Advanced Statistical Analyses to Reduce Inconsistency of Bond Strength Data.

This study was designed to clarify the interrelationship of factors that affect the value of microtensile bond strength (µTBS), focusing on nondestructive testing by which information of the specimens can be stored and quantified. µTBS test specimens were prepared from 10 noncarious human molars. Six factors of µTBS test specimens were evaluated: presence of voids at the interface, X-ray absorption coefficient of resin, X-ray absorption coefficient of dentin, length of dentin part, size of adhesion area, and individual differences of teeth. All specimens were observed nondestructively by optical coherence tomography and micro-computed tomography before µTBS testing. After µTBS testing, the effect of these factors on µTBS data was analyzed by the general linear model, linear mixed effects regression model, and nonlinear regression model with 95% confidence intervals. By the general linear model, a significant difference in individual differences of teeth was observed ( P < 0.001). A significantly positive correlation was shown between µTBS and length of dentin part ( P < 0.001); however, there was no significant nonlinearity ( P = 0.157). Moreover, a significantly negative correlation was observed between µTBS and size of adhesion area ( P = 0.001), with significant nonlinearity ( P = 0.014). No correlation was observed between µTBS and X-ray absorption coefficient of resin ( P = 0.147), and there was no significant nonlinearity ( P = 0.089). Additionally, a significantly positive correlation was observed between µTBS and X-ray absorption coefficient of dentin ( P = 0.022), with significant nonlinearity ( P = 0.036). A significant difference was also observed between the presence and absence of voids by linear mixed effects regression analysis. Our results showed correlations between various parameters of tooth specimens and µTBS data. To evaluate the performance of the adhesive more precisely, the effect of tooth variability and a method to reduce variation in bond strength values should also be considered.

Anorexigenic effects of miglitol in concert with the alterations of gut hormone secretion and gastric emptying in healthy subjects.

Although the α-glucosidase inhibitor miglitol (MG) has been reported to have anorexigenic effects, the mechanism remains to be elucidated. The objective of this study was to explore the effects of MG on appetite in relation to concomitant changes in postprandial gut hormone levels. This randomized open-label crossover study included 20 healthy volunteers. The effects of 50 mg MG on glucagon-like peptide-1 (GLP-1), peptide YY (PYY), and ghrelin levels were assessed in conjunction with a simultaneous determination of appetite scores using visual analogue scales (VAS) over 3 h after the ingestion of a 592 kcal test cookie. Additionally, the gastric emptying rate (GER) was measured using breath ¹³CO2 appearance in 10 subjects. 12 subjects were administered 50 mg MG thrice a day for 1 week, and alterations of the gut hormone levels and the VAS scores for appetite were evaluated. MG pre-administration resulted in a significant enhancement of GLP-1 and PYY responses induced by the cookie ingestion. Following MG administration, ghrelin level declined at 1 h, with a persistent suppression during the postprandial phase in contrast to the restoration to the basal level without MG. Furthermore, MG pre-administration suppressed appetite and maintained satiety evaluated using a VAS rating with concomitant inhibition of GER after cookie ingestion. One-week administration of MG did not influence either gut hormone levels before a meal or VAS rating during a whole day. These observations suggest that MG exerts an anorexigenic effects with concomitant alterations of gut hormone secretions and gastric emptying after meal ingestion.

Hyperadiponectinemia protects against premature death in metabolic syndrome model mice by inhibiting AKT signaling and chronic inflammation.

We previously reported that transgenic (Tg) expression of adiponectin significantly prolonged the lifespan of normal mice. The aim of this study was to elucidate the mechanism involved in the longevity effects of adiponectin using KK/Ta mice, a murine model of metabolic syndrome. We established a Tg line of KK/Ta (Tg-KK/Ta) mice expressing human adiponectin in the liver, and assessed their lifespan. The cause of death was determined by macroscopic and microscopic examinations immediately after death. The expressions of SIRT1, C-reactive protein (CRP), inflammatory cytokines, AMPK, and AKT were measured by quantitative real-time PCR, ELISAs, and/or western blotting. KK/Ta mice had lower serum adiponectin levels and shorter lifespan (57.6±13.9 vs 106.5±18.3 weeks, P<0.0001) than C57BL/6N mice. Tg adiponectin expression significantly extended the lifespan of KK/Ta mice (73.6±16.6 weeks, P<0.001) without affecting body weight, daily food consumption, or plasma glucose levels. Neoplasms were observed in only three of 22 KK/Ta mice that died spontaneously because of tumors. Atherosclerotic lesions were not detected in any mice. SIRT1 levels were not significantly different between KK/Ta and Tg-KK/Ta mice. Gene expressions of Crp, Tnfα, Il6, and Nfκb were increased in KK/Ta mice, but they were significantly attenuated in Tg-KK/Ta mice. Phosphorylated AMPK levels were increased and phosphorylated AKT levels were decreased in Tg-KK/Ta mice. The anti-inflammatory effects of adiponectin, achieved by inhibiting the AKT signaling pathway, may explain how adiponectin slows the accelerated aging process associated with the metabolic syndrome.

Restoration of elbow function by intercostal nerve transfer for obstetrical paralysis with co-contraction of the biceps and the triceps.

Among the conservatively treated patients with obstetrical paralysis after delivery in the vertex presentation, cross reinnervation sometimes occurs in the course of their recovery and co-contraction of multiple muscles impairs smooth upper limb motion. Such co-contraction of the biceps and the triceps inhibits normal elbow motion, making it impossible to use the elbow effectively in daily activities, despite adequate strength in these muscles. To overcome biceps/triceps co-contraction, we performed intercostal nerve transfer to the musculocutaneous nerve for three patients of age 11 months, 6 years and 9 years, respectively, and to the motor branches of the triceps for two patients of age 4 and 14 year-old, respectively. During the average follow-up period of 5.6 (range 1-11.5) years, the power of the reinnervated muscle improved to more than M3, and smooth motion of the elbow independently of shoulder motion was restored.

Differences in the composition of intestinal Bifidobacterium species and the development of allergic diseases in infants in rural Japan.

BACKGROUND: Bifidobacterium is a dominant genus in the intestinal microbiota of infants and comprises many different species. A series of studies performed in northern Europe showed differences in the composition of Bifidobacterium species between allergic infants and healthy controls. Additional studies are needed to confirm this observation. OBJECTIVE: To investigate the composition of fecal Bifidobacterium species in allergic infants and healthy controls in Japan, using molecular methods. METHODS: Full-term born babies were followed up to 6 months of age at a local hospital in rural Japan. The presence or absence of allergy was determined based on allergic symptoms and skin prick tests. Fecal Bifidobacterium species in allergic infants (n=10), and healthy controls (n=16) were evaluated using nine Bifidobacterium species-specific or group-specific primers based on 16S rDNA sequences at 1, 3, and 6 months of age. RESULTS: The number of the infants in whom no Bifidobacterium species could be found was four (15.4%) at 1 month, two (7.7%) at 3 months, and one (3.3%) at 6 months of age, all of whom were healthy controls. At 1 month of age, allergic infants had a higher prevalence f the Bifidobacterium catenulatum group than healthy controls (60.0% vs. 6.3%, P<0.01). At 6 months of age, allergic infants had a higher prevalence of B. bifidum than healthy controls (70.0% vs. 12.5%, P<0.01). These differences were not related to feeding method. CONCLUSIONS: Our results in infants in rural Japan support the hypothesis that a compositional difference in intestinal Bifidobacterium species may be associated with the development of allergy in early infancy, although the responsible species might vary among countries or races.

Lower plasma adiponectin concentration predicts the efficacy of pioglitazone in diabetic patients.

AIM: To investigate clinical efficacy of pioglitazone in association with plasma adiponectin concentration in type 2 diabetic patients. METHODS: Ten diabetic patients were treated with 15 or 30 mg of pioglitazone for 8 weeks, and association between plasma adiponectin concentration before treatment and decrease in glycated albumin levels was examined. RESULTS: Treatment with pioglitazone for 8 weeks lowered glycated albumin level (27.1 +/- 1.2 to 23.8 +/- 1.4%, p < 0.05), and inverse relationship between changes in glycated albumin and plasma adiponectin concentration before treatment was revealed (r = -0.66, p < 0.05). Plasma adiponectin concentrations of patients whose glycated albumin level reduced by more than 10% of the levels before treatment were significantly lower than those of patients whose glycaemic control was affected less (5.2 +/- 0.6 vs. 8.0 +/- 1.0 micro g/ml, p < 0.05). CONCLUSION: Lower plasma adiponectin concentration predicts the clinical efficacy of pioglitazone.

Effects of biotin on glucotoxicity or lipotoxicity in rat pancreatic islets.

Biotin (vitamin H) plays an important role as a cofactor in glucose or lipid metabolism. We showed that biotin potentiated glucose-induced insulin release in isolated rat islets, while biotin alone did not affect insulin release. Coculture with biotin in islets for 48 hours significantly enhanced glucose-induced insulin release or islet insulin content. Similarly, preproinsulin or pancreatic/duodenal homeobox-1 (PDX-1) mRNA was also enhanced in islets cultured with biotin for 48 hours. Furthermore, we measured effects of biotin on beta-cell function under glucotoxic or lipotoxic states. In islets cultured with high glucose or palmitate for 48 hours, glucose-induced insulin release or islet insulin content deteriorated. Coculture with biotin significantly restored glucose-induced insulin release or islet insulin content together with the restoration of preproinsulin or PDX-1 mRNA. We conclude that biotin exerts its beneficial effects on beta-cell dysfunction induced by glucose or free fatty acids probably through the enhancement of insulin biosynthesis.

[Specific and rapid identification of Malassezia species based on the DNA sequences of nuclear ribosomal internal transcribed spacer 1 regions].

Species identification of genus Malassezia is important in epidemiological and etiological studies, however, is difficult by the conventional system. A specific and rapid identification system based on sequences of internal transcribed spacer 1 of ribosomal DNA has therefore been developed. Using this system, we could identify two or more species mixed in the clinical samples.

Effects of bezafibrate on beta-cell function of rat pancreatic islets.

Bezafibrate is an activator of peroxisome proliferator-activated receptors (PPAR) alpha. The present study was performed to investigate the effects of bezafibrate and the PPAR alpha activator, 4-Cholro-6-(2.3-xylidino)-2-pyrimidin-ylthio acetic acid (WY14643), on the beta-cell function of rat pancreatic islets in vitro. In islets cultured with 300 microM bezafibrate or WY14643 for 8 h, a low glucose concentration induced insulin release and increased the levels of mRNA for PPAR alpha, acyl CoA oxidase, carnitine palmitoyl transferase-1, pyruvate dehydrogenase E1 alpha or pyruvate carboxylase. In contrast, after a 48-h culture period, a high glucose concentration induced insulin release and islet insulin content, but decreased the levels of mRNA for glucose transporter-2 (GLUT-2), preproinsulin or pancreatic/duodenal homeobox-1. Diazoxide, the KATP channel opener, restored these responses. We conclude that bezafibrate enhances insulin release through the activation of PPAR alpha gene expression during a short culture period, whereas it may contribute to beta-cell dysfunction through the mechanism of "excessive stimulation" during longer culture periods.

The role of cyclic AMP in the pathogenesis of glucose desensitization in rat pancreatic islets.

Adenosine-3',5'-cyclic monophosphate (cyclic AMP) promotes exocytosis of insulin in pancreatic beta cells. This study was performed to investigate the role of cyclic AMP in the pathogenesis of glucose desensitization in rat pancreatic islets. In islets cultured with high glucose for 48 hours, 27 mmol/L glucose-induced insulin release was markedly impaired, while 3.3 mmol/L glucose-or arginine-induced insulin release was enhanced, indicating glucose desensitization. Islet cyclic AMP content was 190% enhanced in high glucose-culture islets for 48 hours. In islets cultured with dibutyryl-cyclic AMP (dbcAMP) or 3-isobutyl methy-xanthine (IBMX), islet insulin content or 27 mmol/L glucose-induced insulin release was deteriorated. In contrast, 3.3 mmol/L glucose- or arginine-induced insulin release was increased, which was similar to glucose-desensitized islets. Wash-out of dbc AMP for the last 24 hours of the 48-hour culture period restored impaired high glucose-induced insulin release in the same manner as wash-out of high glucose. Diazoxide, the KATP channel opener, also restored impaired high glucose-induced insulin release from dbcAMP-cultured islets. The data suggest that enhancement of cyclic AMP in high glucose-culture islets may be one of the pathogenesis of glucose desensitization.

Effects of free fatty acids on beta-cell functions: a possible involvement of peroxisome proliferator-activated receptors alpha or pancreatic/duodenal homeobox.

It is well known that acute administration of fatty acids enhances insulin release from beta cells, although chronic exposure to fatty acids inhibits insulin release (lipotoxicity). The mechanism for these reciprocal effects of fatty acids on insulin release remains to be elucidated. The present study was performed to investigate the effects of fatty acids on gene expression related to glucose metabolism or insulin biosynthesis. In islets cultured with palmitate for 8 hours, glucose-induced insulin release was enhanced together with increment of pyruvate carboxylase (PC) mRNA or peroxisome proliferator-activated receptors (PPAR)alpha. In contrast, by extending the culture period up to 48 hours, glucose-induced insulin release or islet insulin content was significantly impaired by the coexistence of palmitate. Concomitantly, PC, PPARalpha, GLUT-2, glucokinase (GK), preproinsulin, or pancreatic/duodenal homeobox-1 (PDX-1) mRNA were significantly suppressed in those islets cultured for 48 hours with palmitate. These data may imply that during short-term culture period palmitate promotes PPARalpha gene expression, which enhances PC mRNA expression leading to the enhancement of insulin release, whereas during long-term culture period, palmitate rather inhibits PPARalpha mRNA, which reduces PC mRNA expression. Furthermore, palmitate reduces GLUT-2, GK, or preproinsulin mRNA expression probably through the inhibition of PDX-1 mRNA.

Multiple constrictions of the radial nerve without external compression.

We report a patient with multiple constrictions within the main trunk of the radial nerve that was found after epineurectomy and speculate that the etiology is an inflammatory response. (J Hand Surg 2000; 25A:134-137.

Aminoguanidine exerts a beta-cell function-preserving effect in high glucose-cultured beta-cells (INS-1).

We investigated the effects of aminoguanidine (AG) on beta-cell functions in an insulin secreting cell line (INS-1). Culture with 27mM glucose for one week markedly decreased both insulin release and insulin content compared to culture in 0.8 mM or 3.3 mM glucose. Relative to culture at 27 mM glucose alone, the co-exposure to 1 mM AG almost doubled basal as well as glucose or 25 mM KCl-stimulated insulin release and increased insulin content by 42%. AG failed to affect release and content in cells cultured at 0.8 or 3.3 mM glucose. Preproinsulin mRNA content in 27mM glucose-cultured cells was 52% suppressed compared to 0.8mM glucose-cultured cells, and AG treatment partially counteracted this decline. Advanced glycosylation end product (AGE)-associated fluorescence (370nm excitation and 440 nm emission) of cells' extracts did not differ between 27mM and 0.8mM glucose-cultured cells after 1 week of culture and fluorescence was unaffected by AG. Accumulation of nitrite into culture media was markedly increased from 27mM glucose-cultured cells, and this accumulation was 33% suppressed by AG. In conclusion, AG partially protects against glucotoxic effects in INS-1 cells. These beneficial effects may involve a decrease in early glycation products and/or nitric oxide synthase (NOS) activity. The effects which were obtained after one week of high glucose exposure may supplement AGE-associated effects seen after chronically elevated glucose.

Helicobacter pylori in the natural environment.

The presence of Helicobacter pylori (H. pylori) in the natural environment has been demonstrated in a number of studies. However, its route of infection into humans is unknown. To study this further, we attempted to detect H. pylori in the natural environment in a region of Japan with a high infection rate. Tap and well water and field soil samples were collected from around the residences of subjects who had participated in an epidemiological survey in 1996. Samples of water from rivers and ponds, and specimens of flies and cow faeces were collected in the region. DNA was extracted from the water, field soil and faecal samples after selective collection of H. pylori by the immunomagnetic-bead separation technique. H. pylori-specific DNA was detected in water, field soil, flies and cow faeces by nested polymerase chain reaction (PCR), and the ureA partial sequences of the PCR products were aligned. The nucleotide sequences of the samples amplified by PCR were highly homologous (96-100%) with the H. pylori sequence in the GenBank database and the H. pylori-specific DNA sequences were highly homologous with each other. These findings suggest the existence of H. pylori in the natural environment and a possible transmission route.

A positioning device to allow rotation for cine-MRI of the distal radioulnar joint.

'Cine-mode' magnetic resonance imaging (MRI) is useful in the diagnosis and treatment of patients with disorders of joint motion. We have designed a device for imaging of the distal radioulnar joint by cine-MRI. Five normal wrists and eight patients with rheumatoid arthritis were investigated prior to surgery for subluxation of the distal radioulnar joint. Normally, the radius moves in a constant arc around the ulna, whereas in rheumatoid wrists the centre of rotation varies continuously with increase in rotation. The device should prove helpful in imaging disorders of the distal radioulnar joint.

Interactions between vitamin E and glucose on B-cell functions in the rat: an in vivo and in vitro study.

Hyperglycemia exerts negative effects on B-cell functions that may involve oxidative stress. This was tested for by investigating effects of D-alpha-tocopherol (vitamin E), known as a free radical scavenger, on B-cell functions. For in vivo testing, rat pancreatic islets were transplanted syngeneically under the kidney capsule in streptozotocin-induced diabetic rats. Transplanted rats were treated with D-alpha-tocopherol (40 mg/kg, intraperitoneally injected every other day) or soybean oil for 2 or 6 weeks. Graft-bearing kidneys were then perfused and insulin release was measured after stimulation sequentially with glucose (27 mmol/L) and L-arginine (10 mmol/L). D-alpha-tocopherol treatment for 2 or 6 weeks failed to restore glucose-induced insulin secretion, whereas treatment for 2 but not for 6 weeks enhanced basal insulin release (by 24%, p < 0.05) and arginine-induced release (by 79%, p < 0.05). Treatment did not affect graft content of preproinsulin mRNA. The presence of D-alpha-tocopherol (50 microg/ml) in vitro enhanced glucose (27 mmol/L)-stimulated insulin release from batch-type incubated islets by 33% (p < 0.05). Exposing islets to D-alpha-tocopherol for 1 day in the presence of 38 mmol/L glucose enhanced glucose-stimulated insulin release (by 25%, p < 0.05), L-arginine-stimulated release (by 37%, p < 0.05) and elevated islet insulin content (by 20%, p < 0.05). These effects of exposure to D-alpha tocopherol were lost when the culture period was extended up to 3 weeks. It is concluded that D-alpha-tocopherol exerts moderate beneficial effects on B-cell functions during short to intermediate length of high glucose exposure. These effects are, however, insufficient to support a major role for oxidative stress behind glucotoxicity toward B cells.

Phylogenetic classification and species identification of dermatophyte strains based on DNA sequences of nuclear ribosomal internal transcribed spacer 1 regions.

The mutual phylogenetic relationships of dermatophytes of the genera Trichophyton, Microsporum, and Epidermophyton were demonstrated by using internal transcribed spacer 1 (ITS1) region ribosomal DNA sequences. Trichophyton spp. and Microsporum spp. form a cluster in the phylogenetic tree with Epidermophyton floccosum as an outgroup, and within this cluster, all Trichophyton spp. except Trichophyton terrestre form a nested cluster (100% bootstrap support). Members of dermatophytes in the cluster of Trichophyton spp. were classified into three groups with ITS1 homologies, with each of them being a monophyletic cluster (100% bootstrap support). The Arthroderma vanbreuseghemii-Arthroderma simii group consists of A. vanbreuseghemii, A. simii, Trichophyton mentagrophytes isolates from humans, T. mentagrophytes var. quinckeanum, Trichophyton tonsurans, and Trichophyton schoenleinii. Arthroderma benhamiae, T. mentagrophytes var. erinacei, and Trichophyton verrucosum are members of the Arthroderma benhamiae group. Trichophyton rubrum and Trichophyton violaceum form the T. rubrum group. This suggests that these "species" of dermatophytes have been overclassified. The ITS1 sequences of 11 clinical isolates were also determined to identify the species, and all strains were successfully identified by comparison of their base sequences with those in the ITS1 DNA sequence database.

Multiple origins of fungal group I introns located in the same position of nuclear SSU rRNA gene.

The archiascomycetous fungus Protomyces pachydermus has two group I introns within the nuclear small subunit (nSSU) rRNA gene. One of these introns has an internal open reading frame (ORF) that encodes a predicted protein of 228 amino acid residues. On the other hand, Protomyces macrosporus has two group I introns that insert at the same positions as P. pachydermus, which have no ORF. Each alignment was constructed with Protomyces group I introns located in the same position and other introns retrieved by the BLAST Search. Each phylogenetic tree based on the alignment shows that Protomyces introns are monophyletic but the relationships among fungal introns do not reflect on the fungal phylogeny. Therefore, it is suggested that two different horizontal transfers of group I introns occurred at the early stage of Protomyces species diversification.

Incorporation of an artificial receptor into a native protein: new strategy for the design of semisynthetic enzymes with allosteric properties.

The sugar-facilitated structure and enzymatic activity change of engineered myoglobins bearing a phenylboronic acid moiety, which were semisynthesized by a cofactor reconstitution method, were studied by the denaturation experiment, spectrophotometric titration of the pKa shift of the axial H2O, circular dichloism (CD), and the kinetics of the myoglobin-catalyzed-aniline hydroxylation reaction. Both boronophenylalanine-appended myoglobin [Mb(m-Bphe)2] and phenylboronic acid-appended myoglobin [Mb(PhBOH)2] were stabilized by approximately 2 kcal/mol upon complexation with D-fructose. CD spectral changes and the sugar-induced pKa shift suggested that the microenvironment of the active site of these myoglobins was re-formed from a partially disturbed state to that comparable to the native state upon D-fructose binding. The correlation of pKa with kcat (for the aniline hydroxylase activity) and the delta GDH2O-kcat profile showed that these structural changes of Mb-(m-Bphe)2 and Mb(PhBOH)2 were closely related to their sugar-enhanced aniline hydroxylase activity. Thus, the results established that an incorporation of the artificial receptor molecule can be a valid methodology for the design of stimuli-responsive semiartificial enzymes.