<Emphasis Type="Italic">Streptomyces</Emphasis> metabolites in divergent microbial interactions

Streptomyces and related bacteria produce a wide variety of secondary metabolites. Of these, many compounds have industrial applications, but the question of why this group of microorganism produces such various kinds of biologically active substances has not yet been clearly answered. Here, we overview the results from our studies on the novel function and role of Streptomyces metabolites. The diverged action of negative and positive influences onto the physiology of various microorganisms infers the occurrence of complex microbial interactions due to the effect of small molecules produced by Streptomyces. The interactions may serve as a basis for the constitution of biological community.     

Gibberellic acid improved shoot multiplication in Cephaelis ipecacuanha

The internodes of Cephaelis ipecacuanha elongated when cultured on Gamborg B5 solid medium supplemented with 0.5 or 1 mg/L gibberellic acid (GA₃). The size of the elongated internode doubled in length compared to the untreated shoots, and the adventitious shoots formed on the elongated internodes. The shoots grew easily into plantlets without the use of auxin for rooting. The ex vitro regenerates cultivated in the greenhouse showed normal characteristics. Emetic alkaloids were detected in the leaves of in vitro shoots and the roots of regenerates cultivated in the greenhouse. This method using GA₃ propagated numerous plants at a rate of more than 100 times compared to the method without GA₃.     

Amperometric biosensor based on enzymes immobilized in hybrid mesoporous membranes for the determination of acetylcholine

Acetylcholine sensor is successfully prepared by using immobilized enzymes, i.e., acetylcholinesterase and choline oxidase within separate hybrid mesoporous silica membranes with 12 nm pore diameter (F127M). The measurement was based on the detection of hydrogen peroxide produced by two sequential enzyme reactions. The determination range and the response time are 6.0–800 μM and within approximately 3 min, respectively. The sensor is very stable compared to free enzymes and 80% of the initial response was maintained even after storage for 80 days. These results show that two enzymes are successfully immobilized and well stabilized, and at the same time, two sequential enzyme reactions efficiently proceed within the separate hybrid mesoporous membranes. Further, we studied the possible detection of organophosphorus pesticides in terms of the inhibition of acetylcholinesterase activity, i.e., the decrease of current response, and demonstrated that the nanomolar concentrations of pesticide (DZN-oxon) can be detected with our sensor.     

Adiponectin ameliorates hypoxia-induced pulmonary arterial remodeling

We have demonstrated that adiponectin has anti-atherosclerotic properties. We also reported hypoadiponectinemia and nocturnal reduction in circulating adiponectin concentrations in patients of severe obstructive sleep apnea-hypopnea syndrome (OSAS). OSAS is often complicated with pulmonary hypertension. In this study, we investigated the effect of adiponectin on chronic hypoxia-induced pulmonary arterial remodeling in mice. Exposure of mice to 3-weeks sustained hypoxia (10% O₂) resulted in significant accumulation of adiponectin in pulmonary arteries. The percentage media wall thickness (%MT), representing pulmonary arterial remodeling, under hypoxic condition, was greater in adiponectin-knockout mice than wild-type mice. Overexpression of adiponectin significantly decreased hypoxia-induced pulmonary arterial wall thickening and right ventricular hypertrophy. These findings demonstrate for the first time that overexpression of adiponectin suppresses the development of hypoxic-induced pulmonary remodeling, and that adiponectin may combat a new strategy for pulmonary vascular changes that underlie pulmonary hypertension in OSAS.     

RhoA induces expression of inflammatory cytokine in adipocytes

Rho GTPase regulates actin cytoskeleton organization and assembly in many cell types, however, its significance in adipose tissue is not well characterized. Here, we demonstrate high RhoA activity in adipose tissues of C57BL/6J mice. To determine the effect of RhoA activation on 3T3-L1 cells, stable cell lines overexpressing G14VRhoA fused to destabilizing domain of FKBP12 (DD-G14VRhoA-L1) were generated. Treatment of DD-G14VRhoA-L1 cells with Shield1 following their differentiation into adipocytes, resulted in the appearance of thick cortical actin filaments, and increased the mRNA expression levels of plasminogen activator inhibitor type-1 (PAI-1) and monocyte chemoattractant protein-1 (MCP-1). The induction of PAI-1 and MCP-1 was inhibited by treatment with a Rho-associated kinase (ROCK) inhibitor, Y-27632. In 3T3-L1 adipocytes, tumor necrosis factor-α activated RhoA and increased mRNA expression of PAI-1 and MCP-1, and their treatment with Y-27632 partially inhibited these changes. The results indicate that RhoA-ROCK pathway induces inflammatory cytokine expression in adipocytes.     

Clofibrate treatment promotes branched-chain amino acid catabolism and decreases the phosphorylation state of mTOR, eIF4E-BP1, and S6K1 in rat liver

Leucine stimulates protein synthesis by modulating the mammalian target of rapamycin (mTOR) signaling pathway. We hypothesized that promotion of the branched-chain amino acid (BCAA) catabolism might influence the leucine-induced protein synthesis. Clofibric acid (an active metabolite of clofibrate) is known to promote the BCAA catabolism by activation of branched-chain alpha-keto acid dehydrogenase complex (BCKDC), the rate-limiting enzyme of the BCAA catabolism. In the present study, we examined the phosphorylation state of mTOR, eukaryotic initiation factor 4E-binding protein-1 (4E-BP1), and ribosomal protein S6 kinase 1 (S6K1) in liver of rats with or without activation of the BCKDC by clofibrate treatment. Clofibrate-treated rats were prepared by oral administration of clofibrate 5 h before sacrifice. In order to stimulate phosphorylation of components in the mTOR signaling pathway, rats were orally administered with leucine 1 h before sacrifice. Clofibrate treatment almost fully activated hepatic BCKDC and significantly decreased the plasma leucine concentration in rats without leucine administration, resulting in decreased mTOR and 4E-BP1 phosphorylation. Similarly, in rats administered with leucine, clofibrate treatment attenuated the predicted increase in plasma leucine concentration as well as the phosphorylation of mTOR, 4E-BP1, and S6K1. These results suggest that BCAA catabolism enhanced by clofibrate treatment has significant influences on the leucine-induced activation of translation initiation processes.     

Steroid hormones as bactericidal agents to Helicobacter pylori

Helicobacter pylori is a unique bacterial species that assimilates various steroids as membrane lipid components. Our group has recently found, however, that certain steroids may impair the viability of H. pylori. In this study, we go on to reveal that estradiol, androstenedione, and progesterone (PS) all have the potential to inhibit the growth of H. pylori. Of these three steroid hormones, progesterone demonstrated the most effective anti-H. pylori action. 17α-hydroxyprogesterone caproate (17αPSCE), a synthetic progesterone derivative, had a much stronger anti-H. pylori action than progesterone, whereas 17α-hydroxyprogesterone, a natural progesterone derivative, completely failed to inhibit the growth of the organism. Progesterone and 17αPSCE were both found to kill H. pylori through their bacteriolytic action. Among five bacterial species investigated, H. pylori was the only species susceptible to the bactericidal action of progesterone and 17αPSCE. The other four species, Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus, and Staphylococcus epiderimidis, all resisted this action. Progesterone and free-cholesterol (FC) obstructed each other's effects against the H. pylori cell. Taken in sum, these results suggest that progesterone and FC may bind to the identical region on the H. pylori cell surface. We expect these findings to contribute to the development of a novel anti-H. pylori steroidal agent.