1/inducible costimulator protein (ICOS) pathway and its ability to regulate the production of IFN-gamma suggested that this pathway may be involved in the CD28-independent activation of T cells required for resistance to T. gondii. In support of this hypothesis, infection of wild-type or CD28(-/-) mice with T. gondii resulted in the increased expression of ICOS by activated CD4(+) and CD8(+) T cells. In addition, both costimulatory pathways contributed to the in vitro production of IFN-gamma by parasite-specific T cells and when both pathways were blocked, there was an additive effect that resulted in almost complete inhibition of IFN-gamma production. Although in vivo blockade of the ICOS costimulatory pathway did not result in the early mortality of wild-type mice infected with T. gondii, it did lead to increased susceptibility of CD28(-/-) mice to T. gondi associated with reduced serum levels of IFN-gamma, increased parasite burden, and increased mortality compared with the control group. Together, these results identify a critical role for ICOS in the protective Th1-type response required for resistance to T. gondii and suggest that ICOS and CD28 are parallel costimulatory pathways, either of which is sufficient to mediate resistance to this intracellular pathogen.     

Kinetic analysis of the effect of (-)-epigallocatechin gallate on the DNA scission induced by Fe(II)

The DNA strand scission induced by Fe(II) in a citrate buffer solution and the effect of (-)-epigallocatechin gallate (EGCg) were kinetically analyzed. The rate of consumption of dissolved oxygen by Fe(II) in each of these solutions was measured and paralleled that DNA scission. Coordinated EGCg accelerated these reactions. Curves of the time-course characteristics of DNA scission were simulated by using the rate constant of oxygen consumption and by assuming that scission with the hydroxyl radical (OH), which was formed from the dissolved oxygen, proceeded competitively with the scavenging of OH by citrate, Cl- ions and EGCg added. Free EGCg acted as a DNA scission inhibitor to scavenge OH, in contrast to the case of the coordinated one. This analysis is useful for estimating the rate constant of the reaction between an antioxidant and OH, and might provide a new method for measuring the OH-scavenging activity.     

Environmental,health and economic conditions perceived by 50 rural communities in Bangladesh

For randomly selected 50 villages in Bangladesh, an interview survey with a structured questionnaire was conducted to reveal their perception on the environmental, health and economic conditions at present and for the past 10-year change. The eight following items were analyzed in this paper: air pollution and water pollution, which represent environmental conditions with close relation to health conditions, soil degradation and deforestation, which represent environmental conditions with close relation to economic conditions, epidemic diseases and malnutrition, which represent health conditions, and poverty and jobless, which represent economic conditions. Among the 50 villages, deforestation was most frequently perceived serious at present and worsened in the past 10 years. Of the remaining seven items, those related to economic conditions were more seriously perceived than those related to health and environmental conditions. As revealed by the cluster analysis for the inter-item relations, epidemic diseases, which formed the same cluster with the environmental items, were recognized less serious whereas malnutrition, which formed the same cluster with the economic items, was recognized more serious. These findings are useful not only for rural development programs but also for mitigation programs toward health and environmental hazards in Bangladesh.     

Identification of volicitin-related compounds from the regurgitant of lepidopteran caterpillars

Volicitin-related compounds were found in the oral secretion of the three noctuid species, Helicoverpa armigera, Mythimna separata and Spodoptera litura, and one sphingid species, Agrius convolvuli. Volicitin [N-(17-hydroxylinolenoyl)-L-glutamine], N-(17-hydroxylinoleoyl)-glutamine, N-linolenoylglutamine and N-linoleoylglutamine were identified in the secretion from the noctuid larvae. In secretions from the sphingid larvae, N-linolenoylglutamine and N-linoleoylglutamine were the main components. Furthermore, there were significant differences in the amounts of the N-acylamino acid conjugates in the secretions from the three noctuid species. These results suggest that the proportion of volicitin-related compounds in the regurgitant was species-specific.     

Changes in distribution and molecular weight of the acrosomal protein acrin2 (MC41) during guinea pig spermiogenesis and epididymal maturation

In this study, we examined the localization and characteristics of an intra-acrosomal protein, acrin2 (MC41), during guinea pig spermiogenesis and post-testicular sperm maturation in the epididymis, using the monoclonal antibody MC41. Immunoelectron microscopy demonstrated not only a specific domain localization of acrin2 in the apical segment of the guinea pig sperm acrosome, but also its dynamic behavior according to the spermatid differentiation and passage through the epididymis, as follows: acrin2 was exclusively localized in the membrane of the endoplasmic reticulum of early-stage spermatids but was not detectable in the developing acrosome until spermatids reached the maturation phase. In the final stage of spermiogenesis, acrin2 became localized in the outer acrosomal membrane (OAM)/matrix-associated materials both in the small region posterior to the dorsal matrix and along the ventral margin of the acrosomal apical segment. The acrosomal location of acrin2 in caput epididymidal sperm was almost identical to that observed in the final step spermatids, but during maturation it became progressively more restricted in area until on distal cauda epididymidal sperm it remained only in the dorsal region. In Western blot analysis, the MC41 antibody recognized a 165-kDa protein in the mature sperm extract. Furthermore, it was demonstrated that molecular weight reduction of the protein occurred during sperm passage through the epididymis. These findings indicate that acrin2 changes progressively in both distribution and size during development and maturation of the acrosome.     

Generation of 2-arachidonoylglycerol, an endogenous cannabinoid receptor ligand, in picrotoxinin-administered rat brain

The levels of 2-arachidonoylglycerol, an endogenous cannabinoid receptor ligand, and other molecular species of monoacylglycerols in rat brain were examined. In this study, we sacrificed the animals in liquid nitrogen to minimize postmortem changes. We found that rat brain contains 0.23 nmol/g tissue of 2-arachidonoylglycerol, which accounts for 10.5% of the total monoacylglycerol present in this tissue. We next investigated the level of 2-arachidonoylglycerol after in vivo stimulation with picrotoxinin. We found that the level of 2-arachidonoylglycerol was elevated markedly in picrotoxinin-administered rat brain (4- to 6-fold over the control level). Changes in the levels of other molecular species were relatively small or negligible. Several cannabimimetic molecules as well as Delta(9)-tetrahydrocannabinol are known to depress neurotransmission and to exert anticonvulsant activities; endogenous 2-arachidonoylglycerol produced during neural excitation may play a regulatory role in calming the enhanced synaptic transmission.     

High density cultivation of two strains of iron-oxidizing bacteria through reduction of ferric iron by intermittent electrolysis

Electrolytic cultivation was applied to Leptospirillum ferrooxidans strains P3A and CF27, which use ferrous iron to respire aerobically. Ferrous iron was supplied to the bacteria by intermittent electrolytic reduction of ferric iron as electron shuttle using an electrode. The yield of L. ferrooxidans and strain CF27 reached 20- and 50-fold, respectively, higher density than were achievable yields without electrolysis. The time required to obtain high density depended not on the growth ratio, but rather on the original growth rate of each strain.