Successful Heart Transplantation After Desensitization in a Patient With Extremely High Panel-Reactive Antibody Levels and Pretransplant Donor-Specific Antibody: A Case Report.

Elevated panel-reactive antibody (PRA) levels serve as a significant risk factor for allograft survival and episodes of rejection after heart transplantation (HTX). Patients with high PRA levels tend to show expressions of donor-specific human leukocyte antigen antibodies (DSA), which can cause catastrophic hyperacute rejection after HTX. Therefore, such highly sensitized patients are required to undergo strategic perioperative desensitization therapy. We describe a successful HTX after desensitization in a patient with extremely high PRA levels and pretransplant DSA positivity.

Dual Gas Treatment With Hydrogen and Carbon Monoxide Attenuates Oxidative Stress and Protects From Renal Ischemia-Reperfusion Injury.

BACKGROUND: Hydrogen (H2) and carbon monoxide (CO) gas are both reported to reduce reactive oxygen species and alleviate tissue ischemia-reperfusion (I-R) injury. The present study was conducted to evaluate the effects of a mixture of H2 gas and CO gas (dual gas) in comparison with hydrogen gas (H2: 2%) alone on I-R renal injury (composition of dual gas; N2: 77.8%; O2: 20.9%; H2: 1.30%; CO: 250 parts per million). METHODS: Adult male Sprague-Dawley rats (body weight 250-280 g) were divided into 5 groups: (1) sham operation control, (2) dual gas inhalation (dual treatment) without I-R treatment, (3) I-R renal injury, (4) H2 gas alone inhalation (H2 treatment) with I-R renal injury, and (5) dual treatment with I-R renal injury. I-R renal injury was induced by clamping the left renal artery and vein for 45 minutes followed by reperfusion, and then contralateral nephrectomy was performed 2 weeks later. Renal function was markedly decreased at 24 hours after reperfusion, and thereafter the effects of dual gas were assessed by histologic examination and determination of the superoxide radical, together with functional and molecular analyses. RESULTS: Pathologic examination of the kidney of I-R rats revealed severe renal damage. Importantly, cytoprotective effects of the dual treatment in comparison with H2 treatment and I-R renal injury were observed in terms of superoxide radical scavenging activity and histochemical features. Rats given dual treatment and I-R renal injury showed significant decreases in blood urea nitrogen. Increased expression of several inflammatory cytokines (tumor necrosis factor-α, interleukin-6, intracellular adhesion molecule-1, nuclear factor-κB, hypoxia inducible factor-1α, and heme oxygenase-1) was attenuated by the dual treatment. CONCLUSIONS: Dual gas inhalation decreases oxidative stress and markedly improves I-R-induced renal injury.

Evaluation of bovine viral diarrhoea virus control strategies in dairy herds in Hokkaido, Japan, using stochastic modelling.

Bovine viral diarrhoea virus (BVDV) infection in cattle can result in growth retardation, reduced milk production, reproductive disorders and death. Persistently infected animals are the primary source of infection. In Hokkaido, Japan, all cattle entering shared pastures in summer are vaccinated before movement for disease control. Additionally, these cattle may be tested for BVDV and culled if positive. However, the effectiveness of this control strategy aiming to reduce the number of BVDV-infected animals has not been assessed. The aim of this study was to evaluate the effectiveness of various test-and-cull and/or vaccination strategies on BVDV control in dairy farms in two districts of Hokkaido, Nemuro and Hiyama. A stochastic model was developed to compare the different control strategies over a 10-year period. The model was individual-based and simulated disease dynamics both within and between herds. Parameters included in the model were obtained from the literature, the Hokkaido government and the Japanese Ministry of Agriculture, Forestry and Fisheries. Nine different scenarios were compared as follows: no control, test-and-cull strategies based on antigen testing of either calves or only cattle entering common pastures, vaccination of all adult cattle or only cattle entering shared pastures and combinations thereof. The results indicate that current strategies for BVDV control in Hokkaido slightly reduced the number of BVDV-infected animals; however, alternative strategies such as testing all calves and culling any positives or vaccinating all susceptible adult animals dramatically reduced those. To our knowledge, this is the first report regarding the comparison of the effectiveness between the current strategies in Hokkaido and the alternative strategies for BVDV control measures.

Prevalence of Baylisascaris Roundworm in Captive Kinkajous in Japan.

Baylisascaris potosis causes larva migrans in animals. The present study evaluated the prevalence of B. potosis in captive kinkajous ( Potos flavus ) and the ability of milbemycin to treat natural infections of B. potosis in 2 female wild-caught kinkajous. In 2012, fecal samples were collected from 16 kinkajous in 6 zoological gardens and 29 imported captive kinkajous from 4 pet traders in Japan. Although all samples from zoological gardens were negative, 8 kinkajous from traders were positive for Baylisascaris eggs, at least 4 of which were wild caught in the Republic of Guyana. No associated human illness was reported from any of the facilities. The 2 infected kinkajous received a single oral administration of Milbemycin® A Tablets, which delivers 0.69-0.89 mg/kg milbemycin oxime. Fecal examinations on days 14 and 30 were negative for Baylisascaris eggs. These results demonstrated that milbemycin oxime has possible anthelmintic efficacy against Baylisascaris roundworms in captive kinkajous. We conclude that Baylisascaris infections are highly prevalent in wild-caught kinkajous in Japan and that most of the infected kinkajous were imported from the Republic of Guyana.

Fer tyrosine kinase oligomer mediates and amplifies Src-induced tumor progression.

c-Src is upregulated in various human cancers, suggesting its role in malignant progression. However, the molecular circuits of c-Src oncogenic signaling remain elusive. Here we show that Fer tyrosine kinase oligomer mediates and amplifies Src-induced tumor progression. Previously, we showed that transformation of fibroblasts is promoted by the relocation of c-Src to non-raft membranes. In this study, we identified Fer and ezrin as non-raft c-Src targets. c-Src directly activated Fer by initiating its autophosphorylation, which was further amplified by Fer oligomerization. Fer interacted with active c-Src at focal adhesion membranes and activated Fer-phosphorylated ezrin to induce cell transformation. Fer was also crucial for cell transformation induced by v-Src or epidermal growth-factor receptor activation. Furthermore, Fer activation was required for tumorigenesis and invasiveness in some cancer cells in which c-Src is upregulated. We propose that the Src-Fer axis represents a new therapeutic target for treatment of a subset of human cancers.

Positioning bedridden patients to reduce interface pressures over the sacrum and great trochanter.

OBJECTIVE: In this study, we evaluated the effect of hip-joint rotation on the interface pressure over the sacrum and greater trochanter with a new protocol for positioning of bedridden elderly patients. METHOD: The interface pressure values over the sacrum and greater trochanter in bedridden patients were evaluated. These were collected in the supine position, 90° lateral position, and 30° and 40° laterally inclined positions with external rotation or neutral positioning of the hip joint. Each interface pressure was assessed with a device measuring pressure distribution, after which, the peak pressure index (PPI) was calculated. RESULTS: In the 17 patients examined, the PPI over the sacrum in the supine position was significantly greater than that in other positions. In the 30° and 40° laterally inclined positions, the PPIs over the greater trochanter were significantly lower in the neutral position of the hip joint compared with those in the external rotation position. CONCLUSION: Our findings revealed the effects of hip-joint rotation on the interface pressure for the greater trochanter, possibly due to the increased distance between the greater trochanter and the sacrum caused by neutral position of the hip joint. The results demonstrate that it is to best place the hip joint in a neutral position when the legs are in contact with the bed in order to distribute the pressure over the greater trochanter in the 30° and 40° laterally inclined positions. These results can be applied to the clinical setting to improve patient positioning and decrease pressure ulcers. DECLARATION OF INTEREST: The authors declare that they have no competing financial interests.

Deletion of Rapgef6, a candidate schizophrenia susceptibility gene, disrupts amygdala function in mice.

In human genetic studies of schizophrenia, we uncovered copy-number variants in RAPGEF6 and RAPGEF2 genes. To discern the effects of RAPGEF6 deletion in humans, we investigated the behavior and neural functions of a mouse lacking Rapgef6. Rapgef6 deletion resulted in impaired amygdala function measured as reduced fear conditioning and anxiolysis. Hippocampal-dependent spatial memory and prefrontal cortex-dependent working memory tasks were intact. Neural activation measured by cFOS phosphorylation demonstrated a reduction in hippocampal and amygdala activation after fear conditioning, while neural morphology assessment uncovered reduced spine density and primary dendrite number in pyramidal neurons of the CA3 hippocampal region of knockout mice. Electrophysiological analysis showed enhanced long-term potentiation at cortico-amygdala synapses. Rapgef6 deletion mice were most impaired in hippocampal and amygdalar function, brain regions implicated in schizophrenia pathophysiology. The results provide a deeper understanding of the role of the amygdala in schizophrenia and suggest that RAPGEF6 may be a novel therapeutic target in schizophrenia.

Excess processing of oxidative damaged bases causes hypersensitivity to oxidative stress and low dose rate irradiation.

Ionizing radiations such as X-ray and γ-ray can directly or indirectly produce clustered or multiple damages in DNA. Previous studies have reported that overexpression of DNA glycosylases in Escherichia coli (E. coli) and human lymphoblast cells caused increased sensitivity to γ-ray and X-ray irradiation. However, the effects and the mechanisms of other radiation, such as low dose rate radiation, heavy-ion beams, or hydrogen peroxide (H2O2), are still poorly understood. In the present study, we constructed a stable HeLaS3 cell line overexpressing human 8-oxoguanine DNA N-glycosylase 1 (hOGG1) protein. We determined the survival of HeLaS3 and HeLaS3/hOGG1 cells exposed to UV, heavy-ion beams, γ-rays, and H2O2. The results showed that HeLaS3 cells overexpressing hOGG1 were more sensitive to γ-rays, OH(•), and H2O2, but not to UV or heavy-ion beams, than control HeLaS3. We further determined the levels of 8-oxoG foci and of chromosomal double-strand breaks (DSBs) by detecting γ-H2AX foci formation in DNA. The results demonstrated that both γ-rays and H2O2 induced 8-oxoguanine (8-oxoG) foci formation in HeLaS3 cells. hOGG1-overexpressing cells had increased amounts of γ-H2AX foci and decreased amounts of 8-oxoG foci compared with HeLaS3 control cells. These results suggest that excess hOGG1 removes the oxidatively damaged 8-oxoG in DNA more efficiently and therefore generates more DSBs. Micronucleus formation also supported this conclusion. Low dose-rate γ-ray effects were also investigated. We first found that overexpression of hOGG1 also caused increased sensitivity to low dose rate γ-ray irradiation. The rate of micronucleus formation supported the notion that low dose rate irradiation increased genome instability.

Stakeholder prioritization of zoonoses in Japan with analytic hierarchy process method.

There exists an urgent need to develop iterative risk assessment strategies of zoonotic diseases. The aim of this study is to develop a method of prioritizing 98 zoonoses derived from animal pathogens in Japan and to involve four major groups of stakeholders: researchers, physicians, public health officials, and citizens. We used a combination of risk profiling and analytic hierarchy process (AHP). Profiling risk was accomplished with semi-quantitative analysis of existing public health data. AHP data collection was performed by administering questionnaires to the four stakeholder groups. Results showed that researchers and public health officials focused on case fatality as the chief important factor, while physicians and citizens placed more weight on diagnosis and prevention, respectively. Most of the six top-ranked diseases were similar among all stakeholders. Transmissible spongiform encephalopathy, severe acute respiratory syndrome, and Ebola fever were ranked first, second, and third, respectively.

Gap-junction-mediated communication in human periodontal ligament cells.

Periodontal tissue homeostasis depends on a complex cellular network that conveys cell-cell communication. Gap junctions (GJs), one of the intercellular communication systems, are found between adjacent human periodontal ligament (hPDL) cells; however, the functional GJ coupling between hPDL cells has not yet been elucidated. In this study, we investigated functional gap-junction-mediated intercellular communication in isolated primary hPDL cells. SEM images indicated that the cells were in contact with each other via dendritic processes, and also showed high anti-connexin43 (Cx43) immunoreactivity on these processes. Gap-junctional intercellular communication (GJIC) among hPDL cells was assessed by fluorescence recovery after a photobleaching (FRAP) analysis, which exhibited dye coupling between hPDL cells, and was remarkably down-regulated when the cells were treated with a GJ blocker. Additionally, we examined GJs under hypoxic stress. The fluorescence recovery and expression levels of Cx43 decreased time-dependently under the hypoxic condition. Exposure to GJ inhibitor or hypoxia increased RANKL expression, and decreased OPG expression. This study shows that GJIC is responsible for hPDL cells and that its activity is reduced under hypoxia. This is consistent with the possible role of hPDL cells in regulating the biochemical reactions in response to changes in the hypoxic environment.

A new calpain inhibitor protects left ventricular dysfunction induced by mild ischemia-reperfusion in in situ rat hearts.

We have previously indicated that a new soluble calpain inhibitor, SNJ-1945 (SNJ), attenuates cardiac dysfunction after cardioplegia arrest-reperfusion by inhibiting the proteolysis of α-fodrin in in vitro study. Nevertheless, the in vivo study design is indispensable to explore realistic therapeutic approaches for clinical use. The aim of the present in situ study was to investigate whether SNJ attenuated left ventricular (LV) dysfunction (stunning) after mild ischemic-reperfusion (mI-R) in rat hearts. SNJ (60 µmol/l, 5 ml i.p.) was injected 30 min before gradual and partial coronary occlusion at proximal left anterior descending artery. To investigate LV function, we obtained curvilinear end-systolic pressure-volume relationship by increasing afterload 60 min after reperfusion. In the mI-R group, specific LV functional indices at midrange LV volume (mLVV), end-systolic pressure (ESP(mLVV)), and pressure-volume area (PVA(mLVV): a total mechanical energy per beat, linearly related to oxygen consumption) significantly decreased, but SNJ reversed these decreases to time control level. Furthermore, SNJ prevented the α-fodrin degradation and attenuated degradation of Ca(2+) handling proteins after mI-R. Our results indicate that improvements in LV function following mI-R injury are associated with inhibition of the proteolysis of α-fodrin in in situ rat hearts. In conclusion, SNJ should be a promising tool to protect the heart from the stunning.

Optimum microcurrent stimulation intensity for galvanotaxis in human fibroblasts.

OBJECTIVE: In this study, we develop methods to measure galvanotaxis of fibroblasts and determined the optimum conditions of electrical stimulation. METHOD: An inverted 35mm dish containing cell suspensions (3×105 primary human skin fibroblasts, DMEM, and 10% FBS) was placed on the centre of a 100mm dish. The 35mm dish was removed 24 hours later, and culture medium was added to the 100mm dish. Fibroblasts were randomised (double-blind) into three groups, where electrical stimulation was given at varying intensities: 0UA (control), 50UA, and 100UA. Electrical stimulation (frequency=0.3Hz) was conducted, for a duration of 4 hours, with platinum electrodes in a CO2 incubator. We took pictures immediately before and 20 hours after stimulation. We calculated the migration ratio to the negative pole by dividing the area of attached fibroblasts after stimulation with that before stimulation. RESULTS: The migration ratio to the negative pole was significantly higher in the 100UA group than in the control group (p<0.05). The ratios were 0.902±0.292 in the control group, 1.128±0.253 in the 50UA group, and 1.24±0.300 in the 100UA group. CONCLUSION: This study observed the change in cell proliferation during the initial 24-hour period after plating and was thus able to quantitatively evaluate the migration. The results suggest that a low-intensity direct current promotes migration to the negative pole of human dermal fibroblasts, which is charged with positive electricity. Several clinical reports using the methods in this study showed the microcurrent efficacy for pressure ulcer healing. Electrical stimulation based on our in vitro experiment might be important for the development of physical therapy for pressure ulcers.

Structural and functional analyses of chicken liver ferritin.

Characterization of ferritins from different species has provided insight into iron regulation mechanisms and evolutionary relationships. Here, we examined chicken liver ferritin, which comprises only H subunit and has 14.8 µg of Fe/100 µg of protein. The chicken H subunit apo homopolymer showed the same iron uptake rate as bovine H subunit homopolymer expressed with a baculovirus expression system (0.31 and 0.28 mmol of Fe/min per micromole of protein for chicken and bovine H subunit, respectively). Chicken H subunit apo homopolymer showed a significantly higher biotinylated hemin-binding activity than liver holoferritin. Although bovine spleen apoferritin, which has an L (liver or light):H (heart or heavy) subunit ratio of 1:1, also shows a significantly higher biotinylated hemin-binding activity than its holoferritin, these biotinylated hemin-binding activities were markedly lower than those of both chicken holo- and apoferritins. Binding of chicken holo- and apoferritin with biotinylated hemin was strongly inhibited by hemin but not iron-free hemin, protoporphyrin IX, or Zn-protoporphyrin. These findings demonstrate that chicken ferritin comprises only an H subunit, possesses ferroxidase activity as in mammalian ferritin H subunits, and binds heme more strongly than mammalian ferritins.

Interactions between canine RAD51 and full length or truncated BRCA2 BRC repeats.

In humans, mutations in the gene for the breast cancer susceptibility protein BRCA2 affect its interactions with the recombinase RAD51 and are associated with an increased risk of cancer. This interaction occurs through a series of eight BRC repeat sequences in BRCA2. A mammalian two-hybrid assay using individual BRC repeats demonstrated that BRC6 did not bind to RAD51, whereas there was strong (BRC1, 2 and 4), intermediate (BRC8), or weak (BRC3, 5 and 7) binding of other BRC repeats to RAD51. In serial deletion mutation experiments, binding strengths were increased when the C-terminal BRC repeat was removed from BRC1-8, BRC1-5 and BRC1-3. These results may provide an insight into the effects of missense or truncation mutations in BRCA2 in canine tumours.

Evaluation of the combined use of ultrasound irradiation and wound dressing on pressure ulcers.

OBJECTIVE: To evaluate the effect of ultrasound irradiation when used alongside standard care in the treatment of pressure ulcers; outcome measures were reduction in wound size and exudate weight. METHOD: Five patients (two male and three female, age range: 76-92 years) with seven ulcers participated in this study. They had National Pressure Ulcer Advisory Panel (NPUAP) stage III or IV pressure ulcers. We conducted an ABABA study (A: standard treatment with dressings that promote a moist wound healing environment; B: ultrasound irradiation administered to the pressure ulcer through the same dressing used in period A; each period lasted 2-4 weeks). Six ulcers each were randomised to either the treatment group or control group. One ulcer was not randomised, but was the first to receive ultrasound in the BABA sequence, with a view to determining if the pilot was feasible. The control group received sham ultrasound in period B. Pulsed ultrasound (20% duty cycle, 0.5W/cm2 on the wound surface, 1MHz or 3MHz, for 10 minutes) was applied five times weekly. RESULTS: In the treatment group, two ulcers markedly decreased in size after 3-4 weeks of US treatment, one ulcer decreased in size soon after initiation of treatment and one ulcer showed no clear reduction in size. The volume of exudate was greater in period B than A in two ulcers that reduced markedly in size after 3-4 weeks of US treatment. None of the ulcers in the control group decreased markedly in size. CONCLUSION: This pilot study suggests that US used alongside standard treatment might promote the healing of pressure ulcers. However, larger studies are required to determine the efficacy and mechanism of US treatment for PUs. DECLARATION OF INTEREST: None.

Optimization of a prominent oxygen-permeable device for pancreatic islets.

BACKGROUND: We have demonstrated a culture bag system that is useful for pancreatic islet transplantation. To improve and simplify islet transplantation procedures from culture to transplantation, we developed a novel device specific for both islet culture and transplantation (TUBERO Device [TD]) using an oxygen-permeable material. MATERIALS AND METHODS: Porcine islets with 30 minutes warm ischemia time were cultured for 24 hours at 37 degrees C in 5% CO2 and humidified air under three different procedures: (1) ordinary culture flask, (2) culture bag suitable for platelets, and (3) TD. Loss of islets during culture, glucose-stimulated insulin release as an islet functional test, and ADP/ATP ratio as an index of islet viability tests were evaluated to compare the devices. TD was further applied in two clinical islet transplantations using non-heart-beating donors in Japan. RESULTS: The loss of islets during culture was considerably lower in the TD group. The stimulation index upon glucose challenge tests was significantly higher in the TD group than the others. The ADP/ATP ratio in TD group was significantly lower than that in the ordinary flask group, suggesting that the apoptotic islets were relatively lower among TD. Most importantly, TD was successfully applied both in the clinical islet cultures and in transplantation, resulting in excellent graft function. CONCLUSIONS: We propose that the TD, a novel product, not only simplifies islet transplantation procedures, but also maintain the quality of isolated islets.

Prevalence of herpes B virus genome in the trigeminal ganglia of seropositive cynomolgus macaques.

Herpes B virus infection is almost asymptomatic in macaques (Macaca spp.), which are the natural hosts of this pathogen, but is the cause of high mortality in humans. Reactivation of the latent virus in the trigeminal ganglia (TG) results in the shedding of infectious particles into the oral mucosal membrane. Saliva contaminated with the reactivated virus from the ganglia of the natural host is considered to be important for viral transmission to humans and other monkeys. In the present study, we investigated the prevalence of the herpes B virus genome in the left and right TG of seropositive asymptomatic cynomolgus macaques. The latent virus genome was detected using a polymerase chain reaction and microplate hybridization assay. We found that the virus DNA was present in one or both TG of 12 of the 30 macaques (40%) tested, with the virus being detected from both TG in five of the 12 macaques and from a single TG in the remaining seven.

Reduced tau expression in gastric cancer can identify candidates for successful Paclitaxel treatment.

A recent study disclosed that breast cancer cases with low 'tau' expression can predict susceptibility to Paclitaxel administration. In the current study, the clinical significance of tau expression in gastric cancer cases was established by identifying candidates with Paclitaxel administration. Tissue specimens from 20 cases of in-operable or noncuratively resected gastric cancer were examined. Subsequent to the administration of 80 mg m(-2) of Paclitaxel in six 3-h intravenous infusions, the clinical effectiveness of Paclitaxel was evaluated by the size of metastatic lesions with computed tomography. The status of the tau expression was determined by immunohistochemistry. Based on a previously reported classification scheme, six were classified as tau-negative expression (0, 1+) cases and 14 were classified as tau-positive expression (2+, 3+) cases. All six (100%) cases of tau-negative expression showed a favourable response (partial response or minor response) to Paclitaxel administration. However, 12 (86%) of the 14 cases of tau-positive expression showed progressive disease (n = 11) or no change (n = 1) after Paclitaxel administration. The serum carcinoembryonic antigen values of the six cases of tau-negative expression were markedly decreased in comparison to the 14 tau-positive cases. These data indicate that tau-negative expression can be used to select gastric cancer patients, which will favourably respond to Paclitaxel treatment.