Gene Cluster Responsible for Secretion of and Immunity to Multiple Bacteriocins,the NKR-5-3 Enterocins

Enterococcus faecium NKR-5-3, isolated from Thai fermented fish, is characterized by the unique ability to produce five bacteriocins, namely, enterocins NKR-5-3A, -B, -C, -D, and -Z (Ent53A, Ent53B, Ent53C, Ent53D, and Ent53Z). Genetic analysis with a genome library revealed that the bacteriocin structural genes (enkA [ent53A], enkC [ent53C], enkD [ent53D], and enkZ [ent53Z]) that encode these peptides (except for Ent53B) are located in close proximity to each other. This NKR-5-3ACDZ (Ent53ACDZ) enterocin gene cluster (approximately 13 kb long) includes certain bacteriocin biosynthetic genes such as an ABC transporter gene (enkT), two immunity genes (enkIaz and enkIc), a response regulator (enkR), and a histidine protein kinase (enkK). Heterologous-expression studies of enkT and ΔenkT mutant strains showed that enkT is responsible for the secretion of Ent53A, Ent53C, Ent53D, and Ent53Z, suggesting that EnkT is a wide-range ABC transporter that contributes to the effective production of these bacteriocins. In addition, EnkIaz and EnkIc were found to confer self-immunity to the respective bacteriocins. Furthermore, bacteriocin induction assays performed with the ΔenkRK mutant strain showed that EnkR and EnkK are regulatory proteins responsible for bacteriocin production and that, together with Ent53D, they constitute a three-component regulatory system. Thus, the Ent53ACDZ gene cluster is essential for the biosynthesis and regulation of NKR-5-3 enterocins, and this is, to our knowledge, the first report that demonstrates the secretion of multiple bacteriocins by an ABC transporter.     

Expression of Set-α during Morphogenesis of Mouse Lower First Molar

The detailed in situ expression pattern of the Set-α gene has been studied. Previously we showed that Set-α is a differentially expressed gene in the embryonic mouse mandible at day 10.5 (E10.5) gestational age. Cells expressing Set-α were widely distributed in both the epithelial and underlying ectomesenchymal cells at E10.5. At E12, they were slightly aggregated in an area where tooth germ of the lower first molar is estimated to be formed. At E13.5, Set-α was strongly expressed in the tooth germ. At the cap stage, Set-α was expressed in the enamel organ and dental papilla. At the bell stage, Set-α was distinctly expressed in the inner enamel epithelial and dental papilla cells facing the inner enamel epithelial layer, which were intended to differentiate into ameloblasts and odontoblasts, respectively. Interestingly, Set-α was also expressed in several embryonic craniofacial tissues derived from the ectoderm. This study is the first report that Set-α is distinctly expressed in the developing tooth germ, and suggests that Set-α plays an important role in both the initiation and the growth of the tooth germ, as well as in the differentiation of ameloblasts and odontoblasts.     

Effects of leptin on hypothalamic arcuate neurons in Wistar and Zucker rats: an in vitro study

Leptin, a product of the ob gene, decreases food intake and body weight in both Wistar and Zucker obese rats when administered centrally or peripherally. To examine whether these leptin effects might be mediated through a neuropeptide Y (NPY) signaling pathway in the medial part of the arcuate nucleus of the hypothalamus (vmARC), the effects of leptin on vmARC neurons in Wistar and Zucker obese rats were examined electrophysiologically using brain slice preparations. Bath application of leptin inhibited about 60% of the vmARC neurons recorded in slices from Wistar rats. Similar inhibitory effects of leptin on vmARC neurons were also observed under low-Ca2+, high-Mg2+ Ringer's solution. However, inhibitory effects were almost absent under Ringer's solution containing a protein kinase C inhibitor, chelerythrine chloride. In slices from Zucker obese rats, leptin inhibited only about 25% of the vmARC neurons recorded, and the proportion of neurons inhibited was significantly smaller for these rats than for Wistar rats. These results suggest that reductions in food intake and body weight induced by leptin in both Wistar and Zucker obese rats are partly mediated via inhibition of an NPY signaling pathway in the vmARC.     

Heme-binding properties of heme detoxification protein from Plasmodium falciparum

The heme detoxification protein of the malaria parasite Plasmodium falciparum is involved in the formation of hemozoin, an insoluble crystalline form of heme. Although the disruption of hemozoin formation is the most widely used strategy for controlling the malaria parasite, the heme-binding properties of heme detoxification protein are poorly characterized. In this study, we established a method for the expression and purification of the non-tagged protein and characterized heme-binding properties. The spectroscopic features of non-tagged protein differ from those of the His-tagged protein, suggesting that the artificial tag interferes with the properties of the recombinant protein. The purified recombinant non-tagged heme detoxification protein had two heme-binding sites and exhibited a spectrum typical of heme proteins. A mechanism for hemozoin formation is proposed.     

Mitochondrials complex I activity is reduced in latent adriamycin-induced cardiomyopathy of rat

We previously reported on the use of enzymatic analysis to impair fatty acid metabolism followed by reduced myocardial energy content, leading to severe heart failure in adriamycin (ADR)-treated rats. The aim of this study is to investigate whether impaired myocardial energy metabolism can also be detected by other methods; i.e. measuring mitochondrial complex I activity and myocardial ¹²⁵I-15-(p-iodophenyl)-3-(R,S)- methylpentadecanoic acid (BMIPP) accumulation in ADR-treated rats. Eight-week-old male Sprague-Dawley rats received 6 intraperitoneal injections of ADR (total 15 mg/kg: group ADR) or saline (control group) over 2 weeks. Left ventricular (LV) ejection fraction was assessed using echocardiography at 3- and 6-weeks after ADR injection (3 weeks and 6 weeks, respectively). Myocardial fatty acid utilization was assessed at 3 weeks and 6 weeks. The myocardial counts of BMIPP were measured after intravenous BMIPP (370 kBq) injection, and ¹²⁵I counts were measured to calculate the uptake ratio. The enzymatic activity of complex I was assessed by monitoring the oxidation of nicotinamide-adenine-dinucleotide-disodium-salt (NADH). In rats treated with ADR, significant decrease in LV ejection fraction was observed only at 6 weeks compared to control (72.5 vs. 84.5%, p < 0.01rpar;. LV ejection fraction at 3 weeks was identical between group ADR and control (81.8 vs. 84.4%). However, at 3 weeks, complex I activity was already reduced significantly in group ADR as compared to control group (p = 0.03), but the reduction in BMIPP accumulation was not (p = 0.15). Our data indicated that reduced complex I activity in a phenomenon occurred in early phase of ADR-induced cardiomyopathy, and it might play an important role in the progression of ADR-induced heart failure.     

Histological observation of the urogenital papillae in the bi-directional sex-changing gobiid fish, Trimma okinawae

The gobiid fish Trimma okinawae changes its sex bi-directionally according to its social status. Morphological changes in the urinogenital papillae (UGP) of this fish have been reported during sex change. However, there have been no detailed observations of such changes. Here, we histologically examined the UGP structure of male- and female-phase fish. UGPs of fish in female and male phase contained both oviducts and sperm ducts. Both ducts were coalesced into one duct within the posterior region of the UGP. Female-phase fish had many longitudinal folds in the hypertrophied tunica mucosa of the oviduct, which was found to be responsible for the transport of eggs and the removal of follicular cells from the oocyte. In contrast, male-phase fish had an immature oviduct and a mature sperm duct in the UGP. In the male-phase fish, the co-existence of spermatozoa and fibrillar secretions was observed in the sperm duct during spermiation.     

Direct Absorption of Methyl Mercury by Lymph

Methyl mercury is contained in fish and seafood products and is taken up into the body in food. While the central nervous system is known as a target organ, methyl mercury also induces autoimmunity and acts as a potent immunosuppressor. The aim of the present study is to know whether methyl mercury is directly absorbed by lymph. Conscious rats were infused with methyl mercury (4 mg/kg) via duodenal tubing as a single pulse infusion, followed by the continuous infusion of saline, and lymphatic fluids were continuously collected from the thoracic lymph duct every 30 min until 360 min after infusion. Mercury was detected immediately after infusion, and total mercury contents in lymph gradually increased until 90–120 min, remained steady, and then gradually decreased until 360 min; however, the amount of mercury collected during 330–360 min was about twofold higher than during 0–30 min. The amount of cumulative mercury in lymph at 360 min was 1.4 μg. In contrast, blood mercury concentration was 2.4 μg/ml 5 min after infusion, with the value at 360 min being 12.6 times higher than at 5 min. Plasma mercury concentration was 56 ng/ml at 5 min, with hundreds of nanograms per milliliter of mercury detected until 360 min. From the present study, it is concluded that some methyl mercury is directly absorbed by lymph and remains steady 6 h after infusion.     

Improved procedure for the enantiometric synthesis of 1-hydroxy/acetoxy-2,6-diaryl-3,7-dioxabicycl

Short enantiomeric syntheses of the 1-hydroxy/acetoxy-3,7-dioxabicyclo[3.3.0]octane lignans, paulownin, and (+)-phrymarin I and II, were accomplished by starting from the chiral synthon, (R)-(+)-3-hydroxybutanolide, and employing photocyclization as the key step.     

Glucagon-like peptide-1 (GLP-1) inhibits advanced glycation end product (AGE)-induced up-regulation of VCAM-1 mRNA levels in endothelial cells by suppressing AGE receptor (RAGE) expression

Glucagon-like peptide-1 (GLP-1) is one of the incretins, a gut hormone secreted from L cells in the intestine in response to food intake. It has been proposed as a potential therapeutic target for the treatment of patients with type 2 diabetes. However, the direct effects of GLP-1 on vascular injury in diabetes are largely unknown. Since there is a growing body of evidence that advanced glycation end products (AGE) and their receptor RAGE axis plays an important role in vascular complications in diabetes, this study investigated whether and how GLP-1 blocked the deleterious effects of AGE on human umbilical vein endothelial cells (HUVEC). GLP-1 receptor (GLP-1R) was expressed in HUVEC. GLP-1 dose-dependently inhibited RAGE gene expression in HUVEC, which was blocked by small interfering RNAs raised against GLP-1R. An analogue of cyclic AMP also decreased RAGE mRNA level in HUVEC. Further, GLP-1 decreased reactive oxygen species generation and subsequently reduced vascular cell adhesion molecule-1 mRNA levels in AGE-exposed HUVEC. Our present study suggests that GLP-1 directly acts on HUVEC via GLP-1R and it could work as an anti-inflammatory agent against AGE by reducing RAGE expression via activation of cyclic AMP pathways.