Intratumoural immune cell landscape in germinoma reveals multipotent lineages and exhibits prognostic significance.

AIMS: Alterations in microenvironments are a hallmark of cancer, and these alterations in germinomas are of particular significance. Germinoma, the most common subtype of central nervous system germ cell tumours, often exhibits massive immune cell infiltration intermingled with tumour cells. The role of these immune cells in germinoma, however, remains unknown. METHODS: We investigated the cellular constituents of immune microenvironments and their clinical impacts on prognosis in 100 germinoma cases. RESULTS: Patients with germinomas lower in tumour cell content (i.e. higher immune cell infiltration) had a significantly longer progression-free survival time than those with higher tumour cell contents (P = 0.03). Transcriptome analyses and RNA in-situ hybridization indicated that infiltrating immune cells comprised a wide variety of cell types, including lymphocytes and myelocyte-lineage cells. High expression of CD4 was significantly associated with good prognosis, whereas elevated nitric oxide synthase 2 was associated with poor prognosis. PD1 (PDCD1) was expressed by immune cells present in most germinomas (93.8%), and PD-L1 (CD274) expression was found in tumour cells in the majority of germinomas examined (73.5%). CONCLUSIONS: The collective data strongly suggest that infiltrating immune cells play an important role in predicting treatment response. Further investigation should lead to additional categorization of germinoma to safely reduce treatment intensity depending on tumour/immune cell balance and to develop possible future immunotherapies.

Pre-mRNA Processing Factor Prp18 Is a Stimulatory Factor of Influenza Virus RNA Synthesis and Possesses Nucleoprotein Chaperone Activity.

The genome of influenza virus (viral RNA [vRNA]) is associated with the nucleoprotein (NP) and viral RNA-dependent RNA polymerases and forms helical viral ribonucleoprotein (vRNP) complexes. The NP-vRNA complex is the biologically active template for RNA synthesis by the viral polymerase. Previously, we identified human pre-mRNA processing factor 18 (Prp18) as a stimulatory factor for viral RNA synthesis using a Saccharomyces cerevisiae replicon system and a single-gene deletion library of Saccharomyces cerevisiae (T. Naito, Y. Kiyasu, K. Sugiyama, A. Kimura, R. Nakano, A. Matsukage, and K. Nagata, Proc Natl Acad Sci USA, 104:18235-18240, 2007, https://doi.org/10.1073/pnas.0705856104). In infected Prp18 knockdown (KD) cells, the synthesis of vRNA, cRNA, and viral mRNAs was reduced. Prp18 was found to stimulate in vitro viral RNA synthesis through its interaction with NP. Analyses using in vitro RNA synthesis reactions revealed that Prp18 dissociates newly synthesized RNA from the template after the early elongation step to stimulate the elongation reaction. We found that Prp18 functions as a chaperone for NP to facilitate the formation of NP-RNA complexes. Based on these results, it is suggested that Prp18 accelerates influenza virus RNA synthesis as an NP chaperone for the processive elongation reaction. IMPORTANCE: Templates for viral RNA synthesis of negative-stranded RNA viruses are not naked RNA but rather RNA encapsidated by viral nucleocapsid proteins forming vRNP complexes. However, viral basic proteins tend to aggregate under physiological ionic strength without chaperones. We identified the pre-mRNA processing factor Prp18 as a stimulatory factor for influenza virus RNA synthesis. We found that one of the targets of Prp18 is NP. Prp18 facilitates the elongation reaction of viral polymerases by preventing the deleterious annealing of newly synthesized RNA to the template. Prp18 functions as a chaperone for NP to stimulate the formation of NP-RNA complexes. Based on these results, we propose that Prp18 may be required to maintain the structural integrity of vRNP for processive template reading.

The efficacy of add-on tacrolimus for minor flare in patients with systemic lupus erythematosus: a retrospective study.

OBJECTIVE: We have assessed the effectiveness of tacrolimus for minor flares in systemic lupus erythematosus (SLE) patients. METHODS: The medical records of 313 patients were retrospectively reviewed over a period of seven years, from 2006 to 2013. We enrolled patients with minor flare treated with add-on tacrolimus, without glucocorticoid (GC) intensification (tacrolimus group). Minor flare was defined as a ≥ 1-point increase in a total score between 3 and 11 in the SLE Disease Activity Index (SLEDAI). We enrolled as controls patients who were administered increased doses of GC for minor flare (GC group). All patients were followed for one year. The primary outcome measure was the proportion of responders. RESULTS: There were 14 eligible patients in the tacrolimus group and 20 eligible patients in the GC group. The mean SLEDAI at flare tended to be higher in the tacrolimus group than in the GC group (7.5 vs. 6.2, p = 0.085). A mean dose of 1.6 mg tacrolimus/day was administered for flare, while the mean GC dose was 13.7 mg/day in the GC group. The proportion of responders was 86% (12/14) in the tacrolimus group and 75% (15/20) in the GC group (p = 0.67). The mean dose of GC at 12 months was higher in the GC group than in the tacrolimus group (9.7 mg/day vs. 7.1 mg/day, p < 0.05). Only one patient discontinued tacrolimus because of fatigue after three months. CONCLUSION: Adding tacrolimus without increasing the GC dose may provide an effective treatment option for minor flares in patients with SLE.

Rehabilitation for paraplegia caused by neuromyelitis optica: a case report.

STUDY DESIGN: Single case report. OBJECTIVE: We present a case of paraplegia due to neuromyelitis optica (NMO) with poor rehabilitation outcome. SETTING: University hospital, Japan. CASE REPORT: A 27-year-old woman with NMO presented with T5 paraplegia of ASIA impairment scale grade A. Spinal cord magnetic resonance imaging revealed a lesion spanning C3 to L1 level. After acute phase treatment, flaccid paraplegia below T5 and a T2-weighted hyperintense lesion from T6 to T10 level remained. Rehabilitation aimed at independence of activities of daily living with wheelchair assistance, including transfer activity, was provided for 19 months. However, flaccid paralysis of the trunk and limbs persisted, and safe independent transfer was not achieved. CONCLUSION: Spinal lesions spanning many vertebral segments, a characteristic of NMO, can cause extensive flaccid paralysis of the trunk and limbs. Rehabilitation may achieve poorer functional recovery than that for spinal cord injury.

High-power laser experiment on developing supercritical shock propagating in homogeneously magnetized plasma of ambient gas origin.

A developing supercritical collisionless shock propagating in a homogeneously magnetized plasma of ambient gas origin having higher uniformity than the previous experiments is formed by using high-power laser experiment. The ambient plasma is not contaminated by the plasma produced in the early time after the laser shot. While the observed developing shock does not have stationary downstream structure, it possesses some characteristics of a magnetized supercritical shock, which are supported by a one-dimensional full particle-in-cell simulation taking the effect of finite time of laser-target interaction into account.

High-power laser experiment forming a supercritical collisionless shock in a magnetized uniform plasma at rest.

We present an experimental method to generate quasiperpendicular supercritical magnetized collisionless shocks. In our experiment, ambient nitrogen (N) plasma is at rest and well magnetized, and it has uniform mass density. The plasma is pushed by laser-driven ablation aluminum (Al) plasma. Streaked optical pyrometry and spatially resolved laser collective Thomson scattering clarify structures of plasma density and temperatures, which are compared with one-dimensional particle-in-cell simulations. It is indicated that just after the laser irradiation, the Al plasma is magnetized by a self-generated Biermann battery field, and the plasma slaps the incident N plasma. The compressed external field in the N plasma reflects N ions, leading to counterstreaming magnetized N flows. Namely, we identify the edge of the reflected N ions. Such interacting plasmas form a magnetized collisionless shock.

Over-expression of the LTC4 synthase gene in mice reproduces human aspirin-induced asthma.

BACKGROUND: The pathogenesis of aspirin-induced asthma (AIA) is presumed to involve the aspirin/non-steroidal anti-inflammatory drug (NSAID)-induced abnormal metabolism of arachidonic acid, resulting in an increase in 5-lipoxygenase (5-LO) metabolites, particularly leukotriene C(4) (LTC(4) ). However, the role of LTC(4) in the development of AIA has yet to be conclusively demonstrated. OBJECTIVE: The aim of this study was to evaluate the contribution of the lipid product LTC(4) secreted by the 5-LO pathway to the pathogenesis of AIA. METHODS: To evaluate antigen-induced airway inflammation, the concentrations of T-helper type 2 cytokine in bronchoalveolar lavage fluid (BALF) obtained from LTC(4) synthase-transgenic (Tg) and wild-type (WT) mice after challenge with ovalbumin were measured. Subsequently, the ex vivo and in vivo effects of the NSAID sulpyrine were investigated in these Tg and WT mice by measuring the secretion of LTC(4) from sulpyrine-treated BAL cells and the levels of LTC(4) in BALF following challenge with sulpyrine. Finally, the sulpyrine-induced airway response by the administration of pranlukast, an antagonist of the cysteinyl (cs)-LT1 receptor, was analysed. RESULTS: The concentrations of IL-4, -5, and -13 in BALF from Tg mice were significantly higher than those in WT mice. In addition, sulpyrine augmented the secretion of LTC(4) in BALF and by BAL cells in Tg mice, but not in WT mice. Additionally, the increased airway resistance induced by sulpyrine could be reduced by treatment with pranlukast. Furthermore, the secretion of LTC(4) from mast cells, eosinophils, and macrophages was increased in the allergen-stimulated LTC(4) synthase gene Tg mice, even in the absence of sulpyrine, as well as in BAL cells after sulpyrine. CONCLUSION AND CLINICAL RELEVANCE: The over-expression of the LTC(4) synthase in a mouse asthma model also replicates the key features of AIA. And our study supports that cys-LTs play a major role in the pathogenesis of AIA in patients with chronic asthma.

Correlation between the pharmacological efficacy of cyclosporine and tacrolimus as evaluated by the lymphocyte immunosuppressant sensitivity test (LIST) and the MTT assay procedure in patients before and after renal transplantation.

OBJECTIVES: Cyclosporine and tacrolimus are calcineurin inhibitors that are used to prevent acute rejection in renal transplant recipients. The lymphocyte immunosuppressant sensitivity test (LIST) can predict the pharmacological efficacy of these immunosuppressive agents for renal transplant recipients. There is a correlation between cyclosporine and tacrolimus pharmacological efficacy as evaluated by LIST by the 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide (MTT) assay procedure prior to renal transplantation. However, the LIST can also evaluate patients before and after the transplantation. MATERIALS AND METHODS: The present study examined the relationship between cyclosporine and tacrolimus pharmacological efficacy by LIST using the MTT assay in 16 renal transplant recipients at 1, 3 and 12 months after transplantation, as well as before the operation. RESULTS: The relationship of cyclosporine and tacrolimus pharmacological efficacy gave a significant Kendall and Spearman's coefficient correlation in these transplant recipients by the LIST using the MTT assay procedure immediately prior to renal transplantation (rk = 0.711, rs = 0.877, p < 0.01). Furthermore, correlations between the cyclosporine and tacrolimus IC50 values were also observed with a significant Kendall and Spearman's coefficient correlation at 1 and 12 months after transplantation (rk1month = 0.65, rs1month = 0.829, p < 0.01, and k12month = 0.433, rs12month = 0.603, p < 0.01, respectively). However, no statistically significant relationship was observed between the pharmacological efficacies of the calcineurin inhibitors at 3 months after transplantation (rk3month = 0.117, rs3month = 0.1, p > 0.05). CONCLUSIONS: Both cyclosporine and tacrolimus exhibit pharmacological efficacy by the inhibition of calcineurin. However, the correlation between cyclosporine and tacrolimus pharmacological efficacies may be altered, due to immunosuppressive therapy or clinical events at 3 months after renal transplantation.

Modulation of exteroceptive suppression periods in human jaw-closing muscles by subanaesthetic dose of propofol.

Exteroceptive suppression (ES) periods in human jaw-closing muscles can be conditioned by a wide range of somatosensory stimuli and cognitive states. The aim of this study was to examine the effects of subanaesthetic doses of midazolam, ketamine and propofol on the short latency (ES1) and long latency (ES2) reflex in the jaw-closing muscles. First, we tried to evaluate the various methodological criteria for ES recording. We then examined the effect of subanaesthetic doses of midazolam (0·035 mg kg(-1)), ketamine (0·30 mg kg(-1)) and propofol (0·35 mg kg(-1)) on these reflexes of recording left masseter and temporalis muscle. ES duration did not differ greatly in the present study, recorded with the correct adjustment of stimulating and recording conditions. None of the subanaesthetic doses of the agents influenced ES1, and no significant effects on ES2 were observed with midazolam and ketamine. However, significant inhibitory change was observed in ES2 with propofol. ES2 is thought to be mediated by afferents, which descend in the spinal trigeminal tract and connect with a polysynaptic chain of excitatory interneurones located in the lateral reticular formation. Our observations indicate that propofol is uniquely effective not only through involvement of the gamma-aminobutyric acid type A receptor, but also through a range of other effects.

Differences in the pharmacokinetics of Cyp3a substrates in TSOD and streptozotocin-induced diabetic mice.

The pharmacokinetics of drugs can change in diabetes mellitus and even among diabetics. They may differ between type I diabetes (T1DM) and type 2 diabetes (T2DM). As triazolam was administered orally to Tsumura, Suzuki, obese, diabetes (TSOD) mice and streptozotocin (STZ) mice, clearance per body (CL/F) in TSOD mice did not differ compared with Tsumura, Suzuki, non-obesity (TSNO) mice. In STZ mice, CL/F was greater than in control mice. Small intestinal cytochrome P450 (Cyp) 3a expression in TSOD mice was significantly lower than in TSNO mice. No significant difference existed in small intestinal Cyp3a expression between STZ mice and control mice. In insulin-treated mice, small intestinal Cyp3a expression was significantly lower than in control mice. These results suggested that the differences in changes in small intestinal Cyp3a expression between T1DM and T2DM may be due to differences in plasma insulin concentrations. This may be a factor in the difference in the drug pharmacokinetics between T2DM and T1DM patients.

Development of a semi-quantitative evaluation system for surveillance of bovine spongiform encephalopathy, using the analytic hierarchy process.

Evaluating surveillance results is important in estimating the risk of bovine spongiform encephalopathy (BSE) cases in a particular country, or in countries from which products are imported. Although various methods have been proposed for quantitative evaluation of surveillance results, no methods focusing on surveillance from a qualitative perspective have yet been established. The authors have developed an objective method for evaluating the qualitative aspects of BSE surveillance, based on the analytic hierarchy process. Factors related to surveillance credibility were selected through expert meetings and arranged in a hierarchical structure. These evaluation factors were also weighted, so that a points system could be used for evaluation. As a result, 13 evaluation factors comprising three-layer hierarchies were generated. When surveillance in Japan before and after a BSE case was evaluated using this evaluation system, an improvement in the quality of the surveillance was observed after the outbreak. Although this study suggests that the selection of the experts had a significant effect on the outcome, the authors believe that this method will also be applicable for establishing qualitative evaluation systems for other diseases.

FcepsilonRI-mediated thymic stromal lymphopoietin production by interleukin-4-primed human mast cells.

A significant increase of mRNA expression of thymic stromal lymphopoietin (TSLP) has been reported in the bronchial mast cells (MCs) of asthmatic subjects; however, the mechanism underlying the upregulation of TSLP mRNA and protein remains unknown. FcepsilonRI-mediated activation of human MCs upregulated TSLP mRNA expression by 5.2+/-2.9-fold, while activation of the MCs using lipopolysaccharide and polyriboinosinic:polyribocytidylic acid failed to upregulate TSLP. Stimulation of MCs with interleukin (IL)-4 alone did not affect the TSLP mRNA expression, while pre-incubation of MCs with IL-4 for 48 h significantly enhanced the FcepsilonRI-mediated TSLP mRNA expression (by 53.7+/-15.9-fold; p<0.05) and the amount of TSLP in the cell pellets increased significantly from 23.4+/-4.3 pg mL(-1) to 121.5+/-3.7 pg mL(-1) (p<0.0001). However, the released TSLP was rapidly degraded by proteases that were released by MCs. We identified the population of cells expressing TSLP in the lungs of 16 asthmatic and 11 control subjects by immunohistochemistry. The percentage of TSLP-positive MCs in the total population of MCs was significantly increased in asthmatic airways (p<0.0001). Thus, MCs are able to store TSLP intracellularly and to produce TSLP following aggregation of FcepsilonRI in the presence of IL-4.

Laser-filament-induced corona discharges and remote measurements of electric fields.

Femtosecond laser pulses were used to make plasma filaments near an isolated positively or negatively highly biased electrode. The electrode was well positioned to sustain a high voltage up to U(max)=+/-400 kV to avoid the induced breakdown or a glow discharge; the shape of the electrode was chosen to reduce the corona effects at the maximal voltage. The filament's UV emission is shown to be very sensitive to the voltage applied: it increases nonlinearly with the electrode potential. Along with nanosecond filament-induced flashes at both polarities, long, about a half microsecond, corona flashes were observed at the negative polarity.

Altered expression of CYP in TSOD mice: a model of type 2 diabetes and obesity.

To investigate the pharmacokinetic characteristics in TSOD (Tsumura, Suzuki, obese, diabetes) mice, a model of type 2 diabetes and obesity, the expressions of major hepatic CYP enzymes in TSOD and TSNO (Tsumura, Suzuki, non-obesity; control) mice were compared. The 7-month-old TSOD mice, which represented severe obesity/diabetes-related pathophysiology, showed higher expressions of Cyp2c and Cyp3a compared with TSNO mice, while those of Cyp1a and Cyp2e were lower. Cyp3a metabolic activity was also higher in TSOD mice. In the 7-month-old liver, pregnane X receptor (PXR) (nuclear receptor) and peroxisome proliferator-activated receptor-gamma coactivator-1alpha (PGC-1alpha) (cofactor) mRNA expression were higher in TSOD mice, possibly playing a role in the altered expression of Cyp3a. This specifically altered CYP expression in TSOD mice suggests that the biotransformation of drugs metabolized by these CYP enzymes differs from that in normal animals. Based on these findings, further investigation on the relationship between altered CYP expression and pathophysiology may be useful in elucidating changes in pharmacokinetics in obese/diabetic patients.

Intestinal flora induces the expression of Cyp3a in the mouse liver.

In order to determine the effects of intestinal flora on the expression of cytochrome P450 (CYP), the mRNA expression of CYP was compared between specific pathogen-free (SPF) and germ-free (GF) mice. Most of the major CYP isozymes showed higher expression in the livers of SPF mice compared with GF mice. Nuclear factors such as pregnane X receptor (PXR) and constitutive androstane receptor (CAR), as well as transporters and conjugation enzymes involved in the detoxification of lithocholic acid (LCA), also showed higher expression in SPF mice. The findings suggest that in the livers of SPF mice, LCA produced by intestinal flora increases the expression of CYPs via activation of PXR and CAR. Drugs such as antibiotics, some diseases and ageing, etc. are known to alter intestinal flora. The present findings suggest that such changes also affect CYP and are one of the factors responsible for individual differences in pharmacokinetics.

Combining the EndoFlex tube with fiberoptic bronchoscopy in difficult intubation.

We applied a combination technique using the EndoFlex tube with fiberoptic bronchoscopy for a 69-year-old man presenting with limited mouth opening and neck movement. Awake nasotracheal intubation was performed under conscious sedation with propofol and fentanyl. After positioning the tip of the EndoFlex tube in the oropharynx, the fiberoptic bronchoscope was inserted into the tube until the tip reached the bevel of the tube. Anterior flexion of the distal tip of the EndoFlex tube facilitated uncomplicated insertion of the tube into the trachea without impingement on the arytenoids. Fiberoptic visualization confirmed that the distal-tip flexing mechanism of the EndoFlex tube corrected the direction of the tube tip anteriorly, allowing entry into the trachea. We present a case where this technique proved valuable for tracheal intubation in a patient with limitations of mouth opening and neck movement.

Cyclosporine pharmacological efficacy estimated by lymphocyte immunosuppressant sensitivity test before and after renal transplantation.

OBJECTIVE: Lymphocyte immunosuppressant sensitivity test (LIST) is useful for predicting the pharmacological efficacy of immunosuppressive agents. In this study, the pharmacological efficacy of cyclosporine was estimated by LIST before and after renal transplantation. METHODS: Lymphocyte immunosuppressant sensitivity test was performed by the 3-(4,5-dimethylthiazol-2-yl)-2,5diphenyltetrazolium bromide (MTT) assay before and at 1, 3, and 12 months after transplantation in 19 consecutive renal transplant recipients. RESULTS: There was wide intersubject variability in cyclosporine IC50 before transplantation [Mean (SD) of 593.9 (1067.6) ng/mL]. This variability worsened 1 month after transplantation [525.7 (1532.7) ng/mL] but decreased at 3 months (193.5 (347.9) ng/mL) and 12 months (75.4 (95.4) ng/mL). In this small study, observed differences in IC50 values for the individual subjects at various time intervals was not associated with the occurrence of rejection, graft loss, and infection episodes. CONCLUSION: Lymphocyte sensitivity to cyclosporine assessed by the LIST assay showed a high level of inter-subject variability particularly before and 1 month after transplantation. The observed difference in IC50 values was not associated with clinical outcome in this small study.

DUSP22/LMW-DSP2 regulates estrogen receptor-alpha-mediated signaling through dephosphorylation of Ser-118.

In the previous study, we demonstrated the involvement of dual specificity phosphatase 22 (DUSP22/LMW-DSP2) in regulating the leukemia inhibitory factor/interleukin-6/signal transducer and activator of transcription 3-mediated signaling pathway. In this study, we show beta-estradiol (E2)-induced DUSP22 mRNA expression in estrogen receptor alpha (ERalpha)-positive breast cancer cells, whereas E2-induced phosphorylation and activation of ERalpha was suppressed by overexpression of DUSP22 but not catalytically inactive mutants. Furthermore, small-interfering RNA-mediated reduction of DUSP22 expression enhanced ERalpha-mediated transcription and endogenous gene expression. In fact, DUSP22 associated with ERalpha in vivo and both endogenous proteins interacted in ERalpha-positive breast cancer T47D cells. These results strongly suggest that DUSP22 acts as a negative regulator of the ERalpha-mediated signaling pathway.