The influence of CYP2C19 polymorphisms on exacerbating effect of rabeprazole in celecoxib-induced small bowel injury.

BACKGROUND: Simultaneous use of proton pump inhibitors (PPIs) has been shown to increase the risk of nonsteroidal anti-inflammatory drug (NSAID)-induced small bowel injury. AIM: To investigate whether polymorphisms of the cytochrome P450 2C19 gene (CYP2C19), encoding a key metabolising enzyme for PPIs, are associated with small bowel injury induced by celecoxib in combination with the PPI rabeprazole. METHODS: Study participants included 55 healthy Japanese volunteers, who participated in the PPI-NSAID Kyushu University Study using video capsule endoscopy. For 2 weeks, 26 subjects were treated with celecoxib plus rabeprazole (rabeprazole group), and 29 subjects received celecoxib plus placebo (placebo group). All subjects were genotyped for CYP2C19 using real-time fluorescent polymerase chain reaction. Subjects were sub-classified as poor metabolizers or extensive metabolizers. The incidence and number of small bowel injuries were compared between poor metabolizers and extensive metabolizers in each group. RESULTS: In the rabeprazole group, the incidence of small bowel injuries was significantly higher in poor metabolizers than in extensive metabolizers (85.7% vs 31.6%, P=.026). The number of mucosal injuries in the rabeprazole group was also significantly higher in poor metabolizers compared with extensive metabolizers (median [range] 3 [0-31] vs 0 [0-7], P=.01). In addition, we found a significant interaction between CYP2C19 genotype and concomitant use of rabeprazole in subjects at risk for celecoxib-induced small bowel injury. CONCLUSIONS: The CYP2C19 genotype might be associated with the risk of small bowel injury when celecoxib is combined with rabeprazole.

Disulfide bond formation of thiols by using carbon nanotubes.

Clarification of the interactions between carbon nanotubes (CNTs) and proteinogenic amino acids is a key approach to understanding CNT-protein interactions. Previous studies have addressed the mechanism of the physical adsorption of amino acids onto CNTs. However, little is known about their chemical reactions in aqueous solutions. Here, we established dispersant-free systems to clarify intrinsic CNT-thiol interactions. We demonstrated that the redox reaction of CNTs with cysteine, containing a thiol group, leads to disulfide bond formation between cysteine molecules, even under acidic conditions. The generality of the redox reaction is validated using other thiols such as dithiothreitol and glutathione. The present results suggest that structures of proteins and peptides containing free thiol groups are chemically modified and misfolded on CNT surfaces by this disulfide bond formation in biological systems.

Nisin A extends the shelf life of high-fat chilled dairy dessert, a milk-based pudding.

AIMS: The aims of this study were to evaluate the effectiveness of nisin A to control the growth of spore-forming bacteria, Bacillus and Paenibacillus, in chilled high-fat, milk pudding and to reduce heat treatment to improve aroma and flavour. METHODS AND RESULTS: Nisin A was added to milk pudding containing 5·0 and 7·5% fat to final concentrations of 40, 80, 120 and 240 IU ml(-1). Spores from Bacillus thuringiensis, Bacillus cereus and Paenibacillus jamilae were inoculated into samples at 10 spores ml(-1) prior to pasteurization at 130°C for 2 s. Milk pudding without inoculation was pasteurized using less heat condition (100, 110 and 120°C for 2 s) to measure the effect of adjusting the ingredients to prevent naturally occurring bacteria. The viable cells during storage at 15, 20 and 30°C showed nisin A inhibited spiked bacteria to varying degrees depending on species, sensitivities to nisin A concentration and fat content, and inhibited natural populations at 80 IU g(-1) nisin A in 5·0% fat and at 120 IU g(-1) in 7·5% fat milk pudding. An aroma compound analysis and organoleptic assessment showed processing at 110 and 120°C decreased the temperature-dependent unpleasant odours, for example, reduced dimethyl sulfide and dimethyl disulfide by 1·2-1·5 times and increased rankings in taste tests compared with 130°C treated pudding. CONCLUSIONS: Nisin A was found to be effective as a natural preservative to control spoilage bacteria in high-fat milk pudding and extend its shelf life, when using reduced heat treatments to improve the flavour and aroma without compromising food safety. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report showing nisin A is effective in reducing spoilage bacteria in high-fat, chilled dessert, milk pudding. Therefore, nisin A can be used to improve milk puddings to satisfy both industry and consumer demand for food quality and safety.

Phase stability of a garnet-type lithium ion conductor Li7La3Zr2O12.

The phase stability of Li7La3Zr2O12 (LLZ) was investigated using high temperature X-ray diffraction (HT-XRD). An Al-free tetragonal LLZ phase transformed into a non-quenchable cubic phase around 650 °C. The phase transformation process between the tetragonal phase and the new cubic phase showed perfect reversibility. The thermal analysis showed a pair of endothermic and exothermic peaks around 640 °C that is in good agreement with the phase transformation process observed in the HT-XRD study. The non-quenchable high temperature cubic phase showed high ionic conductivity with extraordinarily low activation energy (0.117 eV). The tetragonal phase showed another phase transformation to a low temperature (LT) cubic phase around 150-200 °C in air by absorbing CO2 into the structure. The preferred temperature for the CO2 absorption process was around 200 °C and the absorbed CO2 was extracted once the temperature reached 450 °C or above resulting in the phase transformation back to the tetragonal phase. On the other hand the high temperature (HT) cubic phase which shows high ionic conductivity was stabilized by Al substitution. A Li-poor LLZ containing impurity phases such as La2Zr2O7 and La2O3 effectively reacted with γ-Al2O3 resulting in the formation of a pure Al-stabilized cubic LLZ, while the stoichiometric LLZ took a much longer time to complete the Al-substitution. The result suggested that the formation of Li vacancies is the primary step in the formation of the Al-stabilized cubic phase.

Hiatal hernia following total gastrectomy with Roux-en-Y reconstruction.

Hiatal hernias after total gastrectomy for advanced gastric cancer are very rare. We review a case of a 44-year-old male who presented with dyspnea and chest pain 2 days after total gastrectomy, lower esophagectomy, and splenectomy with retrocolic Roux-en-Y reconstruction approached by a left thoracoabdominal incision for gastric cancer at the cardia. Plain and cross-sectional imaging identified a large hiatal hernia protruding into the right thorax containing left-sided transverse colon and small intestine. Our patient underwent a laparotomy, and after hernia reduction the hiatal defect was repaired by direct suturing. He experienced anastomotic leakage and right pyothorax, but recovered. The potential cause is discussed here and the published literature on this rare complication is reviewed briefly.

Role of c-Jun N-terminal kinase isoforms in the cellular activity of melanoma cell lines.

BACKGROUND: The c-Jun N-terminal kinase (JNK) is thought to be involved in inflammation, proliferation and apoptosis. AIM: To examine the role of JNK isoforms in metastasis, proliferation, migration and invasion of the malignant melanoma (MM) cell lines SK-MEL-28, SK-MEL-3 and WM164, using a kinase-specific inhibitor or isoform-specific small interfering (si)RNAs. RESULTS: SK-MEL-3, a cell line established from metastatic MM, showed slightly increased phosphorylation of both JNK1 and JNK2, whereas WM164, a cell line derived from primary MM, showed significant phosphorylation of JNK1. A JNK inhibitor, SP600125, inhibited cell proliferation of SK-MEL-3 but not SK-MEL-28 or WM164. Transfection of JNK1-specific siRNA reduced the migratory activity of WM164 cells, while silencing of either JNK1 or JNK2 strongly suppressed the invasive activity of SK-MEL-3. CONCLUSIONS: Our study suggests that JNK isoforms have different roles in MM. Metastasis of MM may be regulated by JNK2, while invasion is regulated by both JNK1 and JNK2. JNK1 and JNK2 respectively mediate cell migration and cell proliferation. Further understanding of the specific roles of JNK isoforms in the pathogenesis of MM may lead to the development of therapies targeting specific isoforms.

Axonal protection by Nmnat3 overexpression with involvement of autophagy in optic nerve degeneration.

Axonal degeneration often leads to the death of neuronal cell bodies. Previous studies demonstrated the crucial role of nicotinamide mononucleotide adenylyltransferase (Nmnat) 1, 2, and 3 in axonal protection. In this study, Nmnat3 immunoreactivity was observed inside axons in the optic nerve. Overexpression of Nmnat3 exerts axonal protection against tumor necrosis factor-induced and intraocular pressure (IOP) elevation-induced optic nerve degeneration. Immunoblot analysis showed that both p62 and microtubule-associated protein light chain 3 (LC3)-II were upregulated in the optic nerve after IOP elevation. Nmnat3 transfection decreased p62 and increased LC3-II in the optic nerve both with and without experimental glaucoma. Electron microscopy showed the existence of autophagic vacuoles in optic nerve axons in the glaucoma, glaucoma+Nmnat3 transfection, and glaucoma+rapamycin groups, although preserved myelin and microtubule structures were noted in the glaucoma+Nmnat3 transfection and glaucoma+rapamycin groups. The axonal-protective effect of Nmnat3 was inhibited by 3-methyladenine, whereas rapamycin exerted axonal protection after IOP elevation. We found that p62 was present in the mitochondria and confirmed substantial colocalization of mitochondrial Nmnat3 and p62 in starved retinal ganglion cell (RGC)-5 cells. Nmnat3 transfection decreased p62 and increased autophagic flux in RGC-5 cells. These results suggest that the axonal-protective effect of Nmnat3 may be involved in autophagy machinery, and that modulation of Nmnat3 and autophagy may lead to potential strategies against degenerative optic nerve disease.

Autologous adipose-derived regenerative cells are effective for chronic intractable radiation injuries.

Effective therapy for chronic radiation injuries, such as ulcers, is prone to infection. Stiffness is expected since the therapeutic radiation often involves wider and deeper tissues and often requires extensive debridement and reconstruction, which are not sometimes appropriate for elderly and compromised hosts. Autologous adipose-derived regenerative cells (ADRCs) are highly yielding, forming relatively elderly aged consecutive 10 cases, 63.6±14.9 y (52-89 y), with mean radiation dose of 75.0±35.4 Gy (50-120 Gy) were included with at least 10-month follow-up. Minimal debridement and ADRC injection in the wound bed and margin along with the injection of mixture of fat and ADRCs in the periphery were tested for efficacy and regenerated tissue quality by clinically as well as imaging by computed tomography and magnetic resonance imaging. Uncultured ADRCs of 1.6±1.3×10(7) cells were obtained. All cases healed uneventfully after 6.6±3.2 weeks (2-10 weeks) post-operatively. The done site morbidity was negligible and without major complications, such as paralysis or massive haematoma. The regenerated tissue quality was significantly superior to the pre-operative one and the mixture of fat and ADRCs connected to the intact tissue was very soft and pliable. Mean follow-up at 1.9±0.8 y (0.9-2.9 y) revealed no recurrence or new ulceration after treatment. Thus, the ADRCs treatment for decades-long radiation injuries is effective, safe and improves the quality of wounds.

Advances in characterization of neuroprotective peptide, humanin.

Humanin (HN), a short amino acid peptide, protects neurons as well as other cells from amyloid ß-induced toxicities and other stresses. A number of HN binding proteins have been identified and their involvements in HN-mediated neuroprotection have been suggested in some cases. However, the way HN binds to the target molecules has never been clarified. Here we will review the structures of HN and HN analogs in solution as a function of solvent conditions and attempt to relate their structural characteristics to the functional properties.

HtrA2/Omi-immunoreactive intraneuronal inclusions in the anterior horn of patients with sporadic and Cu/Zn superoxide dismutase (SOD1) mutant amyotrophic lateral sclerosis.

AIMS: HtrA2/Omi is a mitochondrial serine protease that promotes the apoptotic processes, but the relationship between HtrA2/Omi and amyotrophic lateral sclerosis (ALS) is still unknown. The purpose of the present study was to determine whether abnormal expression of HtrA2/Omi occurs in patients with ALS. METHODS: We prepared autopsied spinal cord tissues from 7 control subjects, 11 patients with sporadic ALS (SALS) and 4 patients with Cu/Zn superoxide dismutase (SOD1)-related familial ALS (FALS). We then performed immunohistochemical studies on HtrA2/Omi using formalin-fixed, paraffin-embedded sections from all of the cases. RESULTS: In the control subjects, the anterior horn cells were mildly to moderately immunostained with HtrA2/Omi. In the patients with SALS, strong HtrA2/Omi immunoreactivity was found in some skein-like inclusions and round hyaline inclusions as well as many spheroids, but Bunina bodies were immunonegative for HtrA2/Omi. In the patients with SOD1-related FALS, Lewy body-like hyaline inclusions were observed in three cases and conglomerate inclusions were observed in the remaining case, and both types of inclusions were intensely immunopositive for HtrA2/Omi. CONCLUSIONS: These results suggest that abnormal accumulations of HtrA2/Omi may occur in several types of motor neuronal inclusions in the anterior horn from SALS and SOD1-linked FALS cases, and that HtrA2/Omi may be associated with the pathogenesis of both types of ALS.

Potential application of arginine in interaction analysis.

Aqueous solution of 0.1-2 M arginine at mildly acidic to neutral pH is widely used in biotechnology and protein research, including protein refolding, purification, and formulation. This is largely because of its ability to suppress non-specific protein-protein and protein-surface interactions. Here we propose potential applications of arginine in interaction analysis for proteins. One of the important goals of such analysis is discovery of small molecule antagonistic or agonistic ligands that bind to target proteins and thereby modulate their function. Such research is often hampered by the low solubility of the small molecules, the instability of target proteins and the non-specific protein-ligand interactions. Aqueous arginine solution increases the solubility of small molecules, which should give an alternative to conventional dissolution method of small molecules by organic solvents. Arginine may also directly impact on the analysis of protein-protein or protein-ligand interactions by suppressing weak non-specific interactions.

Anticancer drugs up-regulate HspBP1 and thereby antagonize the prosurvival function of Hsp70 in tumor cells.

The 70-kDa heat shock protein (Hsp70) is up-regulated in a wide variety of tumor cell types and contributes to the resistance of these cells to the induction of cell death by anticancer drugs. Hsp70 binding protein 1 (HspBP1) modulates the activity of Hsp70 but its biological significance has remained unclear. We have now examined whether HspBP1 might interfere with the prosurvival function of Hsp70, which is mediated, at least in part, by inhibition of the death-associated permeabilization of lysosomal membranes. HspBP1 was found to be expressed at a higher level than Hsp70 in all normal and tumor cell types examined. Tumor cells with a high HspBP1/Hsp70 molar ratio were more susceptible to anticancer drugs than were those with a low ratio. Ectopic expression of HspBP1 enhanced this effect of anticancer drugs in a manner that was both dependent on the ability of HspBP1 to bind to Hsp70 and sensitive to the induction of Hsp70 by mild heat shock. Furthermore, anticancer drugs up-regulated HspBP1 expression, whereas prevention of such up-regulation by RNA interference reduced the susceptibility of tumor cells to anticancer drugs. Overexpression of HspBP1 promoted the permeabilization of lysosomal membranes, the release of cathepsins from lysosomes into the cytosol, and the activation of caspase-3 induced by anticancer drugs. These results suggest that HspBP1, by antagonizing the prosurvival activity of Hsp70, sensitizes tumor cells to cathepsin-mediated cell death.

Clinicopathologic investigation of a family with expanded SCA8 CTA/CTG repeats.

We investigated a family manifesting progressive ataxia, with expanded SCA8 CTA/CTG repeats. Neuropathologically, degeneration of Purkinje, inferior olivary, and nigral neurons and periaqueductal gliosis were evident. The sites of Purkinje cell loss were occupied by fibrillary accumulations. The remaining Purkinje cells showed somatic sprouts, and intracytoplasmic 1C2-positive granular structures were recognizable. This characteristic distribution of neurodegeneration and Purkinje cell cytopathology were distinct from those of other hereditary spinocerebellar ataxias previously reported.

S-carboxymethylcysteine normalises airway responsiveness in sensitised and challenged mice.

S-carboxymethylcysteine (S-CMC) has been used as a mucoregulator in respiratory diseases. However, the mechanism of action of S-CMC on allergic airway inflammation has not yet been defined. In the present study, BALB/c mice were initially sensitised and challenged to ovalbumin (OVA) and, weeks later, re-challenged with OVA (secondary challenge). S-CMC (5-100 mg.kg-1) was administered from 2 days before the secondary challenge through to the day of assay. Mice developed airway hyperresponsiveness (AHR) 6 h after the secondary challenge and increased numbers of neutrophils were present in the bronchoalveolar lavage (BAL) fluid. At 72 h after secondary challenge, mice again developed AHR, but the BAL fluid contained large numbers of eosinophils. S-CMC treatment was found to reduce AHR and neutrophilia at 6 h, as well as eosinophilia and AHR at 72 h. These effects appeared to be dose dependent. Goblet cell hyperplasia, observed at 72 h, was reduced by S-CMC. In BAL fluid, increased levels of interferon-gamma, interleukin (IL)-12 and IL-10 and decreased levels of IL-5 and IL-13 were detected. In conclusion, the data indicate that S-carboxymethylcysteine is effective in reducing airway hyperresponsiveness and airway inflammation at two distinct phases of the response to the secondary allergen challenge in sensitised mice.

Expression of a CD44 variant and VEGF-C and the implications for lymphatic metastasis and long-term prognosis of human breast cancer.

The aim of the present study was to determine whether expression of the CD44 variant v7-v8 (CD44v7-v8) or vascular endothelial growth factor-C (VEGF-C) is associated with long-term prognosis in breast cancer patients. A 10-year follow-up of 91 patients with primary breast cancer who were previously assessed for CD44 expression was undertaken. Immunohistochemical evaluation of VEGF-C expression was performed in 87 of these patients and their long-term prognosis was assessed. The disease-free and overall survival rates were significantly poorer for the CD44v7-v8-positive patients than for the patients negative for this marker. VEGF-C expression was detected in 38 out of the 87 patients (43.7%) with primary human breast cancer. There were no significant differences in tumor size, histological type, axillary lymph node status, presence of lymphatic or venous invasion, or presence of estrogen receptors and progesterone receptors between the VEGF-C-positive and -negative patients. There were also no significant differences in the disease-free or overall survival rates in these patient groups. In conclusion after the 10-year follow-up, expression of CD44v7-v8 was associated with poor prognosis for breast cancer patients. However, there was no association between VEGF-C expression and the clinicopathological factors or prognosis of breast cancer patients.

Frontotemporal and motor neurone degeneration with neurofilament inclusion bodies: additional evidence for overlap between FTD and ALS.

We present the case of a patient who had clinical frontal lobe dementia without apparent motor neurone disease (MND), with pathologic findings not typical of any single currently classified frontotemporal degeneration (FTD). At autopsy, the brain had frontal and temporal atrophy with neuronal loss, gliosis, and superficial spongiosis, typical of all FTDs. There were at least three different morphologic types of intracytoplasmic neuronal inclusions in a variety of brain and brainstem regions, including the hippocampal dentate gyrus and pyramidal neurones, the neocortex (in particular, the motor cortex), basal ganglia, thalamus, subthalamic nucleus, basis pontis, and inferior olivary nuclei. Inclusions had the morphologies of Pick-like bodies, pleomorphic inclusions, and hyaline conglomerate (HC)-like inclusions. None of these were positive with tau immunostains. Pick-like bodies in the dentate gyrus were labelled with ubiquitin. The pleomorphic inclusions in the neocortex and dentate gyrus and the HC-like inclusions in the motor and parietal cortex were strongly positive with immunostains for neurofilament. We discuss the differential diagnosis and compare this case with those disorders to which it is most similar. In particular, we compare the unique neurofilament-positive inclusions to the inclusions of FTD-MND, to Pick bodies, and to the basophilic and HC inclusions that are occasionally seen in amytrophic lateral sclerosis (ALS). Although FTD-MND may be found in ALS, the findings in this case may have additional implications for a link between FTD and ALS.