Ostreopsis sp., a possible origin of palytoxin (PTX) in parrotfish Scarus ovifrons.

A clone of toxic dinoflagellate Ostreopsis sp. and six specimens of a parrotfish Scarus ovifrons were collected in October 1997 at Tokushima Prefecture, Japan. Ostreopsis sp. was cultured in ESM medium for 16 days, and after rearing the cell pellet (about 4.0x10(5) cells) was extracted with 50% methanol, partitioned between an aqueous layer and 1-butanol layer, and biochemically tested. Similarly, the crude toxin from S. ovifrons was extracted, and tested. The mice injected with each 1-butanol layer from Ostreopsis sp. and S. ovifrons showed the common symptoms of convulsion, drowsiness and collapse, and died within 48 h. The lethal potency of Ostreopsis sp. was calculated to be 1.0x10(-4) MU/cell. All specimens of S. ovifrons were found to be toxic, where the highest potency was determined as 2 MU/g in muscle of one specimen. After being injected with toxins, the serum creatine phosphokinase levels of mice were found to be elevated. Toxins from Ostreopsis sp. and S. ovifrons showed delayed haemolytic activity with mouse and human erythrocytes, which was inhibited by an anti-palytoxin (PTX) antibody antibody and ouabain. Toxins from Ostreopsis sp. and S. ovifrons thus resembled each other, and strongly suggested to be PTX or its akin substance. Additionally, a considerable number of adherent Ostreopsis sp. was found in the gut contents of S. ovifrons during the heavy occurrence of Ostreopsis sp. in October 1997 at Tokushima Prefecture. From the above results, it can be strongly postulated that the dinoflagellate Ostreopsis sp. is the origin of PTX which is sequestered by the parrotfish S. ovifrons through food chain.     

Role of SCOX in determination of Drosophila melanogaster lifespan

In man, COX (cytochrome c oxidase) deficiency is reported to be related to mutation of the SCO2 (synthesis of cytochrome c oxidase 2) gene, which encodes one of the copper-donor chaperones involved in the assembly of mitochondrial cytochrome c oxidase. Such COX deficiency due to the genetic condition leads to heart disease and the Leigh syndrome and is frequently fatal in childhood. Synthesis of cytochrome c oxidase X (SCOX) is a Drosophila orthologue of human SCO2. Here, we generated SCOX-knockdown flies and the full length SCOX transgenic flies to investigate the in vivo roles of SCOX. Our results demonstrated knockdown of SCOX gene in all cells and tissues to be associated with lethality at larval or pupal stages and this correlated with a decrease in ATP level. In contrast, the full length SCOX transgenic flies showed a longer lifespan than wild type flies and control flies carrying Act5C-GAL4 alone and this correlated with an increase in ATP level. Finally, when cultured on paraquat-added medium, full length SCOX transgenic flies also exhibited an elongated lifespan. Therefore, we hypothesized that SCOX plays an important role in ATP production and consumption, which helps to prevent production of mitochondrial reactive oxygen species and/or impairment of mitochondrial activity under oxidative stress.     

Evaluation of in vivo genotoxicity induced by N‐ethyl‐N‐nitrosourea,benzo[a]pyrene,and 4‐nitroquinoline‐1‐oxide in the Pig‐a and gpt assays

The recently developed Pig‐a mutation assay is based on flow cytometric enumeration of glycosylphosphatidylinositol (GPI) anchor‐deficient red blood cells caused by a forward mutation in the Pig‐a gene. Because the assay can be conducted in nontransgenic animals and the mutations accumulate with repeat dosing, we believe that the Pig‐a assay could be integrated into repeat‐dose toxicology studies and provides an alternative to transgenic rodent (TGR) mutation assays. The capacity and characteristics of the Pig‐a assay relative to TGR mutation assays, however, are unclear. Here, using transgenic gpt delta mice, we compared the in vivo genotoxicity of single oral doses of N‐ethyl‐N‐nitrosourea (ENU, 40 mg/kg), benzo[a]pyrene (BP, 100 and 200 mg/kg), and 4‐nitroquinoline‐1‐oxide (4NQO, 50 mg/kg) in the Pig‐a (peripheral blood) and gpt (bone marrow and liver) gene mutation assays. Pig‐a assays were conducted at 2, 4, and 7 weeks after the treatment, while gpt assays were conducted on tissues collected at the 7‐week terminal sacrifice. ENU increased both Pig‐a and gpt mutant frequencies (MFs) at all sampling times, and BP increased MFs in both assays but the Pig‐a MFs peaked at 2 weeks and then decreased. Although 4NQO increased gpt MFs in the liver, only weak, nonsignificant increases (two‐ or threefold above control) were detected in the bone marrow in both the Pig‐a and the gpt assay. These findings suggest that further studies are needed to elucidate the kinetics of the Pig‐a mutation assay in order to use it as an alternative to the TGR mutation assay. Environ. Mol. Mutagen. 54:747–754, 2013. © 2013 Wiley Periodicals, Inc.     

Clinical Potential of Lomerizine,a Ca2+ Channel Blocker as an Anti‐Glaucoma Drug: Effects on Ocular Circulation and Retinal Neuronal Damage

Glaucoma is defined as an optic neuropathy with characteristic changes in the optic nerve head and ultimate loss of visual field. Previous studies have suggested that (a) mechanical damage due to raised intraocular pressure and (b) a compromised tissue circulation in the optic nerve head play significant roles in the development of glaucomatous damage in the optic nerve head. Recently, we found that lomerizine, a new Ca²⁺ channel blocker, increased ocular circulation and protected neuronal cells against retinal neurotoxicity both in vitro and in vivo with minimal cardiovascular side effects. We examined the effect of lomerizine on the ocular circulation and compared it with those of other Ca²⁺ channel blockers in normal rabbits and in rabbits with an endothelin‐1‐disturbed circulation in the optic nerve head. In anesthetized rabbits, lomerizine and the other Ca²⁺ channel blockers increased the ocular circulation and also inhibited the hypoperfusion induced in optic nerve head tissue by an intravitreous injection of endothelin‐1. Whereas the other Ca²⁺ channel blockers produced changes in blood pressure and heart rate, the effects of lomerizine on these parameters were slight. In healthy humans, lomerizine increased blood velocity in the optic nerve head, without significantly altering blood pressure or heart rate. Moreover, lomerizine reduced retinal damage in rats both in vitro and in vivo, presumably through a Ca²⁺ channel blocking effect via an action that may involve a direct protection of retinal neurons as well as an improvement in the ocular circulation. These results indicate that lomerizine may be useful as a therapeutic drug against ischemic retinal diseases (such as glaucoma and retinal vascular occlusive diseases) that involve a disturbance of the ocular circulation.     

High concentrations of human β-defensin 2 in gastric juice of patients with Helicobacter pyloriinfection

AIM: Human beta-defensin (HBD)-1 and HBD-2 are endogenous antimicrobial peptides. Unlike HBD-1, the HBD-2 expression is augmented by Helicobacter pylori (H pylori). We sought to determine HBD-1 and HBD-2 concentrations in gastric juice during H pylori infection. METHODS: HBD-1 and HBD-2 concentrations were measured by radioimmunoassay in plasma and gastric juice of 49 H pylori-infected and 33 uninfected subjects and before and after anti-H pylori treatment in 13 patients with H pylori-associated gastritis. Interleukin (IL)-1beta and IL-8 concentrations in gastric juice were measured by enzyme-linked immunosorbent assay (ELISA). Histological grades of gastritis were determined using two biopsy specimens taken from the antrum and corpus. Reverse phase high performance liquid chromatography (RP-HPLC) was used to identify HBD-2. RESULTS: HBD-2 concentrations in gastric juice, but not in plasma, were significantly higher in H pylori-positive than -negative subjects, albeit the post-treatment levels were unchanged. Immunoreactivity for HBD-2 was exclusively identified in H pylori-infected mucosa by RP-HPLC. HBD-2 concentrations in gastric juice correlated with histological degree of neutrophil and mononuclear cell infiltration in the corpus. IL-1beta levels correlated with those of IL-8, but not HBD-2. Plasma and gastric juice HBD-1 concentrations were similar in H pylori-infected and uninfected subjects. CONCLUSION: Our results place the beta-defensins, especially HBD-2, in the front line of innate immune defence. Moreover, HBD-2 may be involved in the pathogenesis of H pylori-associated gastritis, possibly through its function as immune and inflammatory mediator.