Localized NMR Mediated by Electrical-Field-Induced Domain Wall Oscillation in Quantum-Hall-Ferromagnet Nanowire.

We present fractional quantum Hall domain walls confined in a gate-defined wire structure. Our experiments utilize spatial oscillation of domain walls driven by radio frequency electric fields to cause nuclear magnetic resonance. The resulting spectra are discussed in terms of both large quadrupole fields created around the wire and hyperfine fields associated with the oscillating domain walls. This provides the experimental fact that the domain walls survive near the confined geometry despite of potential deformation, by which a localized magnetic resonance is allowed in electrical means.

Optically Induced Nuclear Spin Polarization in the Quantum Hall Regime: The Effect of Electron Spin Polarization through Exciton and Trion Excitations.

We study nuclear spin polarization in the quantum Hall regime through the optically pumped electron spin polarization in the lowest Landau level. The nuclear spin polarization is measured as a nuclear magnetic field B(N) by means of the sensitive resistive detection. We find the dependence of B(N) on the filling factor nonmonotonic. The comprehensive measurements of B(N) with the help of the circularly polarized photoluminescence measurements indicate the participation of the photoexcited complexes, i.e., the exciton and trion (charged exciton), in nuclear spin polarization. On the basis of a novel estimation method of the equilibrium electron spin polarization, we analyze the experimental data and conclude that the filling factor dependence of B(N) is understood by the effect of electron spin polarization through excitons and trions.

Anti-PD-1 antibody monotherapy versus anti-PD-1 plus anti-CTLA-4 combination therapy as first-line immunotherapy in unresectable or metastatic mucosal melanoma: a retrospective, multicenter study of 329 Japanese cases (JMAC study).

BACKGROUND: Anti-programmed cell death protein 1 (PD-1) antibody monotherapy (PD1) has led to favorable responses in advanced non-acral cutaneous melanoma among Caucasian populations; however, recent studies suggest that this therapy has limited efficacy in mucosal melanoma (MCM). Thus, advanced MCM patients are candidates for PD1 plus anti-cytotoxic T lymphocyte-associated antigen-4 (CTLA-4) combination therapy (PD1 + CTLA4). Data on the efficacy of immunotherapy in MCM, however, are limited. We aimed to compare the efficacies of PD1 and PD1 + CTLA4 in Japanese advanced MCM patients. PATIENTS AND METHODS: We retrospectively assessed advanced MCM patients treated with PD1 or PD1 + CTLA4 at 24 Japanese institutions. Patient baseline characteristics, clinical responses (RECIST), progression-free survival (PFS), and overall survival (OS) were estimated using Kaplan-Meier analysis, and toxicity was assessed to estimate the efficacy and safety of PD1 and PD1 + CTLA4. RESULTS: Altogether, 329 patients with advanced MCM were included in this study. PD1 and PD1 + CTLA4 were used in 263 and 66 patients, respectively. Baseline characteristics were similar between both treatment groups, except for age (median age 71 versus 65 years; P < 0.001). No significant differences were observed between the PD1 and PD1 + CTLA4 groups with respect to objective response rate (26% versus 29%; P = 0.26) or PFS and OS (median PFS 5.9 months versus 6.8 months; P = 0.55, median OS 20.4 months versus 20.1 months; P = 0.55). Cox multivariate survival analysis revealed that PD1 + CTLA4 did not prolong PFS and OS (PFS: hazard ratio 0.83, 95% confidence interval 0.58-1.19, P = 0.30; OS: HR 0.89, 95% confidence interval 0.57-1.38, P = 0.59). The rate of ≥grade 3 immune-related adverse events was higher in the PD1 + CTLA4 group than in the PD1 group (53% versus 17%; P < 0.001). CONCLUSIONS: First-line PD1 + CTLA4 demonstrated comparable clinical efficacy to PD1 in Japanese MCM patients, but with a higher rate of immune-related adverse events.

Schwann cell autophagy induced by SAHA, 17-AAG, or clonazepam can reduce bortezomib-induced peripheral neuropathy.

BACKGROUND: The proteasome inhibitor bortezomib has improved the survival of patients with multiple myeloma but bortezomib-induced peripheral neuropathy (BiPN) has emerged as a serious potential complication of this therapy. Animal studies suggest that bortezomib predominantly causes pathological changes in Schwann cells. A tractable system to evaluate combination drugs for use with bortezomib is essential to enable continuing clinical benefit from this drug. METHODS: Rat schwannoma cells were pretreated with vincristine (VCR), histone deacetylase inhibitors, anticonvulsants, or a heat-shock protein 90 (HSP90) inhibitor. To then monitor aggresome formation as a result of proteasome inhibition and the activation of chaperone-mediated autophagy (CMA), we performed double-labelling immunofluorescent analyses of a cellular aggregation-prone protein marker. RESULTS: Aggresome formation was interrupted by VCR, whereas combination treatments with bortezomib involving suberoylanilide hydroxamic acid, 17-allylamino-17-demethoxy-geldanamycin, or clonazepam appear to facilitate the disposal of unfolded proteins via CMA, inducing HSP70 and lysosome-associated membrane protein type 2A (LAMP-2A). CONCLUSIONS: This schwannoma model can be used to test BiPN-reducing drugs. The present data suggest that aggresome formation in Schwann cells is a possible mechanism of BiPN, and drugs that induce HSP70 or LAMP-2A have the potential to alleviate this complication. Combination clinical trials are warranted to confirm the relevance of these observations.

The expression profile of functional regulatory T cells, CD4+CD25high+/forkhead box protein P3+, in patients with ulcerative colitis during active and quiescent disease.

Regulatory T cells (T(reg)) have an essential role in maintaining immune tolerance in the gut. The functional CD4(+) T(reg) express the transcription factor forkhead box protein 3 (FoxP3) or a CD25(high) in humans. Further, depletion of elevated granulocytes/monocytes by extracorporeal adsorption (GMA) induces immunomodulation in patients with ulcerative colitis (UC). We investigated the impact of GMA on T(reg). Thirty-one UC patients, clinical activity index (CAI) 12.1 +/- 2.97, refractory to conventional medications including intravenous corticosteroid and 13 healthy controls (HC), were included. Patients received five GMA sessions over 5 weeks. Biopsies from the rectal mucosa and blood samples at baseline and post-GMA were immunostained with anti-CD4/FoxP3 and anti-CD4/CD25 antibodies for immunohistochemistry and flow cytometry. Following GMA, 22 of 31 patients achieved remission (CAI

An organotypic culture system of Merkel cells using isolated epidermal sheets.

BACKGROUND: Merkel cells (MCs) exist in the epidermal basal layer, in contact with keratinocytes. This direct contact seems critical for maintaining MCs in vitro. OBJECTIVES: To estimate the effects of nerve cells on the maintenance of MCs within epidermal sheets in a new organotypic culture system of MCs. METHODS: We developed a new organotypic culture system of MCs, using MC-containing epidermal sheets embedded in collagen gel. To estimate the effects of nerve cells on the maintenance of MCs within the epidermal sheets, we cocultured nerve cells and MC-containing epidermal sheets. In these culture assemblies, cellular behaviour was analysed by histochemistry, immunohistochemistry, electron microscopy and enzyme-linked immunosorbent assay. RESULTS: This culture, even in the absence of neurotrophin (NT)-3 and nerve growth factor (NGF) (which are crucial for MC biology), retained cytokeratin (CK)-20-positive and neuroendocrine granule-containing MCs within the sheets for over 2 weeks. Coculture of MCs with PC-12 nerve cells significantly increased the number of MCs within the epidermal sheets, and the keratinocytes had almost identical expression levels of CK1, CK10, CK14 and the progenitor marker p63 to those produced by keratinocytes in vivo. Uptake of the growth marker bromodeoxyuridine by MCs and levels of NT-3 and NGF in the culture supernatants were undetectable in this system, regardless of the presence or absence of PC-12. CONCLUSIONS: The data suggest, first, that direct contact between MCs and keratinocytes may be critical for retaining MCs in vitro; second, that nerve cell-affected maintenance of keratinocyte differentiation, but not NT-3 and NGF, may contribute to MC maintenance; and third, that MCs are not able to grow, at least in our system. Our method would be useful for studying MC biology.

Prediction of hypotension during spinal anesthesia for elective cesarean section by altered heart rate variability induced by postural change.

BACKGROUND: Maternal hypotension is a common complication during cesarean section performed under spinal anesthesia. Changes in maternal heart rate with postural changes or values of heart rate variability have been reported to predict hypotension. Therefore, we hypothesized that changes in heart rate variability due to postural changes can predict hypotension. METHODS: A total of 45 women scheduled to undergo cesarean section under spinal anesthesia were enrolled. A postural change test was performed the day before cesarean section. The ratio of the power of low and high frequency components contributing to heart rate variability was assessed in the order of supine, left lateral, and supine. Patients who exhibited a ⩾two-fold increase in the low-to-high frequency ratio when moving to supine from the lateral position were assigned to the postural change test-positive group. RESULTS: According to the findings of the postural change test, patients were assigned to the positive (n=22) and negative (n=23) groups, respectively. Hypotension occurred in 35/45 patients, of whom 21 (60%) were in the positive group and 14 (40%) were in the negative group. The incidence of hypotension was greater in the positive group (P<0.01). The total dose of ephedrine was greater in the positive group (15±11 vs. 7±7mg, P=0.005). The area under the receiver operating characteristic curve was 0.76 for the postural change test as a predictor of hypotension. CONCLUSION: The postural change test with heart rate variability analysis may be used to predict the risk of hypotension during spinal anesthesia for cesarean section.

Relation between negative U waves in precordial leads on the admission electrocardiogram and time course of left ventricular wall motion in anterior wall acute myocardial infarction.

This study indicates that patients with anterior wall acute myocardial infarction showing negative U waves in the precordial leads on the admission electrocardiogram have greater improvement in left ventricular wall motion in the infarct region between 1 and 6 months after acute myocardial infarction. This suggests that these patients have a larger amount of stunned myocardium in the infarct region.

Significance of spontaneous normalization of negative T waves in infarct-related leads during healing of anterior wall acute myocardial infarction.

This study was conducted to elucidate the significance of spontaneous normalization of negative T waves in infarct-related leads during the chronic phase of anterior wall acute myocardial infarction. Results of this study indicate that patients with spontaneous normalization of negative T waves in infarct-related leads between 1 and 6 months after anterior wall acute myocardial infarction have smaller infarct size, decreased left ventricular dysfunction, and greater improvement in left ventricular wall motion in the infarct area, suggesting that T-wave normalization represents functional recovery of viable myocardium in the infarct area.

Relation of QT dispersion to infarct size and left ventricular wall motion in anterior wall acute myocardial infarction.

Previous studies have shown that QT dispersion increases during acute myocardial infarction (AMI). However, the relation of QT dispersion to infarct size and left ventricular (LV) function in AMI has not yet been fully clarified. Accordingly, this study was conducted to elucidate this relation at 1 month after anterior wall AMI. We examined 94 patients with first anterior wall AMI (< or = 6 hours) who underwent coronary arteriography at admission, 1 month, and 6 months after AMI, and left ventriculography at 1 and 6 months after AMI. Mean QT dispersion on the chronic phase (about 1 month after AMI) electrocardiogram was 79 +/- 33 ms. There were no significant correlations between QT dispersion and peak creatine phosphokinase levels, LV ejection fraction, and regional wall motion in the infarct region at 1 month after AMI (r = 0.06, p = 0.57; r = 0.11, p = 0.29; r = -0.05, p = 0.63, respectively). In conclusion, the findings of this study suggest that QT dispersion on the resting electrocardiogram at 1 month after anterior wall AMI is unrelated to infarct size estimated by the peak creatine phosphokinase level and the degree of LV dysfunction.

Significance of negative U waves in the precordial leads during anterior wall acute myocardial infarction.

This study was conducted to clarify the clinical significance of negative U waves in the precordial leads during anterior wall acute myocardial infarction (AMI). In all, 141 patients with first anterior wall AMI (< or = 6 hours) were classified into 2 groups according to the presence (group A, n = 31) or absence (group B, n = 110) of negative U waves in the precordial leads on the admission electrocardiogram (ECG). The number of leads showing ST elevation > or = 1 mm on the admission ECG was smaller in group A than in group B (5.2 +/- 1.3 vs 6.2 +/- 1.7, p < 0.01). Emergent coronary arteriography revealed that group A had a higher incidence of good collateral circulation than group B (39% vs 19%, p < 0.05). Peak creatine kinase activity was lower in group A than in group B (1,708 +/- 1,271 vs 2,735 +/- 1,865 IU/L, p < 0.01). The number of abnormal Q waves on the predischarge ECG was smaller in group A (2.0 +/- 1.5 vs 3.4 +/- 2.0, p < 0.01). Group A had a greater left ventricular ejection fraction and better regional wall motion in the anterobasal, anterolateral, and apical regions in the chronic phase than group B. In conclusion, patients with anterior wall AMI having negative U waves in the precordial leads on admission had a relatively smaller mass of necrotic myocardium than those without the waves. Therefore, negative U waves during anterior wall AMI may be a useful marker for identifying patients with smaller infarction partly due to better collateral circulation.

Emergent coronary angiographic findings of patients with ST depression in the inferior or lateral leads, or both, during anterior wall acute myocardial infarction.

In conclusion, the present study indicates that there are several distinctive differences in emergent coronary angiographic findings according to the presence or absence of ST depression in the inferior or lateral leads, or both, and location of the leads showing ST depression on admission electrocardiograms in patients with anterior AMI. The coronary angiographic features of patients with this ECG finding greatly support a poor prognosis. In patients with anterior AMI, analysis of ST depression on an admission electrocardiogram should be routinely performed because it is useful in predicting coronary anatomy, the extent of infarction, and its prognosis.

Metabolism of FK506, a potent immunosuppressive agent, by cytochrome P450 3A enzymes in rat, dog and human liver microsomes.

The oxidative metabolism of FK506 by liver microsomes and purified cytochrome P450 (P450) enzymes from rats, dogs and humans was studied. The major metabolite formed by liver microsomes from all species was 13-demethylated FK506, named M-I. In adult rats, liver microsomal metabolic activity toward FK506 was higher in males than in females and was stimulated by treatment with P450 3A inducers such as dexamethasone and phenobarbital. In a reconstituted monooxygenase system containing various forms of purified P450 3A enzymes, rat P450 3A2, dog P450 DPB-1 (a form of the P450 3A family) and human P450 3A4 catalyzed FK506 oxidation efficiently in the presence of cytochrome b5, a mixture of phospholipids (dilauroylphosphatidylcholine, dioleoylphosphatidylcholine and phosphatidylserine), and sodium cholate. Rat P450 2C6 and 2D1 and human P450 2CMP also metabolized FK506, with significant lower activity than the P450 3A enzymes, and other rat P450 1A, 2A, 2B, 2C and 2E families including C11 did not show catalytic activities for FK506. Anti-P450 3A2 and anti-P450 3A4 antibodies strongly inhibited FK506 oxidation catalyzed by rat and human liver microsomes, respectively. The formation rate of M-I correlated well with testosterone 2 beta- and 6 beta-hydroxylase activities in rat liver microsomes and with immunoquantified P450 3A4 content, nifedipine oxidase activity, and testosterone 6 beta-hydroxylase activity in human liver microsomes. These in vitro findings indicate that the P450 3A enzymes in liver microsomes from various species of animals, including human, play a major role in the first step oxidation of FK506.

Isolation of antigen from the circulating immune complex in mice infected with Plasmodium berghei.

Circulating immune complex (CIC) is known to play a role in pathological glomerular alterations in malaria. However, the nature of the antigens comprising the CIC is still not fully understood. We report here the isolation of the antigen in CIC and its localisation in mice infected with Plasmodium berghei NK65. The antigen was successfully isolated from CIC extracted from the blood of mice infected with P. berghei, by using C1q-coated microplates. The molecular mass of the antigen separated from CIC bound to C1q was found to be 78 kDa. Furthermore, localisation of the antigen was examined by the fluorescent antibody technique and immunoelectron microscopy. The antigen was detected in the parasitised erythrocyte and the mesangial matrix by both methods. These results suggest that the 78 kDa protein might be associated with the glomerular alterations in malaria infection.

High-level expression of human BSF-2/IL-6 cDNA in Escherichia coli using a new type of expression-preparation system.

BSF-2 (B cell stimulatory factor-2/IL-6) is a member of the lymphokine family and responsible for B cell differentiation. Expression plasmids of human BSF-2 cDNA were constructed using a trp promotor/operator and a trpA terminator. In an extract of Escherichia coli HB101 holding "direct" expression plasmid pBSF-2D, activity of BSF-2 was detected, but overproduction was not observed. A "fused" expression system was therefore developed to prepare the recombinant protein. In this system, cDNA was expressed as a fused protein with human IL-2 N-terminal peptide. In the case of the fused BSF-2 expression plasmid, pBSF-2F, inclusion bodies were observed and overproduction of the protein occurred. As this fused protein had a Phe-Arg-Ala sequence at the junction of hIL-2 and BSF-2, it was possible to process mature BSF-2 from the fused BSF-2 by treatment with kallikrein and aminopeptidase P. From 1 liter of E. coli culture, 45 mg of mature BSF-2 was purified; it had a relative biological activity equal to that of natural BSF-2 purified from T cells.

Renal hematuria caused by "nutcracker" phenomenon: a more logical surgical management.

A case of renal hematuria caused by the "nutcracker" phenomenon in a young man is reported. Resection of a preaortic fibrous tissue, renocaval venous reimplantation, and placement of a synthetic wedge into the bifurcation of the superior mesenteric artery were performed to resume an unstagnant flow of the left renal vein.

A non-competitive solid-phase radioimmunoassay for human aldolase A.

A solid-phase, non-competitive radioimmunoassay for aldolase A in human serum has been developed. Human aldolase A was purified from muscle, and specific antisera to the purified aldolase A were obtained from chickens. Specific IgG anti-human aldolase A was purified by affinity chromatography. Disposable polypropylene plates were coated with specific IgG antibody and used for radioimmunoassay with 125I-specific IgG antibody to aldolase A. The non-specific binding was minimized by saturating the binding sites of the plates with 2% ovalbumin in 0.1% Tween 20. This radioimmunoassay is specific for the aldolase A subunit, with no cross-reactivity with human aldolase B subunit or homopolymeric human aldolase C(C4). The serum aldolase A immunoreactivities of 33 normal subjects ranged from 124 to 212 ng/ml with a mean of 178 +/- 41 ng/ml (+/- 2 SD). Ninety-three patients' sera were assayed with both a solid-phase non-competitive radioimmunoassay and a competitive double antibody radioimmunoassay developed in our laboratory and the results showed a high degree of correlation (r = 0.912; p less than 0.001). Rapidity and simplicity of the solid-phase assay makes it superior to other methods for the measurement of serum aldolase isozymes.

Radioimmunoassay for human aldolase A.

A radioimmunoassay was developed for the direct quantification of aldolase A in human serum. The method is a double antibody radioimmunoassay using radioiodinated aldolase A4 homopolymer as ligand, chicken antibodies to aldolase A, and rabbit antibodies to chicken IgG. The lowest measurable amount by this method was 2 ng (0.01 U). The radioimmunoassay was shown to be specific for the aldolase A subunit, with no cross-reactivity with human aldolase B subunits or homopolymeric human aldolase C (C4). The immunoreactive aldolase A in the sera of 41 normal healthy subjects ranged from 130 to 210 ng/ml (0.81-1.31 U/1), with a mean of 171 /+- 39 ng/ml.

Distribution and protein binding of FK506, a potent immunosuppressive macrolide lactone, in human blood and its uptake by erythrocytes.

The distribution of FK506 in the blood was estimated in-vitro. At a drug level of 5 ng mL-1, FK506 mainly distributed in erythrocytes (95-98%) in dog, monkey and human blood, and its distribution was affected by drug concentration, temperature, and haematocrit values. In erythrocytes most of FK506 was distributed in cytoplasmic components and was bound strongly to a protein having a molecular weight of 10-11 kDa. The molecular weight of this protein agrees with FK506-binding protein found in various cells. Greater than 98.8% of FK506 was bound to the plasma proteins in all species studied. FK506 bound to various plasma proteins such as lipoproteins, globulins, alpha 1-acid glycoprotein and albumin.

Conditionally immortalized brain capillary endothelial cell lines established from a transgenic mouse harboring temperature-sensitive simian virus 40 large T-antigen gene.

Five immortalized brain capillary endothelial cell lines (TM-BBB1-5) were established from 3 transgenic mice harboring temperature-sensitive simian virus 40 large T-antigen gene (Tg mouse). These cell lines expressed active large T-antigen and grew well at 33 degrees C with a doubling time of about 20 to 30 hours. TM-BBBs also grew at 37 degrees C but not at 39 degrees C. However, growth was restored when the temperature of the culture was lowered to 33 degrees C. Although significant amounts of large T-antigen were shown to be present in the cell culture at 33 degrees C, there was less of this complex at 37 degrees C and 39 degrees C. TM-BBBs expressed the typical endothelial marker, von Willebrand factor, and exhibited acetylated low-density lipoprotein uptake activity. The alkaline phosphatase and gamma-glutamyltranspeptidase activity in TM-BBBs were -10% and 50% to 80% of brain capillary fraction of normal mice, respectively. D-mannitol transport in the both apical-to-basal and basal-to-apical directions across the TM-BBB was 2-fold greater than for inulin. TM-BBBs were found to express GLUT-1 but not GLUT-3, and exhibited concentration-dependent 3-O-methyl-D-glucose (3-OMG) uptake activity with a Michaelis-Menten constant of 6.59 +/- 1.16 mmol/l. Moreover, P-glycoprotein (P-gp) with a molecular weight of -170 kDa was expressed in all TM-BBBs. Both mdr1a and mdr1b mRNA were detected in TM-BBB4 using reverse transcription-polymerase chain reaction (RT-PCR) analysis. [3H]-Cyclosporin A uptake by TM-BBB was significantly increased in the presence of 100 micromol/l verapamil and vincristine, suggesting that TM-BBB exhibits efflux transport activity via P-gp. In conclusion, conditional brain capillary endothelial cell lines were established from Tg mice. This cell line expresses endothelial markers and transporters at the BBB and is able to regulate cell growth, due to the amount of active large T-antigen in the cell, by changing the culture temperature.