Differential Expression and Roles of Secreted Frizzled-Related Protein 5 and the Wingless Homolog Wnt5a in Periodontitis.

The Wingless/integrase-1 (Wnt) family of protein ligands and their functional antagonists, secreted frizzled-related proteins (sFRPs), regulate various biological processes ranging from embryonic development to immunity and inflammation. Wnt5a and sFRP5 comprise a typical ligand/antagonist pair, and the former molecule was recently detected at the messenger RNA (mRNA) level in human periodontitis. The main objective of this study was to investigate the interrelationship of expression of Wnt5a and sFRP5 in human periodontitis (as compared to health) and to determine their roles in inflammation and bone loss in an animal model. We detected both Wnt5a and sFRP5 mRNA in human gingiva, with Wnt5a dominating in diseased and sFRP5 in healthy tissue. Wnt5a and sFRP5 protein colocalized in the gingival epithelium, suggesting epithelial cell expression, which was confirmed in cultured human gingival epithelial cells (HGECs). The HGEC expression of Wnt5a and sFRP5 was differentially regulated by a proinflammatory stimulus (lipopolysaccharide [LPS] from Porphyromonas gingivalis) in a manner consistent with the clinical observations (i.e., LPS upregulated Wnt5a and downregulated sFRP5). In HGECs, exogenously added Wnt5a enhanced whereas sFRP5 inhibited LPS-induced inflammation, as monitored by interleukin 8 production. Consistent with this, local treatment with sFRP5 in mice subjected to ligature-induced periodontitis inhibited inflammation and bone loss, correlating with decreased numbers of osteoclasts in bone tissue sections. As in humans, mouse periodontitis was associated with high expression of Wnt5a and low expression of sFRP5, although this profile was reversed after treatment with sFRP5. In conclusion, we demonstrated a novel reciprocal relationship between sFRP5 and Wnt5a expression in periodontal health and disease, paving the way to clinical investigation of the possibility of using the Wnt5a/sFRP5 ratio as a periodontitis biomarker. Moreover, we showed that sFRP5 blocks experimental periodontal inflammation and bone loss, suggesting a promising platform for the development of a new host modulation therapy in periodontitis.

Subtotal glossectomy preserving half the tongue base prevents taste disorder in patients with tongue cancer.

Most patients diagnosed with tongue carcinoma undergo surgical resection and reconstructive surgery to preserve tongue mobility and swallowing functions. Twenty-four patients who underwent a total or subtotal glossectomy and surgical reconstruction were evaluated for residual taste sensation. The graded filter paper test for all four tastes (sweet, salt, sour, and bitter) was performed on the posterior wall of the oropharynx and on tongue remnants if they were visible from the mouth. Eleven of the 24 patients were aware of their taste disorder after surgery. Four patients with more than 1/2 residual tongue base had no taste complaints, whereas seven of 14 patients with less than 1/3 residual tongue base reported taste abnormalities. Patients who could only tolerate a poor diet or tube feeding tended to have taste complaints (P=0.017). The taste test showed that the taste threshold of the residual tongue was significantly lower compared to controls. The taste threshold was significantly correlated with the remaining volume of tongue base. Patients with >1/2 the tongue base remaining had good taste sensation, whereas those with <1/3 residual tissue had impaired taste. This study suggests that glossectomy strategies aimed at preserving at least half the tongue base may substantially reduce dysgeusia in the patients.

Short-interval intracortical inhibition in Parkinson's disease using anterior-posterior directed currents.

Reduced short-interval intracortical inhibition (SICI) is reported in Parkinson's disease (PD) and is considered to reflect abnormal GABAergic inhibitory system of the primary motor cortex in PD. We have recently shown, however, that SICI using anterior-posterior directed currents in the brain was normal in focal dystonia even though that using posterior-anterior currents was abnormal, indicating that the GABAergic system of the primary motor cortex is largely normal in dystonia. Here, we studied SICI in PD to clarify whether the GABAergic system is completely impaired in PD. We used paired-pulse transcranial magnetic stimulation to study SICI at interstimulus intervals of 3 and 4 ms with anterior-posterior or posterior-anterior directed currents in eight PD patients and ten healthy volunteers. The amount of SICI with posterior-anterior directed currents was reduced in PD patients compared with healthy volunteers; in contrast, SICI studied with anterior-posterior directed currents was normal in PD patients. These observations may be due to the difference in I-wave composition generated by the two directed currents and/or the difference in responsible inhibitory interneurons for the inhibition between the two current directions. We suggest that some or a part of inhibitory interneurons are not involved in PD. This discrepancy between SICI using posterior-anterior and anterior-posterior directed currents experiments may provide additional information about the circuits of the motor cortex.

Triad stimulation frequency for cortical facilitation in cortical myoclonus.

BACKGROUND: Abnormally enhanced cortical rhythmic activities have been reported in patients with cortical myoclonus. We recently reported a new triad-conditioning transcranial magnetic stimulation (TMS) method to detect the intrinsic rhythms of the primary motor cortex (M1). Triad-conditioning TMS revealed a 40-Hz intrinsic rhythm of M1 in normal subjects. In this investigation, we study the motor cortical facilitation induced by rhythmic triple TMS pulses (triad-conditioning TMS) in patients with cortical myoclonus. METHODS: Subjects were 7 patients with cortical myoclonus (28-74 years old) and 13 healthy volunteers (30-71 years old). Three conditioning stimuli over M1 at the intensity of 110% active motor threshold preceded the test TMS at various interstimulus intervals corresponding to 10-200 Hz. The resulting amplitudes of conditioned motor evoked potentials recorded from the contralateral hand muscle were compared with those evoked by the test stimulus alone. RESULTS: The facilitation at 25 ms (40 Hz) observed in normal subjects was absent in patients with cortical myoclonus. Instead, triad-conditioning TMS induced facilitation at a 40 ms interval (25 Hz) in cortical myoclonus. DISCUSSIONS: This change in the timing of facilitation may be explained by a shift of the most preferential intrinsic rhythm of M1, or by some dysfunction in the interneuronal network in cortical myoclonus.

A comparison of the effect on QT interval between thiamylal and propofol during anaesthetic induction*.

SUMMARY: The aim of this study was to determine the effect of thiamylal and propofol on heart rate-corrected QT (QTc) interval during anaesthetic induction. We studied 50 patients undergoing lumbar spine surgery. Patients were administered 3 microgxkg(-1) fentanyl and were randomly allocated to receive 5 mgxkg(-1) thiamylal or 1.5 mgxkg(-1) propofol as an induction agent. Tracheal intubation was performed after vecuronium administration. Heart rate, mean arterial pressure, bispectral index score, and 12-lead electrocardiogram were recorded at the following time points: just before (T1) and 2 min after (T2) fentanyl administration; 2 min after anaesthetic administration (T3); 2.5 min after vecuronium injection (T4); and 2 min after intubation (T5). Thiamylal prolonged (p < 0.0001), but propofol shortened (p < 0.0001), the QTc interval.

Effects of STN stimulation on the initiation and inhibition of saccade in Parkinson disease.

OBJECTIVES: The basal ganglia (BG) play an important role in controlling saccades. Deep brain stimulation (DBS) of the subthalamic nucleus (STN) is widely used as a treatment of Parkinson disease (PD) by altering the function of the BG. Nevertheless, the effects of STN DBS on saccade performance are not fully clarified in a systematic manner. In this study, we examined the effects of bilateral STN DBS on both the initiation and inhibition of saccades in PD. METHODS: Thirty-two patients with PD performed 4 oculomotor tasks. Two tasks (visually guided saccades and gap saccades) were reflexive and 2 (memory-guided saccades [MGS] and antisaccades) were volitional. While taking their regular doses of antiparkinsonian drugs, patients performed these tasks under 2 conditions: during DBS (DBS-on condition) and without DBS (DBS-off condition). Fifty-one age-matched subjects served as controls. RESULTS: In the DBS-on condition, parameters of saccade initiation were improved in all tasks, with shorter latencies and increased amplitudes, except for MGS latency. STN DBS improved the ability to suppress unwanted saccades to the cue stimulus in the MGS task. However, it did not suppress prosaccades during the antisaccade task. CONCLUSIONS: These results suggest that deep brain stimulation (DBS) of the subthalamic nucleus (STN) affects the neural pathway common to both reflexive and volitional saccades, possibly by acting on the STN-substantia nigra pars reticulata-superior colliculi pathway. STN DBS may set the functional level of the superior colliculi appropriate for both saccade initiation and inhibition through this pathway. These findings provide novel insights into the pathophysiology of Parkinson disease and may yield better treatment strategies.

Molecular and biological characterization of gtf regulation-associated genes in Streptococcus mutans.

INTRODUCTION: Surface protein antigen (PAc) and glucosyltransferases (GTF) are major adhesive molecules of Streptococcus mutans, though the mechanism of their regulation has not been fully elucidated. METHODS: To investigate the regulation mechanism, we determined a nucleotide sequence in the upstream region of the pac locus in S. mutans and identified two open reading frames (ORF), designated as orf1 and orf2. Each ORF was inactivated and functional analyses were performed. RESULTS: Western blot analyses revealed that the expression level of PAc was unaffected, while that of cell-associated GTF was diminished in both mutant strains. Furthermore, they showed higher hydrophobicity levels and an impaired sucrose-dependent adherence to smooth surfaces. RNA dot blot analysis demonstrated that transcriptions of the gtfB and the gtfC genes, which encode GTF-I and GTF-SI, respectively, were downregulated, while that of pac was comparable to the wild-type strain. In addition, the GTF activities of the mutant strains were significantly lower than those of the wild-type, though a greater amount of total glucan produced by the mutants was noted in culture supernatants. CONCLUSION: These findings suggest that orf1 and orf2 are associated with positive regulation of the gtfB and gtfC genes.

The effect of sevoflurane-induced hypotension in combination with acute hypervolaemic haemodilution on middle cerebral artery flow velocity in surgical patients.

BACKGROUND AND OBJECTIVE: This study was carried out to clarify the effect of the combination of acute hypervolaemic haemodilution and hypotensive anaesthesia induced with sevoflurane on human middle cerebral artery flow velocity using transcranial Doppler ultrasonography. METHODS: Thirty patients who were maintained with N2O-O2-sevoflurane anaesthesia undergoing hip surgery were randomly divided into two groups (no controlled hypotension group, Group A, and controlled hypotension group, Group B). Haemodilution was produced by acute preoperative infusion of 1000 mL of hydroxyethylstarch without removing blood in both groups. Mean arterial pressure was maintained at approximately 95 mmHg in Group A and at 55 mmHg for 80 min by increasing the inspired concentration of sevoflurane in Group B. Middle cerebral artery flow velocity was measured before haemodilution, after haemodilution, 80 min after starting hypotension, and 60 min after recovery from hypotension. RESULTS: Middle cerebral artery flow velocity significantly increased in both groups after haemodilution; by 28%, in Group A, P < 0.05 vs. before haemodilution and by 30% vs. before haemodilution in Group B, P < 0.05). During controlled hypotension, it decreased towards the pre-haemodilution value (P < 0.05 vs. after haemodilution). CONCLUSIONS: Sevoflurane-induced hypotension to a mean arterial pressure of 55 mmHg would reduce middle cerebral artery flow that had been increased by acute hypervolaemic haemodilution, such as haematocrit value of 26%, whereas it could preserve the flow in pre-haemodilution condition during normocapnia.

Difference in intracortical inhibition of the motor cortex between cortical myoclonus and focal hand dystonia.

OBJECTIVE: The short interval intracortical inhibition (SICI) of the motor cortex (M1) is reduced in both cortical myoclonus and focal hand dystonia. This reduction has been attributed to the dysfunction of GABAergic system within the motor cortex. However, the precise mechanisms underlying the reduction may not be entirely identical in these two disorders, being due to primary pathological involvement in M1 or secondary to functional changes outside M1. The aim of this study was to elucidate possible differences in intracortical inhibition between these two disorders. METHODS: Subjects were 11 patients with benign myoclonus epilepsy, 7 with focal hand dystonia, and 11 normal volunteers. We studied SICI using anterior-posterior (AP) directed and posterior-anterior (PA) directed induced currents in the brain. RESULTS: In both disorders, SICI with PA-directed currents was reduced as reported previously. In contrast, SICI studied with AP currents was normal in patients with focal hand dystonia, but reduced in patients with cortical myoclonus. CONCLUSIONS: The difference between the two disorders might reflect the underlying pathological difference. In cortical myoclonus, the inhibitory interneurons of the motor cortex are affected, whereas the same interneurons are intact in dystonia. The difference in SICI induced by AP and PA directed currents in dystonia may be explained by the following possibilities: the difference in composition of I-waves contributing to EMG generation and the difference in modulation of the interneuronal activity by voluntary contraction. These changes may be secondary to dysregulation of the motor cortex by the basal ganglia or related cortices in dystonia. SIGNIFICANCE: The SICI using AP directed currents together with the conventional SICI using PA directed currents was able to demonstrate some difference in the intrinsic circuits of M1 between myoclonus and focal hand dystonia. SICI using AP directed currents can provide additional information about the motor cortical excitability changes over those obtained by the previously reported methods.

Evaluation of corticospinal tracts in ALS with diffusion tensor MRI and brainstem stimulation.

OBJECTIVE: To assess corticospinal tract involvement in patients with amyotrophic lateral sclerosis (ALS) by correlating diffusion tensor imaging (DTI) measures with intra- and extracranial central motor conduction time (CMCT) and clinical features of the patients. METHODS: We investigated 31 patients with ALS and 31 normal volunteers by DTI and measured fractional anisotropy (FA) within the corticospinal tracts and in the extramotor white matter. We measured CMCT for the first dorsal interosseous muscle and segmented it into cortical-brainstem (CTX-BS CT) and brainstem-cervical root (BS-CV CT) conduction times by magnetic brainstem stimulation at the foramen magnum level. Clinical status of each patient was evaluated with the ALS Functional Rating Scale-Revised (ALSFRS-R) and upper motor neuron (UMN) score devised for this study. RESULTS: We found a significant decrease of mean FA in all regions of the corticospinal tracts in patients with ALS as compared with controls. We found that FA along the corticospinal tract decreased significantly with higher UMN scores. There was no significant correlation between FA and ALSFRS-R, to which both upper and lower motoneuron involvements contribute. FA showed a significant correlation with the intracranial part of the central motor conduction (CTX-BS CT) but not with the extracranial conduction time. CONCLUSIONS: Fractional anisotropy reflects functional abnormality of intracranial corticospinal tracts and can be used for objective evaluation of upper motor neuron impairment in amyotrophic lateral sclerosis.

Inflammatory immune responses by water-insoluble alpha-glucans.

Water-insoluble alpha-glucans are synthesized from sucrose by glucosyltransferase-I of mutans streptococci and play an important role in the development of dental plaque. Several types of beta-glucans in fungal cell wall components and water-soluble alpha-glucans from Streptococcus mutans are known to modulate innate immunity. In the present study, we investigated whether water-insoluble alpha-glucans also induced inflammatory innate immune responses. Our results showed that water-insoluble alpha-glucans synthesized by Streptococcus sobrinus activated mouse peritoneal exudate macrophages to produce pro-inflammatory cytokines. The immunological responses were not due to contamination by sucrose, water-soluble alpha-glucan, lipopolysaccharide, or peptidoglycan. Furthermore, human monocytes stimulated by water-insoluble alpha-glucans produced TNF-alpha and IL-8, while human polymorphonuclear cells were activated by water-insoluble alpha-glucans, resulting in chemotaxis and hydrogen peroxide production. The results demonstrated that water-soluble alpha-glucans modulate macrophage- and granulocyte-induced inflammatory immune responses, and suggest that inflammation induced by those alpha-glucans is associated with the development of periodontal diseases.

Improvement of the activity of arylmalonate decarboxylase by random mutagenesis.

Arylmalonate decarboxylase (EC 4.1.1.76) catalyzes enantioselective decarboxylation of alpha-aryl-alpha-methylmalonates to give optically pure alpha-arylpropionates. Recently, we have succeeded in creating a double mutant enzyme that gave opposite enantionmer as the product. Unfortunately, however, the activity of the mutant decreased far lower than that of the native enzyme. Thus, we performed the directed evolution of the mutant via the random mutagenesis method employing the mutator strain Escherichia coli XL1-Red. About 50,000 mutants were screened on color assay plate, and one mutant with higher activity was obtained. Gene analysis of this mutant indicated that the obtained enzyme had an S36N mutation in addition to its original G74C/C188S mutations. The activity of the triple mutant enzyme was tenfold higher than that of the starting doubly mutated enzyme.

The effect of bisphenol A and chlorinated derivatives of bisphenol A on the level of serum vitellogenin in Japanese medaka (Oryzias latipes).

2,2-bis (4-hydroxyphenyl) propane or Bisphenol A (BPA), has been reported to behave as an endocrine disrupter below acute toxic levels, and is widely present in the water environment. Although BPA is easily chlorinated, very little is reported on the effect of chlorinated BPA to the aquatic organisms. In this study, the estrogenic activities of BPA and its chlorinated derivatives were evaluated by the induction of vitellogenin (VTG) in the serum of mature male Japanese medaka. In addition, the effect of sodium hypochlorite on the decomposition of BPA was tested. The relative potencies of estrogenic activities of chlorinated BPA descended in the order 3,3'-diCIBPA>BPA> or =3-CIBPA>3,3',5-triCIBPA, and no estrogenic activity was observed in 3,3',5,5'-tetraCIBPA. Lowest Observed Effect Concentration (LOEC) and No Observed Effect Concentration (NOEC) for both 3-CIBPA and 3,3'-diCIBPA were 500 microg/L and 200 microg/L, respectively. LOEC for 3,3',5-triCIBPA was >500 microg/L. When BPA was reacted with sodium hypochlorite (24 hours; residual chlorine at 1 ppm), however, complete decomposition of BPA and its chlorinated derivatives was observed. The decrease in BPA and its chlorinated derivatives paralleled the decrease in estrogenic potency evaluated by the induction of vitellogenin (VTG) in the serum of mature male Japanese medaka.

Distribution of emm genotypes and superantigen genes of Streptococcus pyogenes isolated in Japan, 1994-9.

The purpose of this study was to examine characteristic profiles of Streptococcus pyogenes clinical isolates isolated in Japan during 1994-9. Genotyping of the M protein (emm typing) revealed that emm types 12 and 28 were the most common among 316 isolates. Most of the emm12 isolates were isolated from mucosa, while emm58 and emm89 were from skin. Moreover, the emm3 isolates were dominant in invasive infections. The distribution of 6 superantigen genes showed that all isolates harboured the mf gene and many had the speG gene. Invasive isolates were shown to have the ssa gene at a higher rate (76%) than noninvasive (37%). The distribution of superantigens was significantly different between emm types, but not between isolation sites. These results suggest that the distribution of emm types is related to isolation site, whereas superantigen distribution is related to clinical features of S. pyogenes infections.

Growth-associated gene expression profiles by microarray analysis of trophoblast of molar pregnancies and normal villi.

We used microarray analysis to investigate expression profiles of 589 known genes committed to cell growth control to characterize regulatory circuitry for cell proliferation in complete moles (CMs). CMs are characterized by hyperplastic trophoblast and have a high propensity to give rise to choriocarcinoma. Characteristic alterations in gene expression profiles were observed when compared with normal villi. Fifty-seven genes were significantly up-regulated in CMs and involved the Ras-Map kinase 3, Jak-STAT5, and Wnt signal pathways, implicating growth factor or cytokine-mediated signal pathways in the trophoblastic hyperplasia of CMs. Several genes associated with anti-apoptosis, cell structuring, and/or cell attachment were also up-regulated in CMs. In contrast, relatively fewer genes were down-regulated and these involved IGFBPs, versican, interleukin-1, tumor necrosis factor receptor, CD44, and RAD52. Genes identified in this study may elucidate regulation mechanisms of trophoblastic proliferation and mechanisms causing a pathological phenotype in CMs.

Interplay of the main chain, chiral side chains, and solvent in conformational transitions: poly([(R)-3,7-dimethyloctyl]-[(S)-3-methylpentyl]silylene).

Light scattering, sedimentation equilibrium, viscosity, circular dichroism (CD), and UV absorption (UV) measurements were made on dilute solutions of poly([(R)-3,7-dimethyloctyl]-[(S)-3-methylpentyl]silylene)(PRS) as functions of molecular weight. From light scattering and viscosity data, PRS is found to be a very stiff polymer of persistence length q as large as 103 nm at 25 degrees C, essentially a 7(3) helix found in the solid state; q increases only gradually with lowering temperature between -15 and 25 degrees C. The CD data show that PRS undergoes a conformational transition around 3 degrees C in isooctane (transition temperature T(c)). The CD signal is largely positive at low temperatures, passes through zero at T(c), and becomes largely negative at higher temperatures; T(c) is independent of sample's chain length N. This is a highly cooperative helix (M)-to-helix (P) transition depending remarkably on N, as PRS is substantially rodlike. The CD data are converted to the fraction f(P) of P helix as a function of N and analyzed successfully by a statistical mechanical theory based on a helix reversal model, where a polymer chain consists of M and P helices intervened by helix reversals, with the result that the free energy difference DeltaG(h) between P and M shows a temperature dependence similar to that of 2f(P) - 1, whereas the helix reversal energy is substantially constant at 1.2 x 10(4) J mol(-1); the latter value means that the helix reversal occurs only once in 100 Si units or less. This DeltaG(h) change and solvent dependence of T(c) are explained by a double-well potential for the rotation about Si-Si bonds, which incorporates into DeltaG(h) the solvent interactions with the helical grooves of side chains surrounding the main chain. Detailed features of UV absorption spectra at different temperature and molecular weights are also presented.

Decreased sensory cortical excitability after 1 Hz rTMS over the ipsilateral primary motor cortex.

OBJECTIVES: To study changes in the excitability of the sensory cortex by repetitive transcranial magnetic stimulation (rTMS) in humans. METHODS: Somatosensory evoked potentials (SEPs) and antidromic sensory nerve action potentials (SNAPs) were elicited by right median nerve stimulation at the wrist before and after low frequency (1 Hz) rTMS over the left motor cortex, lateral premotor cortex, sensory cortex, and also after sham stimulation. The intensity of rTMS was fixed at 1.1 times the active motor threshold at the hand area of motor cortex. RESULTS: N20 peak (N20p)-P25 and P25-N33 amplitudes were suppressed after rTMS over the motor cortex, whereas the N20 onset (N20o)-N20p and SNAP amplitudes were not affected. They recovered to the baseline about 100 min after the rTMS. rTMS over the premotor cortex or sensory cortex or sham stimulation had no suppressive effect on SEPs. CONCLUSIONS: The reduction of N20p-P25 and P25-N33 components without any changes of N20o-N20p amplitude suggests that the suppression occurs in the sensory cortex. rTMS (1 Hz) of the motor cortex induces a long-lasting suppression of the ipsilateral sensory cortex even at an intensity as low as 1.1 times the active motor threshold, probably via cortico-cortical pathways between motor and sensory cortex.

Fba, a novel fibronectin-binding protein from Streptococcus pyogenes, promotes bacterial entry into epithelial cells, and the fba gene is positively transcribed under the Mga regulator.

In infection by Streptococcus pyogenes, fibronectin (Fn)-binding proteins play important roles as adhesins and invasins. Here, we present a novel Fn-binding protein of S. pyogenes that exhibits a low similarity to other Fn-binding proteins reported. After searching the Oklahoma Streptococcal Genome Sequencing Database for open reading frames (ORFs) with an LPXTG motif, nine ORFs were found among those recognized as putative surface proteins, and one of them was designated as Fba. The fba gene was found in M types 1, 2, 4, 22, 28 and 49 of S. pyogenes, but not in other serotypes or groups of streptococci. Fba, a 37.8 kDa protein, possesses three or four proline-rich repeat domains and exhibits a high homology to FnBPA, the Fn-binding protein of Staphylococcus aureus. Recombinant Fba exhibited a strong binding ability to Fn. In addition, Fba-deficient mutants showed diminished invasive capabilities to HEp-2 cells and low mortality in mice following skin infection. The fba gene was located downstream of the mga regulon and analysis using an mga-inactivated mutant revealed that it was transcribed under the control of the Mga regulator. These results indicate that Fba is a novel protein and one of the important virulence factors of S. pyogenes.