Clonal structure in Ichthyobacterium seriolicida, the causative agent of bacterial haemolytic jaundice in yellowtail, Seriola quinqueradiata, inferred from molecular epidemiological analysis.

Bacterial haemolytic jaundice caused by Ichthyobacterium seriolicida has been responsible for mortality in farmed yellowtail, Seriola quinqueradiata, in western Japan since the 1980s. In this study, polymorphic analysis of I. seriolicida was performed using three molecular methods: amplified fragment length polymorphism (AFLP) analysis, multilocus sequence typing (MLST) and multiple-locus variable-number tandem repeat analysis (MLVA). Twenty-eight isolates were analysed using AFLP, while 31 isolates were examined by MLST and MLVA. No polymorphisms were identified by AFLP analysis using EcoRI and MseI, or by MLST of internal fragments of eight housekeeping genes. However, MLVA revealed variation in repeat numbers of three elements, allowing separation of the isolates into 16 sequence types. The unweighted pair group method using arithmetic averages cluster analysis of the MLVA data identified four major clusters, and all isolates belonged to clonal complexes. It is likely that I. seriolicida populations share a common ancestor, which may be a recently introduced strain.

[Gene Replacement Therapy for Inherited Retinal Dystrophies]. / Genersatztherapie bei hereditären Netzhauterkrankungen.

Characteristics of inherited retinal dystrophies include deficiencies in light perception and nervous conduction within the retina, leading to reduced vision or even blindness. In this context, the loss of function of photoreceptor-specific genes causes a variety of clinically and aetiologically distinct syndromes - each of them belonging to the group of rare diseases. With a prevalence of 1 in 2500, however, inherited retinal diseases are clinically significant and important - especially since these diseases lead to restrictions of a patient's fitness for work and overall quality of life. More than 250 genetic mutations causing the various types of inherited retinal dystrophies have been identified by now (https://sph.uth.tmc.edu/Retnet). In recent years, preclinical research on suitable animal models has yielded important progress in the understanding of the mutations underlying the pathological and molecular biological processes of these diseases. These findings have led to the development of novel and innovative therapeutic strategies for the treatment of inherited retinal dysfunctions, which are still incurable. Meanwhile, many of the successful preclinical studies have led to translational research projects aiming to find treatment options for human patients. However, some preliminary results of these human translational studies indicate the need to optimise and refine the underlying therapeutic concepts.

Search for nucleon decay via n→ν[over ¯]π0 and p→ν[over ¯]π+ in Super-Kamiokande.

We present the results of searches for nucleon decay via n→ν[over ¯]π0 and p→ν[over ¯]π+ using data from a combined 172.8 kt·yr exposure of Super-Kamiokande-I,-II, and-III. We set lower limits on the partial lifetime for each of these modes: τn→ν[over ¯]π0>1.1×10(33) years and τp→ν[over ¯]π+>3.9×10(32) years at a 90% confidence level.

Search for dinucleon decay into kaons in Super-Kamiokande.

A search for the dinucleon decay pp → K+ K+ has been performed using 91.6 kton·yr data from Super-Kamiokande-I. This decay provides a sensitive probe of the R-parity-violating parameter λ112''. A boosted decision tree analysis found no signal candidates in the data. The expected background was 0.28±0.19 atmospheric neutrino induced events and the estimated signal detection efficiency was 12.6%±3.2%. A lower limit of 1.7×10(32) years has been placed on the partial lifetime of the decay O16 → C14K+ K+ at 90% C.L. A corresponding upper limit of 7.8×10(-9) has been placed on the parameter λ112''.

[Gene replacement therapy in achromatopsia type 2]. / Genersatztherapie bei genetisch bedingter Zapfenblindheit.

Achromatopsia is an autosomal recessive inherited retinal disease caused by a complete loss of cone photoreceptor function. About 80 % of achromatopsia patients show mutations in the alpha or beta subunit (A3 and B3) of the cGMP controlled cation channel CNG (cyclic nucleotide-gated channel) of cone photoreceptors. Homologous to the human disease, CNGA3 deficient mice reveal a loss of cone specific functionality leading to degeneration of affected cone photoreceptors. The Institute for Ophthalmic Research in Tübingen has now succeeded in curing achromatopsia ACHM2 in an animal model. In this article, we explain the recombinant adeno-associated virus-based approach in detail. Furthermore, applied non-invasive diagnostic techniques for quality and success control, ERG, SLO and OCT, are described. The success of the therapy is indicated by a restored cone photoreceptor function as well as the neuronal processing of retinal signals resulting in a specific, cone-mediated behaviour. The outstanding results derived from the animal model are the starting point for the first human translation of a gene therapy for achromatopsia in Germany.

Evidence for the appearance of atmospheric tau neutrinos in super-Kamiokande.

Super-Kamiokande atmospheric neutrino data were fit with an unbinned maximum likelihood method to search for the appearance of tau leptons resulting from the interactions of oscillation-generated tau neutrinos in the detector. Relative to the expectation of unity, the tau normalization is found to be 1.42 ± 0.35(stat)(-0.12)(+0.14)(syst) excluding the no-tau-appearance hypothesis, for which the normalization would be zero, at the 3.8σ level. We estimate that 180.1 ± 44.3(stat)(-15.2)(+17.8) (syst) tau leptons were produced in the 22.5 kton fiducial volume of the detector by tau neutrinos during the 2806 day running period. In future analyses, this large sample of selected tau events will allow the study of charged current tau neutrino interaction physics with oscillation produced tau neutrinos.

Retinal degeneration modulates intracellular localization of CDC42 in photoreceptors.

PURPOSE: Rho GTPases such as RAS-related C3 botulinum substrate 1 (RAC1) and cell division cycle 42 homolog (S. cerevisiae; CDC42) have been linked to cellular processes including movement, development, and apoptosis. Recently, RAC1 has been shown to be a pro-apoptotic factor in the retina during light-induced photoreceptor degeneration. Here, we analyzed the role of CDC42 in the degenerating retina. METHODS: Photoreceptor degeneration was studied in a mouse model for autosomal dominant retinitis pigmentosa (VPP) with or without a rod-specific knockdown of Cdc42, as well as in wild-type and Cdc42 knockdown mice after light exposure. Gene and protein expression were analyzed by real-time PCR, western blotting, and immunofluorescence. Retinal morphology and function were assessed by light microscopy and electroretinography, respectively. RESULTS: CDC42 accumulated in the perinuclear region of terminal deoxynucleotidyl transferase dUTP nick end labeling-negative photoreceptors during retinal degeneration induced by excessive light exposure and in the rd1, rd10, and VPP mouse models of retinitis pigmentosa. The knockdown of Cdc42 did not affect retinal morphology or function in the adult mice and did not influence photoreceptor apoptosis or molecular signaling during induced and inherited retinal degeneration. CONCLUSIONS: Retinal degeneration induces the accumulation of CDC42 in the perinuclear region of photoreceptors. In contrast to RAC1, however, lack of CDC42 does not affect the progression of degeneration. CDC42 is also dispensable for normal morphology and function of adult rod photoreceptor cells. RECEIVED: May 25, 2011 ACCEPTED: November 10, 2011.

Search for differences in oscillation parameters for atmospheric neutrinos and antineutrinos at Super-Kamiokande.

We present a search for differences in the oscillations of antineutrinos and neutrinos in the Super-Kamiokande-I, -II, and -III atmospheric neutrino sample. Under a two-flavor disappearance model with separate mixing parameters between neutrinos and antineutrinos, we find no evidence for a difference in oscillation parameters. Best-fit antineutrino mixing is found to be at (Δm2,sin2 2θ)=(2.0×10(-3) eV2, 1.0) and is consistent with the overall Super-K measurement.

Induction of STAT3-related genes in fast degenerating cone photoreceptors of cpfl1 mice.

Cone dystrophies are genetic diseases characterized by loss of cone photoreceptor function and severe impairment of daylight vision. Loss of function is accompanied by a progressive degeneration of cones limiting potential therapeutic interventions. In this study we combined microarray-based gene-expression analysis with electroretinography and immunohistochemistry to characterize the pathological processes in the cone photoreceptor function loss 1 (cpfl1) mouse model. The cpfl1-mouse is a naturally arising mouse mutant with a loss-of-function mutation in the cone-specific Pde6c gene. Cpfl1-mice displayed normal rod-specific light responses while cone-specific responses were strongly diminished. Despite the lack of a general retinal degeneration, the cone-specific functional defect resulted in a marked activation of GFAP, a hallmark of Müller-cell gliosis. Microarray-based network-analysis confirmed activation of Müller-glia-specific transcripts. Unexpectedly, we found up-regulation of the cytokine LIF and the anti-apoptotic transcription factor STAT3 in cpfl1 cone photoreceptors. We postulate that STAT3-related pathways are induced in cpfl1 cone photoreceptors to counteract degeneration.

Rod and cone a-waves in three cases of Bietti crystalline chorioretinal dystrophy.

PURPOSE: To estimate retinal function in Bietti crystalline chorioretinal dystrophy using the electroretinogram. METHODS: In this observational case series, the scotopic and photopic electroretinograms in three Japanese female patients (case 1, 55 years old; case 2, 56 years old; case 3, 47 years old) who showed bilateral crystalline retinal deposits but no corneal deposits were recorded. The rod and cone a-waves were analyzed by using the method described by Hood and Birch (1995, 1997). The parameters Rm(p3) (maximum a-wave amplitude) and S (sensitivity) were calculated. RESULTS: In case 1, the rod Rm(p3) was decreased in both eyes. The rod S in the right eye was within the normal range, but that in the left eye was significantly reduced. Although the cone Rm(p3) was decreased, the cone S was within the normal range. In case 2, the rod and cone Rm(p3) was reduced, but the rod and cone S was within the normal range in both eyes. In case 3, the rod and cone Rm(p3) and S were within the normal range. CONCLUSIONS: Electroretinograms illustrated different disease stages, however, no eye with normal Rm(p3) and decreased S was found in rods and cones. In the early stages of this disease, decreased numbers of photoreceptors and/or outer segment shortening may be present while phototransduction remains normal. As the damage to the retina progresses, phototransduction becomes severely affected. Because reduced cone S was not observed in our cases, cones may be less involved than rods in this disease.

Cholecystoparesis with diabetic autonomic neuropathy.

A 69-year-old male diabetic patient was hospitalized with pneumonia in February 1996. Abdominal ultrasonography performed as a routine examination on admission revealed marked dilatation of gallbladder with a fasting maximal size of 25.7 cm2 compared with 8.8 +/- 2.9 cm2 evaluated in 30 male healthy controls aged 67 +/- 8 years old. Four months after recovery from pneumonia, abnormal gallbladder dilatation remained unchanged (25.0 cm2), while dilatation had not been observed on an examination performed 6 years earlier (9.8 cm2). Because of accompanying neurological defects, we suppose that cholecystoparesis may be caused by progression of autonomic neuropathy in this patient with diabetes mellitus.

Simple obesity with cardiomyopathy of obesity.

A 32-year-old obese female was hospitalized with dyspnea. Echocardiogram revealed left ventricular dilatation. Chest X-ray film showed enlarged heart size and prominent pulmonary congestion. Simple obesity with congestive heart failure (CHF) due to cardiomyopathy of obesity was diagnosed according to the absence of obvious disease that caused obesity or CHF. After diet therapy and medication, subjective symptoms disappeared and body weight was reduced from 137 kg to 85 kg. Although few reports of cardiomyopathy of obesity have been reported in Japan, we propose the possibility that similar cases will be on the increase because Japanese dietary habits are now becoming more similar to those of Caucasians.

Immunohistochemical study of age-dependent brain lesions in mice infected intracerebrally with Kasba (Chuzan) virus.

When Kasba (Chuzan) virus (an orbivirus) was injected intracerebrally into 1-, 2- or 4-week-old mice, non-purulent necrotizing encephalitis developed and the mice showed nervous symptoms and became moribund. The necrotic lesions were more severe in younger animals. In 1-week-old mice, viral titres rose until 7 days post-infection, while in 2- and 4-week-old animals the titres reached a peak on day 3 and then declined gradually. Glial fibrillary acidic protein-positive astrocytes increased in the white matter, hippocampus and subpial area of the cerebral cortex of infected animals, and lectin-RCA-1-positive cells, thought to be microglial cells, increased in the necrotic lesions. The number of these glial cells increased even after viral titres had declined. In this study there were no survivors in any age group, but survival time increased with age.

Comparison of arterial blood gases and acid-base balance in young and aged beagle dogs, with regard to postprandial alkaline tide.

Arterial blood pH and blood gas tensions were measured before and after feeding in seven young (1-year-old) and 6 aged (10 to 12 year-old) conscious beagle dogs with a portable "i-STAT" blood gas analyzer to obtain the fundamental physiological data on the laboratory dogs. Before feeding, arterial blood pH, HCO3- and base excess (BE) were 7.395 +/- 0.022, 20 +/- 1.0 mmol/L and -4 +/- 1.1 mmol/L in young dogs, and 7.408 +/- 0.031, 23 +/- 1.5 mmol/L and -2 +/- 1.8 mmol/L in aged ones. After feeding, the above values changed to 7.449 +/- 0.021, 25 +/- 1.1 and 1 +/- 1.3 in young dogs, and 7.456 +/- 0.022, 28 +/- 1.7 and 4 +/- 1.9 in aged ones. After feeding, each of the values for pH, HCO3- and BE was increased in comparison with those before feeding in both young and aged dogs, and alkalinization of blood, the alkaline tide, was observed in all healthy dogs. The values for PCO2 also rose from 33.6 +/- 1.91 to 36.7 +/- 1.95 (mmHg) in young dogs, and 36.4 +/- 2.97 to 40.3 +/- 3.06 (mmHg) in aged ones, and in 3 of 6 aged dogs slight hypoxia (PO2 < 80 mmHg) was noted. These changes in PCO2 and PO2 were considered to be due to compensatory hypoventilation for the alkalinization of blood. To confirm the cause of the alkaline tide phenomenon, we investigated the association of the alkaline tide with gastric acid secretion by cimetidine, an H2-blocker; and found that it was effective to inhibit postprandial alkaline tide in dogs. After feeding, ionized Ca concentration (iCa) was decreased inversely with an arterial blood pH increase in both young and aged dogs. The mean values of iCa in aged dogs after feeding was lower than those in young dogs. In the present experiments, it was demonstrated that in beagle dogs the acid-base balance and blood gas values were largely affected by feeding, especially in aged dogs. In case blood gases would be evaluated in toxicity studies, we should consider the influence of feeding and aging.

[The role of stimulation of the medullary reticular formation on intracranial pressure in cold injured brain].

This study was carried out to study the effect of stimulation of the reticular formation of the medulla oblongata on intracranial pressure (ICP) and cerebral blood volume (CBV) in injured brain with increased ICP. CBV was measured by the photoelectric method from the parietal lobe. Seventeen hours prior to the experiments, cold induced edema was produced to increase basal ICP. In 15 cats, electric stimulation produced temporary increases in BP, ICP and CBV and progressive intracranial hypertension was never observed (Group A). In 9 animals, progressive increases in CBV and ICP up to 50 to 100 mmHg were evoked after cessation of stimulation (Group B). Prestimulation ICP in Group B was significantly higher than that of Group A. Rapid and simultaneous increases in ICP and CBV following stimulation strongly suggested that global increments of CBV secondary to loss of cerebral vasomotor tonus were responsible for producing progressive intracranial hypertension. In Group B, the stimulation electrodes were invariably located at the area of the nucleus reticularis parvocellularis and gigantocellularis. Our experimental results show that in increased ICP a stimulated or irritable condition of the medullary reticular formation will cause acute progressive intracranial hypertension.

[Endoscopic injection of liposomal adriamycin targeting lymph node metastasis of gastric cancer].

Gastric submucosal injection of 5 mg liposomal adriamycin (L-ADM) close to the main gastric cancer tumor was done in 15 patients by endoscopy. This approach was based on the idea that preoperative adjuvant chemotherapy targeting lymph node metastasis in patients with gastric cancer may be effective for prevention of lymph node recurrence. ADM concentrations in the regional lymph nodes were assessed and compared with those in patients who were administered 5 mg of free adriamycin (F-ADM) i.v. preoperatively. ADM concentrations in Group 7 lymph nodes (according to the General Rules for Gastric Cancer Study) were: After 2 days; 7.21 +/- 5.98 micrograms/g (n = 2) in the L-ADM group and 0.59 +/- 0.23 micrograms/g (n = 3) in the F-ADM group. After 4 days; 4.93 +/- 3.93 micrograms/g (n = 2) in the L-ADM group and 0.36 +/- 0.0 micrograms/g (n = 2) in the F-ADM group. After 6 days; 2.08 +/- 0.49 micrograms/g (n = 2) in the L-ADM group and 0.05 +/- 0.05 micrograms/g (n = 3) in the F-ADM group. L-ADM group: those who had L-ADM injected into the side of the lesser curvature of the stomach. F-ADM group: those who had F-ADM administered i.v. These data demonstrate that gastric submucosal injection of L-ADM is well suited for specific delivery to the regional lymph nodes, suggesting that this type of administration may prevent lymph node recurrence of gastric cancer by targeting lymph node metastasis.

[Clinical application of chemotherapy via the portal vein with liposome-encapsulated adriamycin in inoperable metastatic liver cancer].

We studied the effects of liposome-entrapped adriamycin (L-ADM) administered via the portal vein and the clinical application of this treatment in the therapy and inhibition of liver metastasis, experimentally and clinically. Liposomes composed of egg phosphatidylcholine (cholesterol 50 mol%) were used as drug carriers. We examined the distribution in tissues and antitumor effect of freeze-dried L-ADM administered via the portal vein to rabbits bearing VX2 tumors. The liver concentration of ADM increased after delivery and cardiac uptake decreased compared with free drug treatment. The life span was prolonged by L-ADM treatment compared with the control group and the free ADM group. This L-ADM administration was confirmed to be safe and revealed a decrease in the heart toxicities compared with free adriamycin. Nineteen cases were studied from Jan. 1986 to May 1991 via the portal vein and the clinical effects were evaluated. From Mar. 1988 to date, 10 cases were treated with L-ADM (20-30 mg every 2 weeks/body) in patients with inoperable cases using subcutaneously implanted reservoir. The median survival was 450 days; 275 days for colon cancer, 492 days for gastric cancer, and 1,052 days for uterine cancer (range: 136-1,152 days), compared with 141 days (range: 52-253 days) in 9 cases of historical control treated with free-ADM via the portal vein. These results suggest that chemotherapy via the portal vein with L-ADM for metastatic liver cancer may increase survival time.

[Antitumor effect of liposome-entrapped carboplatin after intraperitoneal administration in rats].

We examined the distribution in tissue and antitumor effects of freeze-dried liposome-entrapped carboplatin (Lipo-CBDCA) after intraperitoneal administration to rats bearing AH 130 tumors. Liposomes composed of egg lecithin and cholesterol were used as drug carriers. The serum concentration of platinum was decreased for a short time after the intraperitoneal administration of Lipo-CBDCA. After at least 3 hours, the serum concentration of platinum was higher with free CBDCA intraperitoneal or intravenous administration. The antitumor effects of Lipo-CBDCA were determined in rats with peritoneal dissemination due to AH 130 tumors. Intraperitoneal Lipo-CBDCA prolonged the life span of the tumor-bearing rats. No side effects of the chemotherapy were demonstrated in biochemical and histological studies in the liver, kidney, spleen and small intestine. These results indicate that intraperitoneal chemotherapy with Lipo-CBDCA may be more effective than that with free CBDCA managing in peritoneal dissemination, and may be therapeutically useful without toxic side effects.

[Efficacy of preoperative adjuvant chemotherapy using adriamycin targeting the regional lymph nodes for gastric cancer--lymph node-targeting delivery of adriamycin in rabbits].

Chemotherapy targeting the regional lymph nodes for gastric cancer may be more effective preoperatively than postoperatively since anticancer drugs can be transported to the regional lymph nodes via the lymphatic flow through the stomach. Distribution of Adriamycin (ADR) among the various organs was assessed following its intravenous injection in rabbits. The delivery index of the drug to each organ was assessed by the ratio of the area under the concentration-time curve (AUC) of each organ to the AUC of the regional lymph nodes following the intravenous administration. The delivery index was 0.14 for the stomach, 0.11 for the heart, 0.53 for bone marrow, 0.74 for the spleen, and 0.14 for the liver. These data suggest that preoperative adjuvant chemotherapy by intravenous administration of ADR may be effective in targeting ADR at the regional lymph nodes. Tissue ADR concentrations in the regional lymph nodes were assessed following gastric submucosal administration of ADR in rabbits. The targeting index for the regional lymph nodes was 8.20, measured by the ratio of AUC following a gastric submucosal injection to AUC after the intravenous injection of ADR. This suggests that it may be possible to selectively target chemotherapy to regional lymph nodes by employing a gastric submucosal administration of ADR.

Knockout of PARG110 confers resistance to cGMP-induced toxicity in mammalian photoreceptors.

Hereditary retinal degeneration (RD) relates to a heterogeneous group of blinding human diseases in which the light sensitive neurons of the retina, the photoreceptors, die. RD is currently untreatable and the underlying cellular mechanisms remain poorly understood. However, the activity of the enzyme poly-ADP-ribose polymerase-1 (PARP1) and excessive generation of poly-ADP-ribose (PAR) polymers in photoreceptor nuclei have been shown to be causally involved in RD. The activity of PARP1 is to a large extent governed by its functional antagonist, poly-ADP-glycohydrolase (PARG), which thus also may have a role in RD. To investigate this, we analyzed PARG expression in the retina of wild-type (wt) mice and in the rd1 mouse model for human RD, and detected increased PARG protein in a subset of degenerating rd1 photoreceptors. Knockout (KO) animals lacking the 110 kDa nuclear PARG isoform were furthermore analyzed, and their retinal morphology and function were indistinguishable from wild-type animals. Organotypic wt retinal explants can be experimentally treated to induce rd1-like photoreceptor death, but PARG110 KO retinal explants were unexpectedly highly resistant to such treatment. The resistance was associated with decreased PAR accumulation and low PARP activity, indicating that PARG110 may positively regulate PARP1, an event that therefore is absent in PARG110 KO tissue. Our study demonstrates a causal involvement of PARG110 in the process of photoreceptor degeneration. Contrasting its anticipated role as a functional antagonist, absence of PARG110 correlated with low PARP activity, suggesting that PARG110 and PARP1 act in a positive feedback loop, which is especially active under pathologic conditions. This in turn highlights both PARG110 and PARP1 as potential targets for neuroprotective treatments for RD.