Acetazolamide enhances the release of urinary exosomal aquaporin-1.

BACKGROUND: Renal aquaporin-1 (AQP1), a water channel protein, is known to be secreted into urine, conveyed by nano-sized extracellular vesicles called exosomes. A previous study has demonstrated that acetazolamide (AZ), a diuretic that inhibits carbonic anhydrases, alters the expression level of AQP1 in cultured cells. Here we investigated whether AZ alters the release of urinary exosomal AQP1 in vivo. METHODS: The effect of AZ on urinary exosomal AQP1 secretion was examined in rats and compared with furosemide (another diuretic), NaHCO3 (an alkalizing agent) and NH4Cl (an acidifying agent). Urine, blood and kidney samples were obtained 2 h after each treatment. Urinary exosomes were isolated by a differential centrifugation technique and urinary exosomal proteins were analyzed by immunoblotting. RESULTS: The release of exosomal AQP1 into urine was markedly increased after treatment with AZ, accompanied by alkaluria and metabolic acidosis. Immunohistochemistry clearly demonstrated that AZ increased the apical membrane expression of AQP1 in the proximal tubules. AZ did not affect the release of exosomal marker proteins (tumor susceptibility gene 101 protein and apoptosis-linked gene 2 interacting protein X). Treatment with furosemide did not change, whereas NaHCO3 and NH4Cl decreased the exosomal release of AQP1. CONCLUSION: The present findings indicate that AZ increases the release of exosomal AQP1 into urine in association with enhanced apical membrane expression of AQP1.

Changes in Compression Pressure of Elastic Stockings for the Lower Limbs During Cesarean Section: A Prospective Observational Study.

Background Postpartum peripheral nerve injuries can impact recovery. Elastic stockings are recommended for thromboembolism prevention, although concerns about entrapment neuropathy exist. In this prospective observational study, we investigated the differential compressions caused by wearing elastic stockings before and after anesthesia, as well as changes in the diameters of the lower leg and ankle in parturient women undergoing spinal anesthesia for elective cesarean section (CS). Methods Eighteen pregnant women, classified by the American Society of Anesthesiologists as having physical status 2, underwent lower leg measurements taken before a CS. Elastic stockings were applied, and compression pressure was measured at pre-anesthesia, post-surgery, and six hours post-return to a hospital room. Fluid, blood loss, urine output, and neuropathy presence were recorded. For all parameters, changes at the three time points were compared for the primary analysis. For secondary analysis, participants were categorized as having intraoperative blood loss greater than (group P) or less than 1,000 g (group N), and factors were compared with pre-anesthesia and six hours post-return to a room. Data were analyzed and presented using a one-way analysis of variance with Bonferroni correction for multiple comparisons or unpaired two-tailed t-tests for pairwise comparison. Results None of the women had postoperative entrapment neuropathy. Six patients had >1,000 g of blood loss. Compression significantly increased from pre-anesthesia (left 13.6 ± 2.4, 95% CI: 12.18 to 14.52; right 13.4 ± 2.4, 95% CI: 12.41 to 14.69) to post-surgery (left, 17.4 ± 2.6, 95% CI: 15.68 to 18.12; right, 16.9 ± 2.6, 95% CI: 16.20 to 18.70) (p < 0.01). Compression pressure at post-surgery differed significantly between group P (left, 15.3 ± 1.3; right, 14.7 ± 1.8; 95% CI: -4.98 to -0.32) and group N (left, 18.1 ± 2.9; right, 17.8 ± 2.4; 95% CI: -5.38 to -0.26) (p < 0.05). The results are expressed as mean ± standard deviation, with P-values <0.05 indicating statistical significance. Conclusions In this study, no neuropathy occurred; however, over-compression risk with elastic stockings, especially when exceeding recommended pressure levels, was highlighted. Balancing thromboembolism prevention and over-compression risks is crucial for patients undergoing CSs with spinal anesthesia.

Excretion of urinary exosomal AQP2 in rats is regulated by vasopressin and urinary pH.

Urinary exosomes are small vesicles secreted into urine from all renal epithelial cell types and known to contain proteins that are involved in renal secretion and reabsorption. Among these proteins, urinary exosomal aquaporin-2 (AQP2) has been suggested to be useful for diagnosis of renal disease. However, the mechanisms underlying the excretion of urinary exosomal AQP2 are largely unknown. In this study, we examined the mechanisms of urinary exosomal AQP2 excretion in vivo, using diuretics including furosemide (FS), an inhibitor of the sodium-potassium-chloride symporter; acetazolamide (ACTZ), an inhibitor of carbonic anhydrase; OPC-31260 (OPC), a vasopressin type 2 receptor antagonist; and NaHCO3, a urinary alkalizing agent. Samples of urine from rats were collected for 2 h just after treatment with each diuretic, and urinary exosomes were isolated by ultracentrifugation. Urinary exosomal AQP2 excretion was dramatically increased by treatment with FS accompanied by urine acidification or with ACTZ accompanied by urine alkalization. Immunohistochemistry showed that apical localization of AQP2 was clearly evident and the plasma vasopressin level was increased after each treatment. Although treatment with OPC alone had no significant effect, coadministration of OPC completely inhibited the FS-induced and partially reduced the ACTZ-induced responses, respectively. Treatment with NaHCO3 increased the excretion of urinary exosomal AQP2 accompanied by urine alkalization. This increased response was partially inhibited by coadministration of OPC. These data suggest that an increased plasma level of vasopressin promoted the excretion of urinary exosomal AQP2 and that urine alkalinization also increased it independently of vasopressin.

Fugacity model incorporating computational fluid dynamics for analyzing the behavior of an insecticide sprayed indoors.

Fugacity models are used widely to predict the time-dependent behaviors of chemicals in environments containing several media (e.g., air, sediment, soil, and water). However, these fugacity models work on the assumption that the concentration of a chemical in each medium is uniform, so they cannot describe the spatial distribution of the chemical. We developed a new fugacity model, termed InPestCFD, incorporating computational fluid dynamics to describe both the time-dependent distribution and the spatial distribution of a chemical in a medium. InPestCFD was used to calculate the behavior of an insecticide released from an aerosol canister in a room. Indoor airflow and aerosol particle behavior were calculated via computational fluid dynamics and using a Lagrangian dispersion model. Transport of the insecticide among media (aerosol particles, air, ceiling, floor, and walls) was calculated using the fugacity model. The time-dependent distributions and spatial distributions of the insecticide in the media agreed well with real measurements.

Loss of disability-adjusted life years due to heat-related sleep disturbance in the Japanese.

The purpose of this study was to quantify the sleep disturbances caused by climate change using disability-adjusted life years (DALY). The revised sleep quality index for daily sleep (SQIDS2), a self-administered questionnaire for daily sleep quality, was developed to assess daily sleep disturbances. This questionnaire referenced and simplified the Pittsburgh Sleep Quality Index (PSQI). This study was conducted in Nagoya City in August 2011 and 2012. Sleep quality was measured using SQIDS2 and PSQI. A total of 574 participants in 2011 and 710 in 2012 responded to the survey. The sleep disturbance prevalence calculated from the SQIDS2 score was correlated with the daily minimum temperature (p = 0.0067). This score increased when the daily minimum temperature was above 24.8 °C. When correcting for the PSQI score, DALY loss due to heat-related sleep disturbances in Nagoya City (population: 2,266,851) was estimated to be 81.8 years in 2012. This value was comparable to the DALY loss due to heatstroke. Sleep disturbance due to climate change was quantified using the DALY based on the PSQI. Legislators must recognize the critical impact of the damage caused by sleep disturbances due to high temperatures at night. Additionally, a daily minimum temperature of 25 °C should be the starting point when establishing a goal or guideline for nighttime temperature.

First Report of Spinal Anesthesia for Cesarean Delivery in a Parturient With Jeune Syndrome: A Case Report.

Jeune syndrome, also known as asphyxiating thoracic dystrophy, is a rare form of autosomal recessive skeletal dysplasia. Respiratory distress due to thoracic and lung dysplasia is the primary complication associated with this disorder in neonates. Women with Jeune syndrome seldom conceive and give birth, as only a few survive until adulthood. Herein, we report the world's first case of a cesarean delivery under spinal anesthesia in a pregnant woman with Jeune syndrome with a history of chest wall reconstruction and spinal fusion surgeries.

The protective effect of radicicol against renal ischemia--reperfusion injury in mice.

Overexpression of heat shock protein 70 kDa (HSP70) is known to confer cellular protection against ischemia-reperfusion (I/R) injury. Radicicol, a HSP90 inhibitor, has been reported to induce the expression of HSP70 protein. Here we studied whether radicicol attenuated renal I/R injury in vivo. Treatment of mice with radicicol ameliorated renal I/R injury and increased renal HSP70 mRNA and protein. Administration of radicicol with quercetin, an inhibitor of HSP70 induction, eliminated the renoprotective effect of radicicol. Our results suggest that the up-regulation of renal HSP70 protein by radicicol leads to a novel drug therapy against renal I/R injury.

Long-term intake of miso soup decreases nighttime blood pressure in subjects with high-normal blood pressure or stage I hypertension.

The present study aimed to investigate the effects of the combination of Marukome Nenrin miso, which has natriuretic effects, and Marukome MK-34-1 miso, which has potent angiotensin converting enzyme inhibitory effects, on blood pressure (BP) in humans. A total of 40 subjects aged 40-69 years with high-normal BP or stage I hypertension were randomly assigned to two groups: 1) the miso group (32 g 2:1 w/w Nenrin and MK-34-1 with 3.8 g salt/day) or 2) the control soy food group (14.4 g soy food with 0.2 g salt/day). The levels of major nutrients were equal in the miso and control food servings, except for the fiber and Na levels, which were higher in the miso food serving. Daytime and nighttime BP were measured with an automated BP monitor. Compared with the soy food intake, miso intake for 8 weeks did not affect daytime clinical BP but significantly decreased nighttime BP without affecting pulse rate (PR). Moreover, miso shifted the nighttime BP profile to lower levels than those at baseline. Soy food intake did not change the nighttime BP profile after 8 weeks. Miso intake also tended to reduce nighttime BP in a subgroup with stage 1 hypertension compared with the results of the soy food group participants and shifted the nighttime BP profile toward lower levels than those recorded at baseline. Miso intake did not influence lipid or glucose metabolism. In conclusion, this is the first report showing that miso reduces nighttime BP in humans. Miso may do so by shrinking the fluid spaces in the body and/or deactivating the adrenergic nervous system.

Publisher Correction: Stand age and species richness dampen interannual variation of ecosystem-level photosynthetic capacity.

In the version of this Article originally published, the wrong Supplementary Information pdf was uploaded, in which the figures did not correspond with those mentioned in the main text and the R code was not presented properly. This has now been replaced.

Robust colonoscope tracking method for colon deformations utilizing coarse-to-fine correspondence findings.

PURPOSE: Polyps found during CT colonography can be removed by colonoscopic polypectomy. A colonoscope navigation system that navigates a physician to polyp positions while performing the colonoscopic polypectomy is required. Colonoscope tracking methods are essential for implementing colonoscope navigation systems. Previous colonoscope tracking methods have failed when the colon deforms during colonoscope insertions. This paper proposes a colonoscope tracking method that is robust against colon deformations. METHOD: The proposed method generates a colon centerline from a CT volume and a curved line representing the colonoscope shape (colonoscope line) by using electromagnetic sensors. We find correspondences between points on a deformed colon centerline and colonoscope line by a landmark-based coarse correspondence finding and a length-based fine correspondence finding processes. Even if the coarse correspondence finding process fails to find some correspondences, which occurs with colon deformations, the fine correspondence finding process is able to find correct correspondences by using previously recorded line lengths. RESULT: Experimental results using a colon phantom showed that the proposed method finds the colonoscope tip position with tracking errors smaller than 50 mm in most trials. A physician who specializes in gastroenterology commented that tracking errors smaller than 50 mm are acceptable. This is because polyps are observable from the colonoscope camera when positions of the colonoscope tip and polyps are closer than 50 mm. CONCLUSIONS: We developed a colonoscope tracking method that is robust against deformations of the colon. Because the process was designed to consider colon deformations, the proposed method can track the colonoscope tip position even if the colon deforms.

Temporal changes in dissolved 137Cs concentrations in groundwater and stream water in Fukushima after the Fukushima Dai-ichi Nuclear Power Plant accident.

The concentration of dissolved 137Cs in groundwater and stream water in the headwater catchments in Yamakiya district, located ∼35 km north west of Fukushima Dai-ichi Nuclear Power Plant (FDNPP), was monitored from June 2011 to July 2013, after the earthquake and tsunami disaster. Groundwater and stream water were sampled at intervals of approximately 2 months at each site. Intensive sampling was also conducted during rainstorm events. Compared with previous data from the Chernobyl NPP accident, the concentration of dissolved 137Cs in stream water was low. In the Iboishi-yama catchment, a trend was observed for the concentration of dissolved 137Cs in stream water to decline, which could be divided into two phases by October 2011 (a fast flush of activity as a result of rapid washoff and a slow decline as a result of soil fixation and redistribution processes). The highest 137Cs concentration recorded at Iboishi-yama was 1.2 Bq/L on August 6, 2011, which then declined to 0.021-0.049 Bq/L during 2013 (in stream water under normal water-flow conditions). During the rainfall events, the concentration of dissolved 137Cs in stream water increased temporarily. The concentration of dissolved 137Cs in groundwater at a depth of 30 m at Iboishi-yama displayed a decreasing trend from 2011 to 2013, with a range from 0.039 Bq/L to 0.0025 Bq/L. The effective half-lives of stream water in the initial fast flush and secondary phases were 0.10-0.21 and 0.69-1.5 y, respectively in the three catchments. The effective half-life of groundwater was 0.46-0.58 y at Koutaishi-yama and 0.50-3.3 y at Iboishi-yama. The trend for the concentration of dissolved 137Cs to decline in groundwater and stream water was similar throughout 2012-2013, and the concentrations recorded in deeper groundwater were closer to those in stream water. The declining trend of dissolved 137Cs concentrations in stream water was similar to that of the loss of canopy 137Cs by throughfall, as shown in other reports of forest sites in the Yamakiya district.

Stand age and species richness dampen interannual variation of ecosystem-level photosynthetic capacity.

The total uptake of carbon dioxide by ecosystems via photosynthesis (gross primary productivity, GPP) is the largest flux in the global carbon cycle. A key ecosystem functional property determining GPP is the photosynthetic capacity at light saturation (GPPsat), and its interannual variability (IAV) is propagated to the net land-atmosphere exchange of CO2. Given the importance of understanding the IAV in CO2 fluxes for improving the predictability of the global carbon cycle, we have tested a range of alternative hypotheses to identify potential drivers of the magnitude of IAV in GPPsat in forest ecosystems. Our results show that while the IAV in GPPsat within sites is closely related to air temperature and soil water availability fluctuations, the magnitude of IAV in GPPsat is related to stand age and biodiversity (R2 = 0.55, P < 0.0001). We find that the IAV of GPPsat is greatly reduced in older and more diverse forests, and is higher in younger forests with few dominant species. Older and more diverse forests seem to dampen the effect of climate variability on the carbon cycle irrespective of forest type. Preserving old forests and their diversity would therefore be beneficial in reducing the effect of climate variability on Earth's forest ecosystems.

Stand age and species richness dampen interannual variation of ecosystem-level photosynthetic capacity.

The total uptake of carbon dioxide by ecosystems via photosynthesis (gross primary productivity, GPP) is the largest flux in the global carbon cycle. A key ecosystem functional property determining GPP is the photosynthetic capacity at light saturation (GPPsat), and its interannual variability (IAV) is propagated to the net land-atmosphere exchange of CO2. Given the importance of understanding the IAV in CO2 fluxes for improving the predictability of the global carbon cycle, we have tested a range of alternative hypotheses to identify potential drivers of the magnitude of IAV in GPPsat in forest ecosystems. Our results show that while the IAV in GPPsat within sites is closely related to air temperature and soil water availability fluctuations, the magnitude of IAV in GPPsat is related to stand age and biodiversity (R2 = 0.55, P < 0.0001). We find that the IAV of GPPsat is greatly reduced in older and more diverse forests, and is higher in younger forests with few dominant species. Older and more diverse forests seem to dampen the effect of climate variability on the carbon cycle irrespective of forest type. Preserving old forests and their diversity would therefore be beneficial in reducing the effect of climate variability on Earth's forest ecosystems.

Formation of size-controlled, denaturation-resistant lipid nanodiscs by an amphiphilic self-polymerizing peptide.

Nanodiscs are discoidal particles with a planar phospholipid bilayer enwrapped by proteins such as apolipoprotein A-I. Nanodiscs have been widely used for analyzing structures and functions of membrane proteins by dispersing them in solution. They are expected to be used as drug carriers and therapeutic agents. Amphiphilic peptides are known to form nanodiscs. However, the lipid-peptide nanodiscs are relatively unstable in solution, making them unsuitable for many applications. Here, we report the synthesis of an amphiphilic self-polymerizing peptide termed ASPP1, which polymerizes by intermolecular native chemical ligation reactions. ASPP1 spontaneously formed nanodiscs when added to phospholipid vesicles without using detergents. The diameter of the planar lipid bilayer in the nanodiscs was controlled by the lipid:peptide molar ratio. ASPP1-nanodiscs exhibited greater stability at high temperatures or in the presence of urea than nanodiscs formed by the non-polymerizing amphiphilic peptide or apolipoprotein A-I. Average and maximal degrees of ASPP1 polymerization were 2.4 and 12, respectively. Self-polymerization of the peptide appears to be responsible for stabilization of the nanodiscs. Our results open a new avenue for the development of nanodisc technology.

Cumulative history recorded in the depth distribution of radiocesium in sediments deposited on a sandbar.

We collected sediments deposited on a sandbar from the surface to 20 cm in depth in the Abukuma River to clarify the history of radiocesium derived from the Fukushima Daiichi Nuclear Power Plant (FDNPP) accident. We analyzed the (137)Cs concentration in the sediments from size-fractioned samples as well as bulk samples. The depth distribution of (137)Cs showed the highest concentration in the deepest sediment layer (18-20 cm) studied, which indicates that sediments with a lower (137)Cs concentration were transported and deposited on sediments having a higher (137)Cs concentration. At the same time, the depth distribution suggests a decrease in radioactivity in provenance areas of the sediments. Analysis of the size-fractioned sediments indicated that the three sediment layers at 4-6 cm, 16-18 cm and 18-20 cm intervals had similar size distribution of (137)Cs and grain size composition although the concentration levels of (137)Cs were different according to their bulk concentrations. The size distribution of (137)Cs also supported the possibility that the decrease in (137)Cs concentration in bulk sediments above 18 cm is due to a decrease in the level of radioactivity in the catchment area. A comparison of the size distribution of (137)Cs between the sediment layers above and below 18 cm suggested that the (137)Cs concentration in the transported fine sediment particles decreased more with time than the (137)Cs concentration in the coarse particles, reflecting the selective transport of the finer particles. The results of this study demonstrated that sediment layers deposited on a sandbar retained the cumulative history of the fluvial transport of radiocesium after the FDNPP accident.

Effects of carvedilol on plasma B-type natriuretic peptide concentration and symptoms in patients with heart failure and preserved ejection fraction.

Although the benefits of carvedilol in patients with heart failure and depressed ejection fraction (EF) have been elucidated, those in patients with preserved EF are not understood. We enrolled 40 patients with mild or moderate heart failure and EF >/=45%. They were randomly assigned to carvedilol (n = 19) or conventional therapy (n = 21). After 12 months of treatment, carvedilol significantly improved all end points (plasma concentration of B-type natriuretic peptide [BNP] from 175 (35 to 209) to 106 (52 to 160) pg/ml, mean (95% confidence interval) p <0.01; New York Heart Association functional class from 2.37 (2.13 to 2.61) to 1.56 (1.21 to 1.91), p <0.01; exercise capacity estimated with the Specific Activity Scale from 4.75 (4.50 to 5.00) to 5.68 (5.22 to 6.14) METs, p <0.02), whereas conventional therapy did not (plasma BNP concentration from 150 (114 to 186) to 174 (100 to 248) pg/ml; New York Heart Association functional class from 2.29 (2.08 to 2.50) to 2.11 (1.73 to 2.49); exercise capacity from 4.57 (4.34 to 4.80) to 4.72 (4.41 to 5.03) METs). Univariate regression analyses showed that only the use of carvedilol was correlated with the decrease in plasma BNP concentration (p <0.03). Multivariate analyses demonstrated that an ischemic cause of heart failure (p <0.02), high plasma concentration of BNP (p <0.02), left ventricular dilation (p <0.03), and use of carvedilol (p <0.04) at baseline were predictive of a decrease in plasma concentration of BNP. In conclusion, carvedilol potentially decreased neurohumoral activation, decreased symptoms, and increased exercise capacity in patients with heart failure and preserved EF.

Effects of percutaneous coronary arterial thrombectomy during acute myocardial infarction on left ventricular remodeling.

The benefit of primary angioplasty for acute myocardial infarction (AMI) is limited by the no-reflow phenomenon, resulting in chronic left ventricular (LV) remodeling. The aim of this study was to evaluate the impact of thrombectomy with the Rescue percutaneous thrombectomy catheter on LV function after AMI. We performed a retrospective study comparing conventional angioplasty with the combination of angioplasty and thrombectomy using the Rescue catheter. The study population was comprised of 109 consecutive patients with AMI who underwent angioplasty and thrombectomy and 86 controls treated with conventional angioplasty. Baseline clinical and lesion characteristics were similar in the 2 groups. Postprocedural restoration of normal flow (Thrombolysis In Myocardial Infarction grade 3) was more frequent in the thrombectomy group (82% vs 69%, p = 0.03). No differences were observed in cardiac events, including death, reinfarction, and target vessel revascularization (thrombectomy vs controls, 27% vs 33%; p = 0.44) or changes in ejection fraction (p = 0.22) during 6-month follow-up. The incidence of LV remodeling, defined as an increase in LV end-diastolic volume index of >20%, was significantly lower in the thrombectomy group (22% vs 44%; p = 0.01). Multiple logistic regression analysis revealed that thrombectomy with the Rescue catheter contributed significantly to reduction of both no-reflow and LV remodeling. In the setting of primary angioplasty, adjunctive pretreatment with a rescue catheter reduces the no-reflow phenomenon and protects against LV remodeling.

The use of Sunpatiens (Impatiens spp.) as a bioindicator of some simulated air pollutants--using an ornamental plant as bioindicator.

Sunpatiens were exposed separately or combined to ozone gas (130 ppb), fluoranthene (10 microM) and sulphuric acid mists (pH 3) sprayed as simulated pollutants in chamber conditions for 21 d. The treatments negatively affected the gas (CO(2) and moisture) exchange, leaf chlorophyll fluorescence, and the leaf-quality expressed in chlorophyll value (SPAD). Fluoranthene and the acid individual negative effects on the measured eco-physiological variables were nearly the same on Sunpatiens; their effects became aggravated on combining the duo. The foliar symptom assessments of chlorosis, necrosis and stippling revealed severe damages in ozone containing treatments compared with other treatments. The presence of fluoranthene exuberate ozone negative effects on some of the plant eco-physiological status. Where mannitol (1mM) additions were contained in treatments, mitigation effects of the negative impact of pollutants resulted. These findings indicated that Sunpatiens can be used as an active bioindicator of singular and multiple pollutants in field conditions.

Atmospheric concentrations of nitric acid, sulfur dioxide, particulate nitrate and particulate sulfate, and estimation of their dry deposition on the urban- and mountain-facing sides of Mt. Gokurakuji, Western Japan.

Atmospheric concentrations of nitric acid (HNO3), sulfur dioxide (SO2), particulate nitrate (NO3-) and particulate sulfate (SO4(2-)) on the urban- and mountain-facing sides of Mt. Gokurakuji were measured from November 2002 to October 2003, in order to evaluate the effects of anthropogenic activity on air quality and dry deposited nitrate and sulfate on the surfaces of pine foliage. The results showed that HNO3, SO2 and NO3(-) concentrations were significantly higher (P < 0.05) on the urban-facing side (1.54, 2.48 and 0.65 microg m(-3), respectively) than the mountain-facing side (0.67, 1.19 and 0.37 microg m(-3), respectively), while SO4(2-) concentrations did not differ significantly between the two sides (urban-facing: 2.80 microg m(-3); mountain-facing: 2.05 microg m(-3)). Indirect estimates of dry deposition rates of nitrate and sulfate to the surfaces of pine foliage based on the measured concentrations approximately agreed with the measured values determined by the foliar rinsing technique in a previous study. It was found that HNO3 was the major source (approximately 80%) of dry deposited nitrate on pine foliage, while the contribution from SO4(2-) was about equal to that from SO2. In conclusion, HNO3 and SO2 appear to be dominant species reflecting higher dry deposition rates of nitrate and sulfate on the urban-facing side compared to the mountain-facing side of Mt. Gokurakuji.