IL-4 mediated by HSV vector suppresses morphine withdrawal response and decreases TNFα, NR2B, and pC/EBPß in the periaqueductal gray in rats.

Chronic opiates induce the development of physical dependence. Opioid physical dependence characterized by withdrawal symptoms, may have very long-lasting effects on the motivation for reward, including the incubation of cue-induced drug-seeking behavior. Elucidation of the mechanisms involved in physical dependence is crucial to developing more effective treatment strategies for opioid dependence. Chronic morphine induces production of proinflammatory cytokines in regional-specific sites of the brain. Interleukin-4 (IL-4) is a prototypical anti-inflammatory cytokine that globally suppresses proinflammatory cytokines. Here, we used recombinant herpes simplex virus vector S4IL4 that encode mouse il4 gene to evaluate the therapeutic potential of IL-4 in naloxone-precipitation morphine withdrawal (MW). One week after microinjection of the vector S4IL4 into the PAG LacZ or mouse IL-4 immunoreactivity in the vlPAG was visualized. ELISA assay showed that vector S4IL4 into the PAG induced the expression of IL-4. S4IL4 blunted the morphine withdrawal syndrome. S4IL4 suppressed the upregulated TNFα, NR2B and pC/EBPß in the PAG induced by MW. These results show that inhibition of proinflammatory factor in the PAG suppressed MW. This study may provide a novel therapeutic approach to morphine physical withdrawal symptoms.

Cephalometric evaluation after two-stage palatoplasty combined with a Hotz plate: a comparative study between the modified Furlow and Widmaier-Perko methods.

The effects on craniofacial growth of two different soft palate repair techniques in two-stage palatoplasty were investigated. This was a retrospective, cross-sectional cohort study of 68 children with non-syndromic, complete unilateral cleft lip and palate. Thirty-four patients were treated with the modified Furlow method (F-group) and the remaining 34 with the Widmaier-Perko method (P-group). Craniofacial growth was assessed by analyzing 12 angular and 12 linear measurements on lateral cephalograms. Composite facial diagrams from the two groups were compared with those of a control non-cleft group. Angular and linear measurements did not differ significantly between the two groups, implying that the craniofacial morphology was not affected by the difference in soft palate repair technique. However, small differences in anterior nasal spine and posterior nasal spine were found in cleft patients compared with controls. These findings suggest that the modified Furlow and Widmaier-Perko methods have a similar impact on craniofacial growth. Considering speech function, the modified Furlow method provides better craniofacial growth and speech function. However, the long-term effects of both methods on craniofacial growth after growth cessation remain to be determined.

Early and late outcomes of inflammatory abdominal aortic aneurysms: comparison with the outcomes after open surgical and endovascular aneurysm repair in literature reviews.

AIM: The aim of this study was to analyze the clinical characteristics, operative management and early and late outcomes of Inflammatory abdominal aortic aneurysms (IAAAs) in our vascular service, retrospectively and to compare with the outcomes of open and endovascular aneurysm repair (EVAR) in the patients with IAAAs in literature reviews. METHODS: Between January 1990 and December 2011, this series included twenty-four patients with IAAAs who underwent surgical treatment. The mean length of follow-up was 37.6 months (range 3-108). Kaplan-Meier life tables were used to calculate survival rate. RESULTS: Fifteen patients (60%) were symptomatic including 3 patients with hydronephrosis. Seven patients with severe symptoms underwent emergency operations. Twenty four patients underwent open surgical AAA repair. The 30 - day mortality rates were 0%. All patients had successful post operative courses and followed as outpatients in an elective situation. The ten year cumulative survival rate was 70%. The early and late mortality was the same for open repair compared to non-inflammatory AAA. CONCLUSIONS: IAAA compared with non-IAAA is associated with a higher incidence of preoperative morbidity, however operative mortality rates are lower. EVAR has lower early operative mortality rates, however, there are some problems such as remaining periaortic fibrosis and hydronephrosis in approximately half of IAAA patients after EVAR.

Palmar reconstruction of the triangular fibrocartilage complex for instability of the distal radioulnar joint: a biomechanical study.

We developed a new triangular fibrocartilage complex reconstruction technique for distal radioulnar joint instability in which the palmar portion of the triangular fibrocartilage complex was predominantly reconstructed, and evaluated whether such reconstruction can restore stability of the distal radioulnar joint in seven fresh cadaver upper extremities. Distal radioulnar joint instability was induced by cutting all soft-tissue stabilizers around the distal ulna. Using a palmar approach, a palmaris longus tendon graft was sutured to the remnant of the palmar radioulnar and ulnocarpal ligaments. The graft was then passed through a bone tunnel created at the fovea and was sutured. Loads were applied to the radius, and dorsopalmar displacements of the radius relative to the ulna were measured using an electromagnetic tracking device in neutral rotation, 60° supination and 60° pronation. We compared the dorsopalmar displacements before sectioning, before reconstruction and after reconstruction. Dorsopalmar instability produced by sectioning significantly improved in all forearm positions after reconstruction.

Evaluation of skin cancer chemoprevention potential of sunscreen agents using the Epstein-Barr virus early antigen activation in vitro assay.

In our continuing search for novel cancer chemopreventive compounds of natural and synthetic origin, we have evaluated 14 commonly used ultraviolet (UV) sunscreen agents (designated UV-1 to UV-14) for their skin cancer chemoprevention potential. They belong to 8 different chemical categories: aminobenzoate (UV-5, UV-7, UV-8 and UV-14), benzophenone (UV-1, UV-2, UV-3 and UV-13), benzotriazole (UV-10), benzyloxyphenol (UV-9), cinnamate (UV-6), quinolone (UV-4), salicylate (UV-11) and xanthone (UV-12). In the in vitro assay employed, the sunscreens were assessed by their inhibition of the Epstein-Barr virus early antigen (EBV-EA) activation induced by the tumour promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) in human lymphoblastoid Raji cells. All sunscreens tested were found to exhibit anti-tumour promoting activity: listed in decreasing order, moderate (UV-11, UV-2, UV-7, UV-12, UV-3, UV-9 and UV-14) to weak (UV-1, UV-6, UV-8, UV-16, UV-5, UV-4 and UV-10) with octyl salicylate (UV-11) as the most potent and drometrizole (UV-10) as the least potent among the compounds evaluated. A plausible relationship between the antioxidant property of sunscreens and their ability to promote anti-tumour activity was noted. The results call for a comprehensive analysis of skin cancer chemoprevention potential of currently used UV sunscreen agents around the globe to identify those with the best clinical profile.

Effects of transglucosidase on diabetes, cardiovascular risk factors and hepatic biomarkers in patients with type 2 diabetes: a 12-week, randomized, double-blind, placebo-controlled trial.

In this 12-week, randomized, double-blind, placebo-controlled trial, the efficacy and safety of transglucosidase (TGD) were compared with placebo in patients with type 2 diabetes mellitus (T2DM). At 12 weeks, TGD 300 mg/day and TGD 900 mg/day significantly reduced HbA1c (0.18 and 0.21%) and insulin concentration (19.4 and 25.0 pmol/l), respectively, vs. placebo. TGD 300 mg/day and TGD 900 mg/day also significantly reduced low-density lipoprotein cholesterol (0.22 and 0.17 mmol/l, respectively). TGD 900 mg/day significantly reduced triglyceride by 0.24 mmol/l and diastolic blood pressure by 8 mmHg. Placebo was associated with a significant increase from baseline in body mass index, alanine aminotransferase and aspartate aminotransferase (0.17 kg/m(2) , 3 and 2 U/l, respectively), whereas TGD was not. TGD 300 mg/day significantly increased high-molecular-weight adiponectin by 0.6 µg/ml. Adverse events did not differ significantly between the groups. TGD resulted in lowering of HbA1c and blood insulin level and improvements in metabolic and cardiovascular risk factors in T2DM.

X-ray waveguide mode in resonance with a periodic structure.

We present a novel concept for x-ray waveguiding based on electromagnetism in photonic crystals, using a waveguide consisting of a pair of claddings sandwiching a core with a periodic structure. By confining the x rays undergoing multiple interference in the core by total reflection, a characteristic waveguide mode whose field distribution matches the periodicity of the core is formed. The distinctively low propagation loss enables the single-mode propagation of x rays. This concept opens broad application possibilities in x-ray physics from coherent imaging to x-ray quantum optics.

Complication of adenoid cystic carcinoma and sialolithiasis in the submandibular gland: report of a case and its etiological background.

The authors report a case of adenoid cystic carcinoma (ACC) complicated with sialolithiasis of the submandibular gland. The patient was a 43-year-old female with a history of papillotubular carcinoma of the breast almost at the same time. She had noticed a swelling in her sublingual area for 10 years, which was later diagnosed by her dentist to be due to a sialolith in the left submandibular gland. After several years of observation, the patient was referred to have her left submandibular gland, containing the stone, surgically removed with a diagnosis of atrophic sialadenitis. Histopathologically, the submandibular gland was extensively replaced with fibrous granulation tissue, in which there were small but invasive foci of ACC. The present case indicates that ACC could arise in the background of chronic sialadenitis. It is suggested that long-standing sialadenitis cases should be carefully examined to exclude suspicion of malignancy before surgery.

Expression and maturation of Sendai virus vector-derived CFTR protein: functional and biochemical evidence using a GFP-CFTR fusion protein.

Sendai virus (SeV) vector has been shown to efficiently transduce airway epithelial cells. As a precursor to the potential use of this vector for cystic fibrosis (CF) gene therapy, the correct maturation of the SeV vector-derived CF transmembrane conductance regulator (CFTR) protein was examined using biochemical and functional analyses. We constructed a recombinant SeV vector, based on the fusion (F) gene-deleted non-transmissible SeV vector, carrying the GFP-CFTR gene in which the N terminus of CFTR was fused to green fluorescence protein (GFP). This vector was recovered and propagated to high titers in the packaging cell line. Western blotting using an anti-GFP antibody detected both the fully glycosylated (mature) and the core-glycosylated (immature) proteins, indicating that SeV vector-derived GFP-CFTR was similar to endogenous CFTR. We also confirmed the functional channel activity of GFP-CFTR in an iodide efflux assay. The efficient expression of GFP-CFTR, and its apical surface localization, were observed in both MDCK cells in vitro, and in the nasal epithelium of mice in vivo. We concluded that recombinant SeV vector, a cytoplasmically maintained RNA vector, is able to direct production of a correctly localized, mature form of CFTR, suggesting the value of this vector for studies of CF gene therapy.

Sendai virus-mediated CFTR gene transfer to the airway epithelium.

The potential for gene therapy to be an effective treatment for cystic fibrosis has been hampered by the limited gene transfer efficiency of current vectors. We have shown that recombinant Sendai virus (SeV) is highly efficient in mediating gene transfer to differentiated airway epithelial cells, because of its capacity to overcome the intra- and extracellular barriers known to limit gene delivery. Here, we have identified a novel method to allow the cystic fibrosis transmembrane conductance regulator (CFTR) cDNA sequence to be inserted within SeV (SeV-CFTR). Following in vitro transduction with SeV-CFTR, a chloride-selective current was observed using whole-cell and single-channel patch-clamp techniques. SeV-CFTR administration to the nasal epithelium of cystic fibrosis (CF) mice (Cftr(G551D) and Cftr(tm1Unc)TgN(FABPCFTR)#Jaw mice) led to partial correction of the CF chloride transport defect. In addition, when compared to a SeV control vector, a higher degree of inflammation and epithelial damage was found in the nasal epithelium of mice treated with SeV-CFTR. Second-generation transmission-incompetent F-deleted SeV-CFTR led to similar correction of the CF chloride transport defect in vivo as first-generation transmission-competent vectors. Further modifications to the vector or the host may make it easier to translate these studies into clinical trials of cystic fibrosis.

Positron emission tomography and magnetic resonance imaging in spinocerebellar ataxia type 2: a study of symptomatic and asymptomatic individuals.

Spinocerebellar ataxia type 2 (SCA2) is an autosomal dominant neurodegenerative disorder characterized as an expanded CAG trinucleotide repeats in SCA2 gene resulting in abnormal polyglutamine sequence. We used positron emission tomography (PET) and magnetic resonance imaging (MRI) to clarify metabolic and atrophic changes of the brain in two symptomatic and three asymptomatic individuals who were genetically confirmed for SCA2. PET revealed decreased glucose metabolism in both patients and two of the three asymptomatic carriers in the cerebellum, pons, or both. No PET abnormality was found in the remaining one carrier who had only a very mildly expanded CAG repeat. MRI showed cerebellar and/or pontine atrophic changes in both patients and one of three carriers. The present study suggest that hypometabolism and atrophy of the cerebellum and pons may occur years before the clinical onset of SCA2. PET and MRI may be useful in the early detection of subclinical brain changes associated with SCA2.

Single nucleotide polymorphisms in the gene encoding Krüppel-like factor 7 are associated with type 2 diabetes.

AIMS/HYPOTHESIS: Although genetic susceptibility plays an important role in the pathogenesis of type 2 diabetes, most of the genes that influence susceptibility to type 2 diabetes have yet to be identified. Krüppel-like transcription factors are known to play important roles in development and cell differentiation, and have recently been implicated in the pathogenesis of type 2 diabetes. The present study aimed to examine the associations of single nucleotide polymorphisms (SNPs) in genes encoding members of the Krüppel-like-factor (KLF) family with type 2 diabetes in a large cohort of Japanese subjects. METHODS: We genotyped 33 SNP loci found in 12 KLF genes in subjects with type 2 diabetes and in subjects from the general population using the PCR-Invader assay. We also examined the effects of the overexpression of KLF7 on adipogenesis in 3T3-L1 cells. RESULTS: We identified a significant association between an SNP in KLF7 and type 2 diabetes (A vs C: p=0.004 after Bonferroni's correction, odds ratio=1.59, 95% CI 1.27-2.00). The expression of Klf7 decreased in response to the differentiation of 3T3-L1 adipocytes, and the overexpression of KLF7 resulted in significant inhibition of adipogenesis in 3T3-L1 cells. CONCLUSIONS/INTERPRETATION: These results indicate that the gene encoding KLF7 is a novel candidate for conferring genetic susceptibility to type 2 diabetes.

Bottle choice tests for oxidized oil in rats.

Bottle choice tests in rats are useful for the study of fat appetite. However, little is known about the feeding behavior of rats toward oxidized oil. In this study, 24-h two-bottle choice tests were performed in male Sprague-Dawley (SD) rats for 5 days. When rats were given a choice between thermally oxidized oil and fresh oil, they ingested more fresh oil. A choice test between vapor-added fresh oil and fresh oil showed that rancid flavor is involved in the avoidance of oxidized oil. Results with deodorized oxidized oil indicated that a nonvolatile product also influenced choice. In conclusion, rats could distinguish oxidized oil from fresh oil and preferred fresh oil.

Generation of a recombinant Sendai virus that is selectively activated and lyses human tumor cells expressing matrix metalloproteinases.

Malignant tumor cells often express matrix metalloproteinases (MMPs) at a high level to enable their dissemination and metastasis. Sendai virus (SeV), a nonsegmented negative strand RNA virus, spreads in the target tissues in vivo via cleavage activation of the viral fusion glycoprotein by a tissue-specific, trypsin-like enzyme. By deleting the viral matrix protein, we previously generated a recombinant SeV that does not bud to mature virions, but is highly fusogenic and spreads extensively from cell to cell in a trypsin-dependent manner. Here, we changed the tryptic cleavage site of the fusion glycoprotein of this virus to a site susceptible to MMPs. The resulting recombinant virus was no longer activated by trypsin but spread efficiently in cultured cells supplemented with MMP2 or MMP9 and in human tumor cell lines expressing these MMPs. Furthermore, the virus spread extensively in tumor cells xenotrasplanted to nude mice without disseminating to the surrounding normal cells, leading to the inhibition of the tumor growth in the mice. These results demonstrate the selective targeting and killing of human tumor cells by recombinant SeV technology and greatly advance the reemerging concept of oncolytic virotherapy, which currently appears to rely largely upon a natural preference of certain viruses for cancer cells.

Intraoral grafting of an ex vivo produced oral mucosa equivalent: a preliminary report.

The objective of this study was to assess the efficacy of the use of an ex vivo produced oral mucosa equivalent (EVPOME) for intraoral grafting procedures. Autogenous keratinocytes were harvested from a punch biopsy 4 weeks prior to surgery, placed in a serum-free culture system and seeded onto a human cadaveric dermal equivalent, AlloDerm. Thirty patients with either a premalignant or cancerous lesion were triaged into two groups, depending on the stage of disease: Group 1: EVPOME or Group 2: AlloDerm, control without an epithelial layer. Clinically, EVPOME grafts were easy to handle and showed excellent compliance on grafting. Both, EVPOME and AlloDerm grafts, showed a 100% take rate. At 6 days post-grafting, the EVPOME clinically showed changes indicating vascular ingrowth and had cytologic evidence of the persistence of grafted cultured keratinocytes on the surface. The EVPOME grafts had enhanced maturation of the underlying submucosal layer associated with rapid epithelial coverage when compared to the AlloDerm grafts at biopsies taken at 28 days post-grafting. In summary, EVPOME appears to be an acceptable oral mucosal substitute for human intraoral grafting procedures and results in a more favorable wound healing response than AlloDerm alone.

Three-dimensional structural model analysis of the binding site of lithocholic acid, an inhibitor of DNA polymerase beta and DNA topoisomerase II.

The molecular action of lithocholic acid (LCA), a selective inhibitor of mammalian DNA polymerase beta (pol beta), was investigated. We found that LCA could also strongly inhibit the activity of human DNA topoisomerase II (topo II). No other DNA metabolic enzymes tested were affected by LCA. Therefore, LCA should be classified as an inhibitor of both pol beta and topo II. Here, we report the molecular interaction of LCA with pol beta and topo II. By three-dimensional structural model analysis and by comparison with the spatial positioning of specific amino acids binding to LCA on pol beta (Lys60, Leu77, and Thr79), we obtained supplementary information that allowed us to build a structural model of topo II. Modeling analysis revealed that the LCA-interaction interface in both enzymes has a pocket comprised of three amino acids in common, which binds to the LCA molecule. In topo II, the three amino acid residues were Lys720, Leu760, and Thr791. These results suggested that the LCA binding domains of pol beta and topo II are three-dimensionally very similar.

Catalog of 258 single-nucleotide polymorphisms (SNPs) in genes encoding three organic anion transporters, three organic anion-transporting polypeptides, and three NADH:ubiquinone oxidoreductase flavoproteins.

We present here a series of high-density maps of single-nucleotide polymorphisms (SNPs) detected in genes encoding three organic-anion transporters, three organic anion-transporting polypeptides, and three nicotinamide adenine dinucleotide, reduced:ubiquinone oxidoreductase flavoproteins. A total of 258 SNPs were identified among these nine genes through systematic screening of DNA from 48 Japanese individuals: 17 in 5' flanking regions, three in 5' untranslated regions, 13 in coding regions, 211 in introns, six in 3' untranslated regions, and 8 in 3' flanking regions. By comparing our data with SNPs deposited in the dbSNP database in the National Center for Biotechnology Information, we determined that 236 (91.5%) were novel. In addition, 46 genetic variations of other types were discovered within these loci. These high-resolution maps will serve as a useful resource for analyzing potential associations between variations in these nine genes and differences in human susceptibilities to common diseases or response to drug therapies.

Triterpene constituents from the stem bark of Pinus luchuensis and their DNA topoisomerase II inhibitory effect.

Nine lanostane and serratane-type triterpenes including two unknown compounds were isolated from the stem bark of Pinus luchuensis. These new compounds were characterized as 3-oxolanost-9(11)-ene-24S,25-diol (1) and 29-acetoxy-3beta-methoxyserrat-14-en-21alpha-ol (2) on the basis of spectroscopic evidence. Some of these triterpenes were tested for the inhibitory effect on DNA topoisomerase II activity. Compound 1 showed a slightly less potent inhibitory activity with an IC(50) value of 186 microM.

Catalog of 46 single-nucleotide polymorphisms (SNPs) in the microsomal glutathione S-transferase 1 (MGST1) gene.

A major goal in our laboratory is to understand the role of common genetic variations among individual patients as regards susceptibility to common diseases and differences in therapeutic efficacy and/or side effects of drugs. As an addition to the high-density SNP (single-nucleotide polymorphism) maps of 12 glutathione S-transferase and related genes reported earlier, we provide here an SNP map of the microsomal glutathione S-transferase 1 (MGST1) gene. Among 48 healthy Japanese volunteers examined. we identified a total of 46 SNPs at this locus, 36 of which had not been reported before: 4 in the promoter region, 34 in introns, 3 in the 3' untranslated region, and 5 in the 3' flanking region. No SNP was found in 5'untranslated or coding regions. The ratio of transition to transversion was approximately 1.2:1. Among the 13 insertion-deletion polymorphisms was a 2-bp deletion in the coding region of MGST1 in DNA from one of the volunteers, which resulted in a frame-shift mutation. Since the gene product encoded by this mutant allele would lack the C-terminal half including the MAPEG (membrane-associated proteins in eicosanoid and glutathione metabolism) domain, MGST1 activity is likely to be reduced in the carrier's cells. The SNP map presented here adds to the archive of tools for studying complex genetic diseases, population migration patterns, and a variety of pharmacogenetic possibilities.