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Aim: Previous studies have highlighted the causation between chronic pain and falls in the elderly, but the mediator variables between chronic pain and falls in the frail elderly have not been identified. The purpose of this study was to identify the mediator variable of chronic pain and falls., Methods: This study is a prospective cohort study. Participants were 116 frail elderly individuals living in a community. Chronic pain (pain lasting more than 3 months) was assessed using a questionnaire, and the occurrence of falls was tracked using a falls calendar. Using logistic regression, a model was created with falls as the dependent variable and chronic pain and confounders as independent variables. The mediation analysis was conducted with chronic pain as the independent variable, falls as the dependent variable, and factors that showed significant differences between the chronic pain group and the non-chronic pain group as candidate mediators., Results: Even after adjusting for covariates, chronic pain significantly influenced the occurrence of falls (odds ratio: 3.004, 95% CI [1.226, 7.363], p=0.016). The results of mediation analysis showed a significant direct effect between chronic pain and falls, and a significant indirect effect (partial mediation) of sleep disorders on the relationship between chronic pain and falls., Conclusions: Chronic pain in frail elderly mediates sleep disorders and influences falls. It is suggested that interventions for chronic pain and sleep disorders should focus on preventing falls in the frail elderly., (Copyright © 2021 Elsevier B.V. All rights reserved.)

We herein report a case of Carney complex (CNC) complicated with primary pigmented nodular adrenocortical disease (PPNAD) after unilateral adrenalectomy. A 44-year-old woman was admitted to our hospital for PPNAD surgery. She had previously undergone surgery for cardiac myxoma and had a PRKAR1A mutation with no family history of CNC. She had Cushing's signs, but her metabolic abnormalities were mild. Adrenal insufficiency due to poor medication adherence was a concern, so she underwent unilateral adrenalectomy. Cushing's signs improved postoperatively and without recurrence for five years. Treatment plans for PPNAD should be determined based on the patient's condition, medication adherence, and wishes.

Purpose: Telesimulation, whereby learners telecommunicate and use simulator resources to receive education at an off-site location, has been used to educate surgical trainees about how to perform basic surgical procedures. However, it has not yet been used for advanced surgical procedures. We aimed to develop a telesimulation curriculum to teach a common laparoscopic procedure called transabdominal preperitoneal (TAPP) repair and to explore the feasibility of its use., Methods: Learning objectives were created to develop a telesimulation curriculum that included didactic telelecture and telesimulation training. Pre-tests and post-tests to assess the didactic materials were developed and assessed among surgeons with various experiences. We assessed the feasibility of the telelecture and telesimulation separately. Pre-tests, post-tests, and questionnaires were used to assess the telelectures. We created a TAPP repair telesimulation system and checked for problems during training. Trainees were assessed to determine their skill improvement using previously published assessment tools and questionnaires., Results: A didactic telelecture was developed based on the learning objectives using an expert consensus and pilot-tested among five participants. After the lecture, their test scores improved and they expressed positive opinions about the usefulness of telelectures. The TAPP repair telesimulation training was pilot-tested among three trainees. No technical problems occurred during training. All trainees improved their skills after the telesimulation training and agreed that the training was useful for learning the TAPP repair procedure., Conclusions: We systematically developed a telesimulation curriculum for the TAPP repair procedure and demonstrated its feasibility among learners., Competing Interests: Conflict of interestOn behalf of all authors, the corresponding author states that there is no conflict of interest., (© The Author(s), under exclusive licence to Association for Surgical Education 2022, Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.)

Background: Endosurgery provides several advantages over open surgery in the context of global surgery; however, there are several barriers to its introduction. The preliminary assessment of needs and barriers is essential for carrying out effective support as Global Surgery. However, no report has described the initiation of support for endosurgery based on a preliminary survey of the needs and barriers. The present survey study aimed at determining the needs and barriers of pediatric endosurgery in Nepal. Materials and Methods: A needs assessment survey was conducted among all pediatric surgeons in Nepal via an online platform. This was followed by workshop on pediatric endosurgical skills in Nepal. To assess the skills of participants and effectiveness of the workshop, the skill evaluation tests and the questionnaire survey were conducted. Results: Fourteen pediatric surgeons (response rate: 60.9%) responded to the needs assessment survey. More than 70% of the participants did not have any experience with advanced endosurgical procedures. However, advanced endosurgical procedures were strongly needed. A lack of training was indicated as a major barrier for the introduction of pediatric endosurgery. Fifteen participants completed the workshop. Participants' confidence in their endosurgical skills improved significantly after the workshop. The skill evaluation tests revealed that participants' endosurgical skills also improved significantly after the workshop, although even after the workshop, participants still took an average of 415.6 seconds to place and knot one suture. Conclusions: The needs assessment survey and workshop for Nepalese pediatric surgeons helped clarify their needs for endosurgery and the barriers to its introduction.

Tumor necrosis frequently occurs in malignant tumors, showing rapid growth and invasion. This phenomenon is generally regarded as simple ischemic necrosis due to insufficient tumor vessels and blood supply. However, the necrotic tissue contains high amount of nuclear substances, DNA, and nucleoproteins that may affect the surrounding tumor cells by promoting or suppressing the tumor cell growth in vivo . This study focused on the effects of an externally administered water-soluble nuclear crude extract (SNE) containing nuclear protein and oligonucleotides on several human cancer and noncancer cell lines. The results demonstrated that the SNE suppressed cell growth in cancer and noncancer cells in vitro . Through the flow cytometry analysis of the nuclear DNA content, it was observed that the SNE increased and decreased cell proportion in the S and G2/M phases, respectively, thereby suggesting that the cell growth inhibition was due to cell cycle delay, and not due to apoptosis. These studies suggest that the high-concentration of extracellular nucleotides generated as a result of tumor necrosis and/or released from infiltrated neutrophils could suppress the growth of surrounding cancer and intrinsic cells, which provides us some insights into an alternative anticancer strategy for patients with highly malignant necrotic tumor., Competing Interests: The authors declare no conflict of interest., (© 2021 The Author(s).)

Women are more resistant than men to the development of vascular diseases. However, menopause is a factor leading to deterioration of female vascular integrity, and it is reported that the risk of vascular diseases such as atherosclerosis and abdominal aortic aneurysm is increased in postmenopausal women. Although it is suggested that perivascular adipose tissue (PVAT) is deeply involved in the increased risk of vascular disease development, the effect of menopause on PVAT integrity is unknown. In this study, we aimed to elucidate the effect of menopause on PVAT in ovariectomized (OVX) rats. PVAT was divided into 4 regions based on characteristics. Hypertrophy and increased inflammation of adipocytes in the PVAT were observed in the OVX group, but the effects of OVX were different for each region. OVX induced matrix metalloproteinase (MMP) -9 which degrade extracellular matrix such as elastin and collagen fibers in PVAT. Degeneration of the arterial fibers of the thoracic and abdominal aorta were observed in the OVX group. These results indicate that OVX can cause dysfunction of PVAT which can cause degradation of arterial fibers. Appropriate management of PVAT may play an important role in the prevention and treatment of diseases originating from ovarian hypofunction.

Many organisms reductively assimilate selenite to synthesize selenoprotein. Although the thioredoxin system, consisting of thioredoxin 1 (TrxA) and thioredoxin reductase with NADPH, can reduce selenite and is considered to facilitate selenite assimilation, the detailed mechanism remains obscure. Here, we show that selenite was reduced by the thioredoxin system from Pseudomonas stutzeri only in the presence of the TrxA (PsTrxA), and this system was specific to selenite among the oxyanions examined. Mutational analysis revealed that Cys33 and Cys36 residues in PsTrxA are important for selenite reduction. Free thiol-labeling assays suggested that Cys33 is more reactive than Cys36. Mass spectrometry analysis suggested that PsTrxA reduces selenite via PsTrxA-SeO intermediate formation. Furthermore, an in vivo formate dehydrogenase activity assay in Escherichia coli with a gene disruption suggested that TrxA is important for selenoprotein biosynthesis. The introduction of PsTrxA complemented the effects of TrxA disruption in E. coli cells, only when PsTrxA contained Cys33 and Cys36. Based on these results, we proposed the early steps of the link between selenite and selenoprotein biosynthesis via the formation of TrxA-selenium complexes.

Dietary protamine can ameliorate hyperlipidemia; however, the protamine-derived active peptide and its hypolipidemic mechanism of action are unclear. Here, we report the discovery of a novel anti-obesity and hypocholesterolemic peptide, RPR (Arg-Pro-Arg), derived from protamine in mice fed a high-fat diet for 50 days. Serum cholesterol levels were significantly lower in the protamine and RPR groups than in the control group. White adipose tissue weight was significantly decreased in the protamine and RPR groups. The fecal excretion of cholesterol and bile acid was significantly higher in the protamine and RPR groups than in the control group. We also observed a significant decrease in the expression of hepatic SCD1, SREBP1, and adipocyte FAS mRNA, and significantly increased expression of hepatic PPARα and adipocyte PPARγ1 mRNA in the protamine group. These findings demonstrate that the anti-obesity effects of protamine are linked to the upregulation of adipocyte PPARγ1 and hepatic PPARα and the downregulation of hepatic SCD1 via SREBP1 and adipocyte FAS. RPR derived from protamine has a crucial role in the anti-obesity action of protamine by evaluating the effective dose of adipose tissue weight loss.

Cation-dependent mannose 6-phosphate receptor (CD-MPR) and cation-independent MPR (CI-MPR) belong to the P-type lectin family. Both intracellular and cell surface MPRs can recognize and bind with the terminal mannose 6-phospahte (M6P) residues of N-glycans attached to the mammalian lysosomal enzymes and the related co-factors. Domain9 (Dom9), which is one of the extracytoplasmic region of the CI-MPR, has relatively higher affinity for M6P residues. Here we describe the production of recombinant Dom9-His protein by Pichia pastris, purification, and application as a probe for lectin blotting.

A biaxial tensile testing method has been used to get macroscopic information on elastoplastic deformation of a thin steel specimen and improve the accuracy of plastic processing of steel materials. We newly developed a biaxial tensile testing machine for pulsed neutron experiments (BTM-NEU) to provide the microscopic crystallographic information of steel materials under biaxial load and correlate it with the macroscopic mechanical properties of the materials. The performance of the BTM-NEU was experimentally evaluated with cold-rolled mild steel and hot-rolled high-tensile-strength steel materials and compared with that of a standard biaxial tensile testing machine (BTM-std) as follows. •The BTM-NEU can test an ISO-standardized cruciform specimen as the BTM-std and its performance is equivalent to that of the BTM-std.•The BTM-NEU has excellent long-time reliability and stability necessary for pulsed neutron experiments, especially Bragg-edge neutron imaging experiments.•The BTM-NEU can be applied to pulsed neutron experiments using a Bragg-edge imaging method., (© 2019 The Author(s).)

An electron was found to catalyze the coupling of magnesium diarylamides with aryl iodides giving triarylamines through a radical-anion intermediate. The transformation requires no transition metal catalysts or additives, and a wide array of products are formed in good-to-excellent yields., (© 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

If invasive ventilation can be avoided by performing noninvasive mechanical ventilation (NIV) in patients with acute respiratory failure (ARF), the disease can be effectively managed. It is important to clarify the characteristics of patients with neuromuscular diseases in whom initial NIV is likely to be unsuccessful. We studied 27 patients in stable neuromuscular condition who initially received NIV to manage fatal ARF to identify differences in factors immediately before the onset of ARF among patients who receive continuous NIV support, patients who are switched from NIV to invasive ventilation, and patients in whom NIV is discontinued. Endpoints were evaluated 24 and 72 hours after the initiation of NIV. After 24 hours, all but 1 patient with amyotrophic lateral sclerosis (ALS) received continuous NIV support. 72 hours later, 5 patients were switched from NIV to invasive ventilation, and 5 patients continued to receive NIV support. 72 hours after the initiation of NIV, the proportion of patients with a diagnosis of ALS differed significantly among the three groups (P=0.039). NIV may be attempted to manage acute fatal respiratory failure associated with neuromuscular diseases, but clinicians should carefully manage the clinical course in patients with ALS.

We used high-performance liquid chromatography to separate urine obtained from whole-body gamma-irradiated mice (4 Gy) before analyzing each fraction with matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry to identify radiation-responsive molecules. We identified two candidates: hepcidin antimicrobial peptide 2 (hepcidin-2) and peptide fragments of kidney androgen-regulated protein (KAP). We observed that peak increases of hepcidin-2 in urine were delayed in a dose-dependent manner (1 Gy and above); however, the amount of KAP peptide fragments showed no correlation with radiation dose. In addition, an increase in hepcidin-2 after exposure to relatively low radiation doses (0.25 and 0.5 Gy, respectively) was biphasic (at 8-48 h and 120-168 h, respectively, after irradiation). The increase in hepcidin-2 paralleled an increase in hepcidin-2 gene (Hamp2) mRNA levels in the liver. These results suggest that radiation exposure directly or indirectly induces urinary excretion of hepcidin-2 at least in part by the upregulation of Hamp2 mRNA in the liver., (© The Author 2016. Published by Oxford University Press on behalf of The Japan Radiation Research Society and Japanese Society for Radiation Oncology.)

Amla is one of the most important plants in Indian traditional medicine and has been shown to improve various age-related disorders while decreasing oxidative stress. Mitochondrial dysfunction is a proposed cause of aging through elevated oxidative stress. In this study, we investigated the effects of Amla on mitochondrial function in C2C12 myotubes, a murine skeletal muscle cell model with abundant mitochondria. Based on cell flux analysis, treatment with an extract of Amla fruit enhanced mitochondrial spare respiratory capacity, which enables cells to overcome various stresses. To further explore the mechanisms underlying these effects on mitochondrial function, we analyzed mitochondrial biogenesis and antioxidant systems, both proposed regulators of mitochondrial spare respiratory capacity. We found that Amla treatment stimulated both systems accompanied by AMPK and Nrf2 activation. Furthermore, we found that Amla treatment exhibited cytoprotective effects and lowered reactive oxygen species (ROS) levels in cells subjected to t-BHP-induced oxidative stress. These effects were accompanied by increased oxygen consumption, suggesting that Amla protected cells against oxidative stress by using enhanced spare respiratory capacity to produce more energy. Thus we identified protective effects of Amla, involving activation of mitochondrial function, which potentially explain its various effects on age-related disorders.

Glutathione is a valuable tripeptide widely used in the pharmaceutical, food, and cosmetic industries. In industrial fermentation, glutathione is currently produced primarily using the yeast Saccharomyces cerevisiae. Intracellular glutathione exists in two forms; the majority is present as reduced glutathione (GSH) and a small amount is present as oxidized glutathione (GSSG). However, GSSG is more stable than GSH and is a more attractive form for the storage of glutathione extracted from yeast cells after fermentation. In this study, intracellular GSSG content was improved by engineering thiol oxidization metabolism in yeast. An engineered strain producing high amounts of glutathione from over-expression of glutathione synthases and lacking glutathione reductase was used as a platform strain. Additional over-expression of thiol oxidase (1.8.3.2) genes ERV1 or ERO1 increased the GSSG content by 2.9-fold and 2.0-fold, respectively, compared with the platform strain, without decreasing cell growth. However, over-expression of thiol oxidase gene ERV2 showed almost no effect on the GSSG content. Interestingly, ERO1 over-expression did not decrease the GSH content, raising the total glutathione content of the cell, but ERV1 over-expression decreased the GSH content, balancing the increase in the GSSG content. Furthermore, the increase in the GSSG content due to ERO1 over-expression was enhanced by additional over-expression of the gene encoding Pdi1, whose reduced form activates Ero1 in the endoplasmic reticulum. These results indicate that engineering the thiol redox metabolism of S. cerevisiae improves GSSG and is critical to increasing the total productivity and stability of glutathione.

Parkinson's disease (PD) is an obstinate progressive neurodegenerative disease and characterized by locomotor impairment and dopaminergic neuronal degeneration in the substantia nigra pars compacta (SNc). We examined in here the dietary effect of nucleoprotein (NP) extracted from salmon soft roe on 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-injected PD-like mice model to prevent the symptom as an alternative medicine. Male C57/BL6 mice were given either an artificially modified NP-free diet (NF) or NF supplied with 1.2% NP for 1 week. Then, mice were injected intraperitoneally four times with 20 mg/kg MPTP. Seven days later, locomotor activity was examined, and the brains were immunostained with tyrosine hydroxylase (TH) and Iba1 antibodies. Moreover, in situ detection of superoxide anion (O2(-)) and gene expression of mitochondrial electron transfer chain gene, Cox8b was evaluated in midbrains. NP-fed animals showed significantly reduced locomotor impairment and an increased number of TH-positive cells in the SNc compared with NF animals. The NP-fed animals also showed reduced lower levels of O2(-) and up-regulation of Cox8b levels and Iba1 immunoreactivity, suggesting that inflammation and oxidative stress were suppressed and mitochondrial impairment was relieved in these animals. Supplementation of the diet with NP may serve as a useful preventive measure to slow the onset of PD.

The ability of an activated carbon (AC) to adsorb 18 different cytokines with molecular weights ranging from 8 kDa to 70 kDa and high mobility group box-1 (HMGB1) from inflammatory model plasma at 310 K and the mechanisms of adsorption were examined. Porosity analysis using N2 gas adsorption at 77K showed that the AC had micropores with diameters of 1-2 nm and mesopores with diameters of 5-20 nm. All 18 cytokines and HMGB1 were adsorbed on the AC; however, the shapes of the adsorption isotherms changed depending on the molecular weight. The adsorption isotherms for molecules of 8-10 kDa, 10-20 kDa, 20-30 kDa, and higher molecular weights were classified as H-2, L-3, S-3, and S-1 types, respectively. These results suggested that the adsorption mechanism for the cytokines and HMGB1 in the mesopores and on the surface of the AC differed as a function of the molecular weight. On the basis of these results, it can be concluded that AC should be efficient for cytokine adsorption., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Olfactory systems sense and respond to various odorants. Olfactory receptors, which in most organisms are G protein (heterotrimeric guanine nucleotide-binding protein)-coupled receptors, directly bind volatile or soluble odorants. Compared to the genomes of mammals, the genome of the nematode Caenorhabditis elegans contains more putative olfactory receptor genes, suggesting that in nematodes there may be combinatorial complexity to the receptor-odor relationship. We used RNA interference (RNAi) screening to identify nematode olfactory receptors necessary for the response to specific odorants. This screening identified 194 candidate olfactory receptor genes linked to 11 odorants. Additionally, we identified SRI-14 as being involved in sensing high concentrations of diacetyl. Rescue and neuron-specific RNAi experiments demonstrated that SRI-14 functioned in ASH neurons, specific chemosensory neurons, resulting in avoidance responses. Calcium imaging revealed that ASH neurons responded to high diacetyl concentrations only, whereas another class of chemosensory neurons, AWA neurons, reacted to both low and high concentrations. Loss of SRI-14 function hampered ASH responses to high diacetyl concentrations, whereas loss of ODR-10 function reduced AWA responses to low odorant concentrations. Chemosensory neurons ectopically expressing SRI-14 responded to a high concentration of diacetyl. Thus, nematodes have concentration-dependent odor-sensing mechanisms that are segregated at the olfactory receptor and sensory neuron levels.

Glutathione is a valuable tripeptide that is widely used in the pharmaceutical, food, and cosmetic industries. Intracellular glutathione exists in two forms, reduced glutathione (GSH) and oxidized glutathione (GSSG). Most of the glutathione produced by fermentation using yeast is in the GSH form because intracellular GSH concentration is higher than GSSG concentration. However, the stability of GSSG is higher than GSH, which makes GSSG more advantageous for industrial production and storage after extraction. In this study, an oxidized glutathione fermentation method using Saccharomyces cerevisiae was developed by following three metabolic engineering steps. First, over-expression of the glutathione peroxidase 3 (GPX3) gene increased the GSSG content better than over-expression of other identified peroxidase (GPX1 or GPX2) genes. Second, the increase in GSSG brought about by GPX3 over-expression was enhanced by the over-expression of the GSH1/GSH2 genes because of an increase in the total glutathione (GSH + GSSG) content. Finally, after deleting the glutathione reductase (GLR1) gene, the resulting GPX3/GSH1/GSH2 over-expressing ΔGLR1 strain yielded 7.3-fold more GSSG compared with the parental strain without a decrease in cell growth. Furthermore, use of this strain also resulted in an enhancement of up to 1.6-fold of the total glutathione content compared with the GSH1/GSH2 over-expressing strain. These results indicate that the increase in the oxidized glutathione content helps to improve the stability and total productivity of glutathione.

A 14-year-old girl weighing 32 kg was diagnosed with suprasellar tumor causing hydrocephalus, hypothyroidism, adrenal dysfunction and central diabetes insipidus. She was treated with levothyroxine and hydrocortisone and urged to take fluid to replace urine. She was scheduled to undergo ventricular drainage to relieve hydrocephalus prior to tumor resection. For the first surgery, desmopressin was not started and urine output reached 4,000 to 6,000 ml x day(-1), urine osmolality 64 mOsm x l(-1) and urine specific gravity 1.002. Anesthesia was induced with sevoflurane and maintained with propofol and remifentanil. Maintenance fluid was with acetated Ringer's solution and urine loss was replaced with 5% dextrose. Bradycardia and hypotension occurred after intubation, which was treated with volume load. Infusion volume was 750 ml and urine output was 1100 ml during 133 min of anesthesia. Postoperative day 1 nasal desmopressin was started. Ten days later, partial tumor resection was performed. Anesthesia was induced with propofol and fentanyl and maintained with sevoflurane and remifentanil. Infusion volume was 610 ml, urine output 380 ml, and blood loss 151 ml during 344 min of anesthesia. Hemodynamic parameters were stable throughout the procedure. Pathology of the tumor was revealed to be germinoma. Bradycardia and hypotension experienced during the first surgery was suspected to be caused by preoperative hypovolemia brought by polyuria. Desmopressin was proved to be effective to treat excessive urine output and to maintain good perioperative water balance.

A 70-year-old woman, who had hepatitis-C related liver cirrhosis died suddenly. Autopsy showed a massive retroperitoneal hematoma and ruptured splenic vein, as well as densely bloody ascites. This suggested that chronic and unnoticed retroperitoneal leak from the ruptured vein preceded the acute and fatal outcome of the intra-abdominal bleeding. Spontaneous rupture of the splenic vein is rarely reported in liver cirrhosis despite the presence of portal hypertension. This rare association is discussed with a literature review.

Invasion into the matrix is one of hallmarks of malignant diseases and is the first step for tumor metastasis. Thus, analysis of the molecular mechanisms of invasion is essential to overcome tumor cell invasion. In the present study, we screened for colon carcinoma-specific genes using a cDNA microarray database of colon carcinoma tissues and normal colon tissues, and we found that fermitin family member-1 (FERMT1) is overexpressed in colon carcinoma cells. FRRMT1, FERMT2 and FERMT3 expression was investigated in colon carcinoma cells. Reverse transcription polymerase chain reaction (RT-PCR) analysis revealed that only FERMT1 had cancer cell-specific expression. Protein expression of FERMT1 was confirmed by western blotting and immunohistochemical staining. To address the molecular functions of FERMT genes in colon carcinoma cells, we established FERMT1-, FERMT2- and FERMT3-overexpressing colon carcinoma cells. FERMT1-overexpressing cells exhibited greater invasive ability than did FERMT2- and FERMT3-overexpressing cells. On the other hand, FERMT1-, FERMT2- and FERMT3-overexpressing cells exhibited enhancement of cell growth. Taken together, the results of this study indicate that FERMT1 is expressed specifically in colon carcinoma cells, and has roles in matrix invasion and cell growth. These findings indicate that FERMT1 is a potential molecular target for cancer therapy.

Microglial cells account for approximately 12-15 % of the cells in the central nervous system (CNS). Microglial cells are polarized by pathological stimuli such as cytokines, chemokines, and growth factors, and play important roles in the deterioration and repair of the CNS. Here, we established cultures of primary microglial cells isolated from the brains of adult C57/BL6 mice using Percoll density gradients. The cells were cultured and stained with antibodies against CD11b, glial fibrillary acidic protein, myelin basic protein and NeuN to determine microglial, astroglial, oligodendroglial, and neuronal cells respectively. Moreover, the cells were exposed to interferon-γ (IFNγ) plus interleukin-1β (IL-1β) or IL-4 for 24 h to demonstrate the activating phenotypes with inducible nitric oxide synthase (iNOS), Ym1, and Iba-1 immunoblotting. At least 95 % of the cultured cells were CD11b-positive and -negative for astroglial, neuronal, and oligodendrocyte markers. IFNγ plus IL-1β treatment resulted in classical activation, which was represented by an increase in iNOS. The cells also displayed alternative activation, which increased Ym1 when treated with IL-4. The present study indicates that the microglial cells isolated as described here are a useful tool for elucidating adult microglial function.

Traumatic brain injury (TBI) is a major cause of preventable death and serious morbidity, with subsequent low cerebral blood flow (CBF) considered to be associated with poor prognosis. In the present study, we demonstrated the effect of the free radical scavenger edaravone on regional CBF (rCBF) after TBI. Male mice (C57/BL6) were subjected to TBI using a controlled cortical impactor device. Immediately after TBI, the animals were intravenously administered 3.0 mg/kg of edaravone or a vehicle saline solution. Two-dimensional rCBF images were acquired before and 24 h post-TBI, and were quantified in the ipsilateral and contralateral hemispheres (n = 5 animals per group). CBF in the vehicle-treated animals decreased broadly over the ipsilateral hemisphere, with the region of low rCBF spreading from the frontal cortex to the occipital lobe. The zone of lowest rCBF matched that of the contusion area. The mean rCBF at 24 h for a defined elliptical region between the bregma and lambda was 73.7 ± 5.8 %. In comparison, the reduction of rCBF in edaravone-treated animals was significantly attenuated (93.4 ± 5.7 %, p < 0.05). The edaravone-treated animals also exhibited higher rCBF in the contralateral hemisphere compared with that seen in -vehicle-treated animals. It is suggested that edaravone reduces neuronal damage by scavenging reactive oxygen species (ROS) and by maintaining intact the autoregulation of the cerebral vasculature.

Traumatic brain injury (TBI) is a major cause of death and disability in young people. No effective therapy is available to ameliorate its damaging effects. Our aim was to investigate the optimal therapeutic time window of edaravone, a free radical scavenger which is currently used in Japan. We also determined the temporal profile of reactive oxygen species (ROS) production, oxidative stress, and neuronal death. Male C57Bl/6 mice were subjected to a controlled cortical impact (CCI). Edaravone (3.0 mg/kg), or vehicle, was administered intravenously at 0, 3, or 6 hours following CCI. The production of superoxide radicals (O2 (∙-)) as a marker of ROS, of nitrotyrosine (NT) as an indicator of oxidative stress, and neuronal death were measured for 24 hours following CCI. Superoxide radical production was clearly evident 3 hours after CCI, with oxidative stress and neuronal cell death becoming apparent after 6 hours. Edaravone administration after CCI resulted in a significant reduction in the injury volume and oxidative stress, particularly at the 3-hour time point. Moreover, the greatest decrease in O2 (∙-) levels was observed when edaravone was administered 3 hours following CCI. These findings suggest that edaravone could prove clinically useful to ameliorate the devastating effects of TBI.

A novel extracellular glutathione fermentation method using engineered Saccharomyces cerevisiae was developed by following three steps. First, a platform host strain lacking the glutathione degradation protein and glutathione uptake protein was constructed. This strain improved the extracellular glutathione productivity by up to 3.2-fold compared to the parental strain. Second, the ATP-dependent permease Adp1 was identified as a novel glutathione export ABC protein (Gxa1) in S. cerevisiae based on the homology of the protein sequence with that of the known human glutathione export ABC protein (ABCG2). Overexpression of this GXA1 gene improved the extracellular glutathione production by up to 2.3-fold compared to the platform host strain. Finally, combinatorial overexpression of the GXA1 gene and the genes involved in glutathione synthesis in the platform host strain increased the extracellular glutathione production by up to 17.1-fold compared to the parental strain. Overall, the metabolic engineering of the glutathione synthesis, degradation, and transport increased the total (extracellular + intracellular) glutathione production. The extracellular glutathione fermentation method developed in this study has the potential to overcome the limitations of the present intracellular glutathione fermentation process in yeast.

Glutathione (GSH) is a valuable tri-peptide that is widely used in the pharmaceutical, food, and cosmetic industries. Glutathione is produced industrially by fermentation using Saccharomyces cerevisiae. In this study, we demonstrated that engineering in sulfate assimilation metabolism can significantly improve GSH production. The intracellular GSH content of MET14 and MET16 over-expressing strains increased up to 1.2 and 1.4-fold higher than that of the parental strain, respectively, whereas those of APA1 and MET3 over-expressing strains decreased. Especially, in the MET16 over-expressing strain, the volumetric GSH concentration was up to 1.7-fold higher than that of the parental strain as a result of the synergetic effect of the increases in the cell concentration and the intracellular GSH content. Additionally, combinatorial mutant strains that had been engineered to contain both the sulfur and the GSH synthetic metabolism synergistically increased the GSH production. External addition of cysteine to S. cerevisiae is well known as a way to increase the intracellular GSH content; however, it results a decrease in cell growth. This study showed that the engineering of sulfur metabolism in S. cerevisiae proves more valuable than addition of cysteine as a way to boost GSH production due to the increases in both the intracellular GSH content and the cell growth.

Glutathione is a valuable tripeptide that is widely used in the pharmaceutical, food, and cosmetic industries. Glutathione is industrially produced by fermentation using Saccharomyces cerevisiae. Before the glutathione fermentation process with S. cerevisiae, a glucose extraction process from starchy materials is required. This glucose extraction is usually carried out by converting starchy materials to starch using high-temperature cooking and subsequent hydrolysis by amylases to convert starch to glucose. In this study, to develop an energy-saving glutathione production process by reducing energy consumption during the cooking step, we efficiently produced glutathione from low-temperature cooked rice using amylase-expressing S. cerevisiae. The combination of the amylase-expressing yeast with low-temperature cooking is potentially applicable to a variety of energy-saving bio-production methods of chemicals from starchy bio-resources., (Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Glutathione is a valuable tri-peptide that is widely used in the pharmaceutical, food, and cosmetic industries. Glutathione is produced industrially by fermentation using Saccharomyces cerevisiae, and supplementation of fermentation with several amino acids can increase intracellular GSH content. More recently, however, focus has been given to protein as a resource for biofuel and fine chemical production. We demonstrate that expression of a protease on the cell surface of S. cerevisiae enables the direct use of keratin and soy protein as a source of amino acids and that these substrates enhanced intracellular GSH content. Furthermore, fermentation using soy protein also enhanced cell concentration. GSH fermentation from keratin and to a greater extent from soy protein using protease-displaying yeast yielded greater GSH productivity compared to GSH fermentation with amino acid supplementation. This protease-displaying yeast is potentially applicable to a variety of processes for the bio-production of value-added chemicals from proteinaceous biomass resources.

A unique spiro-type Brønsted acid-Lewis base organocatalyst has been developed. The new bifunctional organocatalyst promoted aza-Morita-Baylis-Hillman reactions to yield adducts with up to 98% ee.

We developed a novel enzymatic glutathione (GSH) production system using Saccharomyces cerevisiae as a whole-cell biocatalyst, and improved its GSH productivity by metabolic engineering. We demonstrated that the metabolic engineering of GSH pathway and ATP regeneration can significantly improve GSH productivity by up to 1.7-fold higher compared with the parental strain, respectively. Furthermore, the combination of both improvements in GSH pathway and ATP regeneration is more effective (2.6-fold) than either improvement individually for GSH enzymatic production using yeast. The improved whole-cell biocatalyst indicates its great potential for applications to other kinds of ATP-dependent bioproduction.

Human leukocyte antigen (HLA) typing is essential to carry out HLA-class I restricted antigenic peptide-based cancer immunotherapy. To establish a one-step polymerase chain reaction-sequence-specific primer (PCR-SSP) method, we designed two novel HLA-A24-specific primer sets and determined the optimal conditions for specific amplification. Then, we performed HLA-A24 typing of two healthy donors' and 17 cancer patients' peripheral blood with serological typing and PCR-SSP typing. Eleven of the 19 cases were determined HLA-A24-positive by the PCR-SSP method precisely; however, five cases showed false positive with serological analysis. Thus, for HLA-A24 typing in the Japanese population, the PCR-SSP method is faster and more accurate than serological typing.

The simplified international diagnostic criteria for autoimmune hepatitis (AIH), re-revised by the International AIH Group in 2008, were investigated in 114 patients with AIH from 15 centers in Japan. While applying of the criteria, we had to pay attention to anti-nuclear antibody measurement methods, and liver histology scoring. Definite and probable AIH were diagnosed in 83 and 22 patients, respectively. The criteria were found to be useful for the diagnosis of AIH in Japan. However, 9 patients who did not meet the diagnostic criteria showed normal immunoglobulin G levels or were negative for autoantibodies. As the criteria were unreliable for diagnosing such atypical cases in the present series, we speculated that we should not rely solely on these, criteria and take a more holistic approach to diagnosis in such cases.

A 54-year-old woman suffered acute hepatitis after she acquired cystitis. Laboratory results on admission showed: AST 925, ALT 1171, ALP 623, gamma-GTP127 IU/l, T-Bil 5.0 mg/dl, antinuclear antibodies negative, smooth muscle antibodies 80, antimitochondrial antibodies (AMA) 80, antimitochondrial M2 antibody (AMA-M2) 117 index, IgG 2210 mg/dl. She also had HLA-DR4 and HLA-DR8. Histological study of a liver biopsy specimen suggested that she had autoimmune hepatitis rather than primary biliary cirrhosis. When prednisolone was administered, her liver function immediately improved and AMA and AMA-M2 levels fell to 20 and 52 respectively. However when cystitis recurred 4 months later, her liver function worsened. Laboratory findings showed AST 174, ALT 183 IU/l. Upon increasing the dosage of prednisolone, her liver function improved again. After the recurrence of hepatitis, AMA and AMA-M2 levels increased to 320 and 149 respectively. We speculate that the urinary tract infection triggered an autoimmune response and her genetic predisposition also played a crucial role in the process.

A glutamine synthetase I family protein, Lengsin, was previously identified as a novel lens-specific transcript in the vertebrate eye. In this report, we show for the first time that Lengsin is a novel tumor-associated antigen expressed ectopically in lung cancer. Interestingly, a novel spliced form of human Lengsin termed 'splicing variant 4', gaining exon 3 that codes extra 63 amino acids, is the dominant transcript form in lung cancer cells. Lengsin mRNA could be detected in 7 of 12 (58%) lung cancer cell lines and 7 of 7 (100%) surgically resected lung cancer tissues. On the other hand, Lengsin transcripts could not be detected in normal major tissues or in other cancer cell lines, including melanoma, colorectal carcinoma, breast carcinoma and hepatocellular carcinoma. In addition, knockdown of Lengsin mRNA with RNAi caused cell death and a decrease of cell viability, suggesting that Lengsin has some essential role in cell survival. Since the lens is an immune-privileged site, we regard Lengsin as a highly immunogenic cancer antigen. Anti-Lengsin autoantibodies were detectable in sera of lung cancer patients, although these patients did not show any lens-related disturbances. Hence, Lengsin splicing variant 4 might be an immunogenic lung cancer-specific antigen that is suitable as a diagnostic marker and for molecular targeting therapy, including immunotherapy.

Identification of tumor-associated antigens may facilitate vaccination strategies to treat patients with malignant diseases. We have found that the centrosomal protein, Cep55/c10orf3 acts as a novel breast carcinoma-associated tumor-associated antigen. Cep55/c10orf3 mRNA was detectable in a wide variety of tumor cell lines. Expression was barely detectable in normal tissues except for testis and thymus. Moreover, Cep55/c10orf3 protein could be detected by a monoclonal anti-Cep55/c10orf3 antibody (# 11-55) in 69.8% of breast carcinoma, 25% of colorectal carcinoma, and 57.8% of lung carcinoma tissues. The expression of Cep55/c10orf3 protein did not show any relationship with the hormone receptors such as estrogen receptor and progesterone receptor or expression patterns of p185 HER2/neu. We designed 11 peptides which displayed a human leukocyte antigen-A24 binding motif. One Cep55/c10orf3-peptide, Cep55/c10orf3&#95;193(10) (VYVKGLLAKI), induced cytotoxic T lymphocytes (CTLs) in 3 of 3 patients with Cep55/c10orf3 (# 11-55)-positive breast carcinoma. A Cep55/c10orf3&#95;193(10)-specific CTL clone could also recognize Cep55/c10orf3 (+) displayed on human leukocyte antigen-A24 (+) cancer cell lines. These data indicate that Cep55/c10orf3 peptides were naturally presented by breast cancer cells and can cause CTL clonal expansion in vivo. Monoclonal antibody # 11-55 and the Cep55/c10orf3&#95;193(10) peptides may be useful as part of a therapeutic strategy for hormonal therapy or anti-p185 HER2/neu monoclonal antibody therapy-resistant breast carcinoma patients.

To achieve good care for stroke patients in the Hokumou Secondary Medical District, we reaffirmed the current status of the services, discovered the keypoints and issues, and searched for solutions to the current problems. The population, its age composition and the number of beds occupied by stroke patients--stratified in acute, convalescent and maintenance stages-were surveyed. The number of patients suffering from stroke under the jurisdiction of this district was compiled, by using the "regional alliance path for stroke," and the severity and outcome of each case were evaluated by using the mRS (modified Rankin Scale). We held seminars for citizens' on the prevention and management of stroke and the attendees were asked to fill out questionnaires, and were organized: networks for medical care and nursing in the Okhotsk Sea area and people's voices were heard at the network meetings; and at the town meetings held at Kitami, Abashiri and Bihoro, the challenges and problems in each area were discussed. The findings obtained from these activities were compiled for the current study. The rate of release of the patients to their home environment was high in the acute stage but low in the convalescent stage. To substantiate the regional management of stroke patients, it is essential that one be familiar with the availability of the limited medical recourses for and dynamics of patients. And yet, there is a limit to the efforts that the medical profession alone can make for the acute and convalescent stages. It is necessary that the health services assist in solving the problems, with the cooperation of the citizens and the nursing network that is involved in homes and nursing facilities for the care of patients in the maintenance stage.

The nuclear mitotic apparatus (NuMA) protein is a component of the nuclear matrix in interphase cells and an essential protein for the formation of mitotic spindle poles. We used herpes simplex virus (HSV), an enveloped DNA virus that replicates in the nucleus, to study the intra-nuclear dynamics of NuMA in infected cells. This study shows that NuMA is extensively modified following HSV infection, including phosphorylation of an unidentified site(s), and that it depends to an extent on viral DNA synthesis. Although NuMA is insoluble in uninfected interphase cells, HSV infection induced solubilisation and dynamic relocalisation of NuMA, whereupon the protein became excluded from viral replication compartments -- sites of virus transcription and replication. Live cell, confocal imaging showed that NuMA localisation dramatically changed from the early stages (diffusely nuclear, excluding nucleoli) to late stages of infection (central diminuition, but remaining near the inner nuclear peripheries). In addition, NuMA knockdown using siRNA suggested that NuMA is important for efficient viral growth. In summary, we suggest that NuMA is required for efficient HSV infection, and identify further areas of research that address how the virus challenges host cell barriers.

The protein encoded by the UL14 gene of herpes simplex virus type 1 (HSV-1) and HSV-2 is expressed late in infection and is a minor component of the virion tegument. An UL14-deficient HSV-1 mutant (UL14D) forms small plaques and exhibits an extended growth cycle at low multiplicities of infection (MOI) compared to wild-type virus. Although UL14 is likely to be involved in the process of viral maturation and egress, its precise role in viral replication is still enigmatic. In this study, we found that immediate-early viral mRNA expression was decreased in UL14D-infected cells. Transient coexpression of UL14 and VP16 in the absence of infection stimulated the nuclear accumulation of both proteins. We intended to visualize the fate of VP16 released from the infected virion and constructed UL14-null (14D-VP16G) and rescued (14R-VP16G) viruses that expressed a VP16-green fluorescent protein (GFP) fusion protein. Synchronous high-multiplicity infection of the viruses was performed at 4 degrees C in the absence of de novo protein synthesis. We found that the presence of UL14 in the virion had an enhancing effect on the nuclear accumulation of VP16-GFP. The lack of UL14 did not significantly alter virus internalization but affected incoming capsid transport to the nuclear pore. These observations suggested that UL14 (i) enhanced VP16 nuclear localization at the immediately early phase, thus indirectly regulating the expression of immediate-early genes, and (ii) was associated with efficient nuclear targeting of capsids. The tegument protein UL14 could be part of the machinery that regulates HSV-1 replication.

Posterior-anterior body weight shift during stance phase of human overground locomotion was investigated by recording sole-floor reaction force from five anatomically discrete points with strain gauge transducers of 14 mm diameter attached firmly to the sole of bare foot. At first the subject was asked to walk straight on the laboratory floor at his/her preferred velocity. Then the subject was asked to walk curved path of about 1m radius. For kicking off the body at the end of stance phase, sole-floor reaction force from 3rd metatarsal was stronger than 1st metatarsal or 5th metatarsal during the straight walking, thus body weight shift is represented from heel to 3rd metatarsal line. When walking along a curved path, two types of strategies were recognized; a group of subjects walked leaning to inner leading foot during stance period as judged by stronger forces recorded from 5th metatarsal combined with stronger force from 1st metatarsal of outer trailing foot. Another group of subjects showed almost the same patterns either in the straight and curved walking, suggesting the subjects changed direction of the foot during the immediately previous swing phase to the tangent direction of the curve and placed the foot without leaning the body weight to either direction. Hemiplegic patients showed strikingly different distribution of sole-floor reaction forces from the five points; strongest forces were recorded from 3rd and 5th metatarsals combined with reduced reaction force from heel, therefore characteristic y-vector patterns were observed.

The first patient is a 60-year-old man who underwent total gastrectomy and splenectomy for type-3 gastric cancer after neoadjuvant chemotherapy. The second patient is a 69-year-old woman who underwent distal gastrectomy for type-2 gastric cancer and pyloric stenosis after neoadjuvant chemotherapy. Some peritoneal dissemination was observed in these two cases. No re-growth of peritoneal dissemination was seen for three years or more from treatment with the oral anticancer drug TS-1. Treatment on an outpatient basis, therefore, greatly contributed to their quality of life. We consider TS-1 as a first-line anti-cancer drug for advanced gastric cancer.

Right and left balance during human locomotion has been estimated by the distance between locations of the sequential right and left heel-strikes, or step width. During the stance phase of human locomotion one leg maintains medial-lateral balance for the progression. We focused our attention on this point, and medial-lateral balance during straight-ahead and circular walking was investigated by recording sole-floor reaction force from five anatomically discrete points of human sole; calcaneus, 1st, 3rd, 5th metatarsals and great toe. Forces from these points were recorded during straight walking and circular walking. Medial-lateral balance was obtained by subtracting force at 5th metatarsal from force at 1st metatarsal (x-axis vector). The foot takes off the floor from medial balance in most steps, although in some steps the foot takes off from lateral balance at slower walking speed at 2 km/h or 4 km/h, showing variable patterns of x-axis vector. At faster walking speed at 6 km/h or 8 km/h body weight shifted to 1st metatarsal before taking off the floor. During circular walking body weight shift to 1st metatarsal in the outer foot, and to 5th metatarsal in the inner foot in most cases.

In order to clarify differences of treadmill from overground locomotion, experiments were carried out on 10 volunteers (five males and five females). Sole-floor reaction force was recorded from five anatomically discrete points with strain gauge transducers of 14 mm diameter attached firmly to the sole of bare-foot. At first the subject was asked to walk on the laboratory floor at his/her preferred velocity. After the average velocity was obtained, the subject was asked to walk on the treadmill at the same velocity of average overground walking. Stance period at treadmill walking shortened to 93.3% (P < 0.01) of the value at overground walking. Coefficient of variation of stance period was significantly smaller at the treadmill walking than at overground walking. Strain gauge-floor contact times were shorter in the treadmill walking; heel 81.2%, first metatarsal 93.5%, third metatarsal 93.6%, fifth metatarsal 90.6% and at great toe 93.2% of overground locomotion. Cadence during treadmill locomotion was significantly larger than overground walking (106.6%, P < 0.05). These results show that when subjects walk on the treadmill and on laboratory floor at the identical speed, stance period shortened by 6.7% while cadence increased by 6.6% on the treadmill.

Role of a central fixation target on the latencies of visually guided manual movement was analyzed on young healthy subjects, age-matched control subjects and patients with Parkinson's disease (Hoehn and Yahr stages II, III, and IV). Two paradigms were used: overlap paradigm where a central fixation target was lighted throughout the test, and gap paradigm where a central fixation target was turned off 200 ms before a peripheral target was lighted. The subject was first asked to fixate the central target then instructed to locate a peripheral target with a laser beam spot, operated with wrist flexion or extension as quickly as possible. Latencies of gap paradigm are always shorter than those of overlap task in all the groups. Latencies of both overlap and gap tasks prolonged from young to elder, from elder to PD II, from PD II to PD III and from PD III to PD IV. Also latencies were extremely prolonged in the overlap tasks and correlated with disease severity. Latencies in the gap tasks were less prolonged as compared with those in the overlap tasks. The visual fixation target prolonged the visuo-motor latency in association with severity of Parkinson's disease.

Sole-floor reaction forces were recorded from five anatomically discrete points to analyze characteristics of human locomotion. Strain gauge of 14 mm diameter were firmly attached to the sole of bare-foot for recording force changes from the following five points: (1) medial process of calcaneus, (2) head of 1st metatarsal, (3) head of 3rd metatarsal, (4) head of 5th metatarsal and (5) great toe. Fifteen healthy adults were asked to walk at 2, 4, 6 and 8 km/h and to run at 8 km/h on the treadmill. Sole-floor reaction forces from 1st to 5th metatarsals show reciprocal changes during stance phase, while force from 1st metatarsal is strong 5th metatarsal shows weak reaction and vice versa. This phenomenon may be an expression of locomotor program to maintain vertical stability of the body during stance phase. There was a linear relation between walking speeds and sum of force from the five points, although sum of forces from three metatarsals did not change significantly during the walking speeds, indicating mainly calcaneus and great toe contribute to increasing walking speed. During running the sum of force from the three metatarsals increased sharply, joining the other two points to increase thrust.

A 26-year-old female with intractable epileptic seizures was studied with I-123 iomazenil cerebral benzodiazepine receptor, I-123 IMP inter-ictal and Tc-99m ECD ictal cerebral blood flow SPECT. The ictal cerebral blood flow SPECT indicated the location of the seizures to be in the left temporal lobe, where increased regional cerebral blood flow was noted in marked contrast to the inter-ictal SPECT. Ictal electroencephalograms (EEGs) recorded with scalp and sphenoidal electrodes also suggested the left temporal lobe as the location of the seizures. On I-123 iomazenil SPECT, however, decreased benzodiazepine receptor density was demonstrated in the right temporal lobe. MRI showed mild atrophy and abnormal signal intensity in the right temporal lobe. Ictal EEGs recorded with intracranial electrodes revealed that abnormal electrical activity of the brain always emerged from the right temporal lobe and then propagated to the contralateral side. Based on the findings of intracranial EEGs, partial resection of the right anterior temporal lobe including hippocampus was performed. After the surgery, no seizure occurred. Pathological examination of the surgical specimens revealed hippocampal sclerosis. This case suggested that cerebral benzodiazepine receptor imaging with I-123 iomazenil can be helpful for correct localization of epileptogenic foci.

Strain gauge transducers were firmly attached to five points of the human sole: calcaneus, 1st, 3rd, and 5th metatarsals and great toe. Forces from these five points were recorded during treadmill walking at different speeds. With this method it is possible to obtain data of several dozen steps successively. Lateral-medial force change (x-vector) during progression was obtained from the 5th and 1st metatarsals and posterior-anterior force change (y-vector) was obtained from the calcaneus and 3rd metatarsal. Lateral balance and medial balance were differentiated in x-vector and rearfoot phase and forefoot phase were distinguished in y-vector. The percentage of the forefoot phase among the stance period shows a linear increase with speed of progression. It was concluded that the phase of body sway forward is regulated by walking speeds.

The insecticidal activity of dinotefuran and 23 related compounds against the housefly, Musca domestica (L) was measured by injection with metabolic inhibitors. Dinotefuran was less active than imidacloprid and clothianidin by a factor of 10 in molar concentrations. Their binding activities to the fly-head membrane preparation were measured by using [125I]alpha-bungarotoxin ([125I]alpha-BGTX) and [3H]imidacloprid ([3H]IMI) as radioligands. The activity of some selected compounds measured with [3H]IMI was 10(4)-fold higher than that measured with [125I]alpha-BGTX. With [3H]IMI as a radioligand, dinotefuran was 13-fold less active than imidacloprid. The inhibitory effect of dinotefuran on the binding of [3H]IMI to the membrane preparation was in a competitive manner. Quantitative analysis of the insecticidal activity of the test compounds with the binding activity measured with [3H]IMI showed that the higher the binding activity, the higher was the insecticidal activity.