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There has been a great deal of research on cell division and its mechanisms; however, its processes have not yet been entirely elucidated. To find novel proteins that regulate cell division, we performed the screening using siRNAs and/or the expression plasmid of the target genes and identified leucine zipper protein 1 (LUZP1). Recent studies have shown that LUZP1 interacts with various proteins and stabilizes the actin cytoskeleton; however, the function of LUZP1 in mitosis is not known. In this study, we found that LUZP1 colocalized with the chromosomal passenger complex (CPC) at the centromere in metaphase and at the central spindle in anaphase and that these LUZP1 localizations were regulated by CPC activity and kinesin family member 20A (KIF20A). Mass spectrometry analysis identified that LUZP1 interacted with death-associated protein kinase 3 (DAPK3), one regulator of the cleavage furrow ingression in cytokinesis. In addition, we found that LUZP1 also interacted with myosin light chain 9 (MYL9), a substrate of DAPK3, and comprehensively inhibited MYL9 phosphorylation by DAPK3. In line with a known role for MYL9 in the actin-myosin contraction, LUZP1 suppression accelerated the constriction velocity at the division plane in our timelapse analysis. Our study indicates that LUZP1 is a novel regulator for cytokinesis that regulates the constriction velocity of the contractile ring.

Background Respiratory compromise (RC) including hypoxia and hypoventilation is likely to be missed in the postoperative period. Integrated pulmonary index (IPI) is a comprehensive respiratory parameter evaluating ventilation and oxygenation. It is calculated from four parameters: end-tidal carbon dioxide, respiratory rate, oxygen saturation measured by pulse oximetry (SpO2), and pulse rate. We hypothesized that IPI monitoring can help predict the occurrence of RC in patients at high-risk of hypoventilation in post-anesthesia care units (PACUs). Methods This prospective observational study was conducted in two centers and included older adults (≥ 75-year-old) or obese (body mass index ≥ 28) patients who were at high-risk of hypoventilation. Monitoring was started on admission to the PACU after elective surgery under general anesthesia. We investigated the onset of RC defined as respiratory events with prolonged stay in the PACU or transfer to the intensive care units; airway narrowing, hypoxemia, hypercapnia, wheezing, apnea, and any other events that were judged to require interventions. We evaluated the relationship between several initial parameters in the PACU and the occurrence of RC. Additionally, we analyzed the relationship between IPI fluctuation during PACU stay and the occurrences of RC using individual standard deviations of the IPI every five minutes (IPI-SDs). Results In total, 288 patients were included (199 elderly, 66 obese, and 23 elderly and obese). Among them, 18 patients (6.3 %) developed RC. The initial IPI and SpO2 values in the PACU in the RC group were significantly lower than those in the non-RC group (6.7 ± 2.5 vs. 9.0 ± 1.3, p p = 0.040, respectively). We used the area under the receiver operating characteristic curves (AUC) to evaluate their ability to predict RC. The AUCs of the IPI and SpO2 were 0.80 (0.69–0.91) and 0.64 (0.48–0.80), respectively. The IPI-SD, evaluating fluctuation, was significantly greater in the RC group than in the non-RC group (1.47 ± 0.74 vs. 0.93 ± 0.74, p = 0.002). Conclusions Our study showed that low value of the initial IPI and the fluctuating IPI after admission to the PACU predict the occurrence of RC. The IPI might be useful for respiratory monitoring in PACUs and ICUs after general anesthesia.

Targeted knock-in supported by the CRISPR/Cas systems enables the insertion, deletion, and substitution of genome sequences exactly as designed. Although this technology is considered to have wide range of applications in life sciences, one of its prerequisites for practical use is to improve the efficiency, precision, and specificity achieved. To improve the efficiency of targeted knock-in, there first needs to be a reporter system that permits simple and accurate monitoring of targeted knock-in events. In the present study, we created such a system using the PIGP gene, an autosomal gene essential for GPI-anchor biosynthesis, as a reporter gene. We first deleted a PIGP allele using Cas9 nucleases and then incorporated a truncating mutation into the other PIGP allele in two near-diploid human cell lines. The resulting cell clones were used to monitor the correction of the PIGP mutations by detecting GPI anchors distributed over the cell membrane via flow cytometry. We confirmed the utility of these reporter clones by performing targeted knock-in in these clones via a Cas9 nickase-based strategy known as tandem paired nicking, as well as a common process using Cas9 nucleases, and evaluating the efficiencies of the achieved targeted knock-in. We also leveraged these reporter clones to test a modified procedure for tandem paired nicking and demonstrated a slight increase in the efficiency of targeted knock-in by the new procedure. These data provide evidence for the utility of our PIGP-based assay system to quantify the efficiency of targeted knock-in and thereby help improve the technology of targeted knock-in.

Targeted knock-in assisted by the CRISPR/Cas9 system is an advanced technology with promising applications in various research fields including medical and agricultural sciences. However, improvements in the efficiency, precision, and specificity of targeted knock-in are prerequisites to facilitate the practical application of this technology. To improve the efficiency of targeted knock-in, it is necessary to have a molecular system that allows sensitive monitoring of targeted knock-in events with simple procedures.We developed an assay, named CD55 correction assay, with which to monitor CD55 gene correction accomplished by targeted knock-in. To create the reporter clones used in this assay, we initially introduced a 7.7-kb heterozygous deletion covering CD55 exons 2-5, and then incorporated a truncating mutation within exon 4 of the remaining CD55 allele in human cell lines. The resultant reporter clones that lost the CD55 protein on the cell membrane were next transfected with Cas9 constructs along with a donor plasmid carrying wild-type CD55 exon 4. The cells were subsequently stained with fluorescence-labeled CD55 antibody and analyzed by flow cytometry to detect CD55-positive cells. These procedures allow high-throughput, quantitative detection of targeted gene correction events occurring in an endogenous human gene.The current study demonstrated the utility of the CD55 correction assay to sensitively quantify the efficiency of targeted knock-in. When used with the PIGA correction assay, the CD55 correction assay will help accurately determine the efficiency of targeted knock-in, precluding possible experimental biases caused by cell line-specific and locus-specific factors.

Tandem paired nicking (TPN) is a method of genome editing that enables precise and relatively efficient targeted knock-in without appreciable restraint by p53-mediated DNA damage response. TPN is initiated by introducing two site-specific nicks on the same DNA strand using Cas9 nickases in such a way that the nicks encompass the knock-in site and are located within a homologous region between a donor DNA and the genome. This nicking design results in the creation of two nicks on the donor DNA and two in the genome, leading to relatively efficient homology-directed recombination between these DNA fragments. In this study, we sought to identify the optimal design of TPN experiments that would improve the efficiency of targeted knock-in, using multiple reporter systems based on exogenous and endogenous genes. We found that efficient targeted knock-in via TPN is supported by the use of 1700–2000-bp donor DNAs, exactly 20-nt-long spacers predicted to be efficient in on-target cleavage, and tandem-paired Cas9 nickases nicking at positions close to each other. These findings will help establish a methodology for efficient and precise targeted knock-in based on TPN, which could broaden the applicability of targeted knock-in to various fields of life science.

Background Intraoperative oxygen management is poorly understood. It was hypothesized that potentially preventable hyperoxemia and substantial oxygen exposure would be common during general anesthesia. Methods A multicenter, cross-sectional study was conducted to describe current ventilator management, particularly oxygen management, during general anesthesia in Japan. All adult patients (16 yr old or older) who received general anesthesia over 5 consecutive days in 2015 at 43 participating hospitals were identified. Ventilator settings and vital signs were collected 1 h after the induction of general anesthesia. We determined the prevalence of potentially preventable hyperoxemia (oxygen saturation measured by pulse oximetry of more than 98%, despite fractional inspired oxygen tension of more than 0.21) and the risk factors for potentially substantial oxygen exposure (fractional inspired oxygen tension of more than 0.5, despite oxygen saturation measured by pulse oximetry of more than 92%). Results A total of 1,786 patients were found eligible, and 1,498 completed the study. Fractional inspired oxygen tension was between 0.31 and 0.6 in 1,385 patients (92%), whereas it was less than or equal to 0.3 in very few patients (1%). Most patients (83%) were exposed to potentially preventable hyperoxemia, and 32% had potentially substantial oxygen exposure. In multivariable analysis, old age, emergency surgery, and one-lung ventilation were independently associated with increased potentially substantial oxygen exposure, whereas use of volume control ventilation and high positive end-expiratory pressure levels were associated with decreased potentially substantial oxygen exposure. One-lung ventilation was particularly a strong risk factor for potentially substantial oxygen exposure (adjusted odds ratio, 13.35; 95% CI, 7.24 to 24.60). Conclusions Potentially preventable hyperoxemia and substantial oxygen exposure are common during general anesthesia, especially during one-lung ventilation. Future research should explore the safety and feasibility of a more conservative approach for intraoperative oxygen therapy.

Background: Various biomarkers were used for sepsis; however, only a few become the standard. We reported that plasma histidine-rich glycoprotein (HRG) levels decreased in septic mice. Moreover, our clinical study on patients with systemic inflammatory response syndrome (SIRS) demonstrated that HRG levels in septic patients were lower than those in noninfective SIRS patients, and HRG could be a biomarker for sepsis within the SIRS population. In this study, we focused on septic patients and assessed the differences in HRG levels between the non-survivors and survivors. Methods: We studied intensive care unit (ICU) patients newly diagnosed with sepsis. Blood samples were collected within 24 h of ICU admission, and HRG levels were determined using an enzyme-linked immunosorbent assay. Findings: Ninety-nine septic patients from 11 institutes were included. HRG levels were significantly lower in non-survivors (n = 16) than in survivors (n = 83) (median, 15·1 [IQR, 12·7-16·6] vs. 30·6 [22·1-39·6] µg/ml; p < 0·01). Survival analysis revealed that the Harrell C-index (predictive power) for HRG was 0·90. Furthermore, when patients were divided into four subgroups according to quartiles of HRG level, the lowest HRG subgroup represented a significantly higher mortality. Interpretation: The initial HRG levels were found to be lower in non-survivors than in survivors. Decreased and classified HRG levels were also found to predict patient mortality. These results suggested that HRG could be a novel prognostic biomarker for sepsis. Trial Registration: UMIN Clinical Trials Registry (UMIN000017651). Funding Statement: This work was supported by grants from Japan Agency for Medical Research and Development (AMED) (AMED 15lk0201014h0003; AMED 18im0210109h0002), and from Secom Science and Technology Foundation to MN. Declaration of Interests: The authors declare no competing interests. Ethics Approval Statement: This was a multicenter, prospective, and observational investigation approved by the Institutional Review Board of the Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical Sciences, and of each institution.

This study considers adult mortality and analyzes the effect of public health investment on economic development, whereby investment increases savings but decreases fertility through a decrease in adult mortality. As the labor force increases, investment temporarily decreases capital per unit of labor. However, the decrease in fertility increases capital per unit of labor in subsequent periods. By considering these two opposing effects of decreasing fertility, we clarify the conditions required for investment to improve economic development via increasing savings and decreasing fertility. We examine panel estimation and present some weak evidence for our model.

We evaluated the anti-inflammatory effect of tyrosol (Tyr) on endotoxin-induced uveitis (EIU) in rats. EIU was induced in male Lewis rats by subcutaneous injection of lipopolysaccharide (LPS). Tyr (10, 50 or 100 mg/kg) was intravenously injected 2 hr before, simultaneously and 2 hr after LPS injection. The aqueous humor (AqH) was collected 24 hr after LPS injection; the infiltrating cell number, protein concentration, and tumor necrosis factor (TNF)-α, prostaglandin (PG)-E2 and nitric oxide (NO) levels were determined. Histopathologic examination and immunohistochemical studies for nuclear factor (NF)-κB, inhibitor of κB (IκB)-α, cyclooxygenase (COX)-2 and inducible NO synthase (iNOS) in the iris-ciliary body (ICB) were performed at 3 or 24 hr after LPS injection. To further clarify the anti-inflammatory effects, RAW264.7 macrophages were stimulated with LPS in the presence or absence of Tyr. Tyr reduced, in a dose-dependent manner, the infiltrating cell number, protein concentration, and TNF-α, PGE2 and NO levels in AqH and improved histopathologic scores of EIU. Tyr also inhibited LPS-induced COX-2 and iNOS expression, IκB-α degradation and nuclear translocation of activated NF-κB in ICB. Tyr significantly suppressed inflammatory mediator production in the culture medium and COX-2 and iNOS expression and activated NF-κB translocation in LPS-stimulated RAW264.7 cells. These results suggest that Tyr suppresses ocular inflammation of EIU by inhibiting NF-κB activation and subsequent proinflammatory mediator production.

Intraoperative oxygen management is poorly understood. It was hypothesized that potentially preventable hyperoxemia and substantial oxygen exposure would be common during general anesthesia.A multicenter, cross-sectional study was conducted to describe current ventilator management, particularly oxygen management, during general anesthesia in Japan. All adult patients (16 yr old or older) who received general anesthesia over 5 consecutive days in 2015 at 43 participating hospitals were identified. Ventilator settings and vital signs were collected 1 h after the induction of general anesthesia. We determined the prevalence of potentially preventable hyperoxemia (oxygen saturation measured by pulse oximetry of more than 98%, despite fractional inspired oxygen tension of more than 0.21) and the risk factors for potentially substantial oxygen exposure (fractional inspired oxygen tension of more than 0.5, despite oxygen saturation measured by pulse oximetry of more than 92%).A total of 1,786 patients were found eligible, and 1,498 completed the study. Fractional inspired oxygen tension was between 0.31 and 0.6 in 1,385 patients (92%), whereas it was less than or equal to 0.3 in very few patients (1%). Most patients (83%) were exposed to potentially preventable hyperoxemia, and 32% had potentially substantial oxygen exposure. In multivariable analysis, old age, emergency surgery, and one-lung ventilation were independently associated with increased potentially substantial oxygen exposure, whereas use of volume control ventilation and high positive end-expiratory pressure levels were associated with decreased potentially substantial oxygen exposure. One-lung ventilation was particularly a strong risk factor for potentially substantial oxygen exposure (adjusted odds ratio, 13.35; 95% CI, 7.24 to 24.60).Potentially preventable hyperoxemia and substantial oxygen exposure are common during general anesthesia, especially during one-lung ventilation. Future research should explore the safety and feasibility of a more conservative approach for intraoperative oxygen therapy.

Japan has been experiencing significant changes in its demographics. This study examines the effects of public care services and in doing so, aims to show that it is possible to improve overall welfare level by substituting family care with public care services, particularly when family care and public care services are interchangeable. In addition, we show that instead of reducing family care to zero, further expanding public care services can help achieve welfare optimization.

Tyrosol (Tyr) is a natural phenolic antioxidant with diverse biological activities. We compared the anti-inflammatory effects of intravenously administered Tyr versus prednisolone (PSL) in an endotoxin-induced uveitis (EIU) rat model. Intravenous administration of 100 mg/kg Tyr was performed 2 hr before, simultaneously and 2 hr after lipopolysaccharide (LPS) injection. Tyr treatment was associated with decreased inflammatory cell number, protein concentration, tumor necrosis factor (TNF)-α, PGE2 and NO levels in AqH and improvements in histopathologic evidence of EIU in ocular tissue at 24 hr after LPS injection. 100 mg/kg Tyr and 1 mg/kg PSL (administered on the same schedule as Tyr) had comparable anti-inflammatory effects. Taken together, Tyr may represent a promising therapeutic agent for the management of intraocular inflammatory diseases.

島根県立中央病院にて主に褥瘡がある患者をほぼ毎週水曜日に回診して得られた実態を報告する。褥瘡回診にて診察した患者の平均年齢は76.2歳で，男女比3：2，1ヵ月あたり平均53名で新規患者は平均20名であった。その予後は治癒42％（平均診察日数15.6日），死亡19％（同26.1日），転院20％（同33.7日），退院19％（同16.3日）であった。死亡例が多いのは救命救急科にての褥瘡発生報告が多いのが要因の一つと考えられた。回診を通した褥瘡の処置，指導，褥瘡患者のデータの集計により褥瘡対策への認識が高まり，褥瘡管理加算は約4倍の算定数となった。

Acute subdural hematoma (ASDH) is a critical condition following the onset of traumatic brain injury, and it is essential to immediately reduce elevated intracranial pressure (ICP). Single burr hole surgery/twist drill craniostomy is commonly performed in patients with ASDH as an emergency surgical intervention, usually preceding decompressive craniotomy. A novel method using a cerebrospinal fluid (CSF) drainage catheter kit for rapid drainage of ASDH is described. Percutaneous twist drill craniostomy using a CAMINO® micro ventricular bolt pressure-temperature monitoring kit was performed in the emergency room in 12 patients with severe ASDH. The kit contained a closed-system CSF drainage and pressure-temperature monitoring catheter, which allowed aspiration of the hematoma and monitoring of the ICP. The tip of the catheter was inserted into the hematoma from the forehead. The mean initial ICP was 61 mmHg, with a range of 31 to 120 mmHg. The liquid hematoma was aspirated, and the ICP was temporarily controlled to the normal range. Pupil dilation recovered immediately after aspiration of the hematoma in 3 patients. No complications occurred either during or after the operation. This new method for craniostomy is easy, safe, and effective to monitor and rapidly control ICP in the emergency room. This technique also offers the possibility of evaluating the patient's prognosis and determining indications for further decompressive craniectomy by the continuation of ICP control under ICP monitoring and evaluation of the reversibility of pupillary findings in ASDH patients.

Lipid peroxidation is caused by reactive oxygen species (ROS) and is involved in traumatic brain injury (TBI). Consequently, a therapeutic strategy for TBI may be to control lipid peroxidation. The only drug approved to date for blocking lipid peroxidation is edaravone (MCI-186), a novel free-radical scavenger shown to exert neuroprotective effects in acute ischemic stroke. Although edaravone scavenges hydroxyl and nitric oxide radicals, its effect on alkoxyl radicals (OR-), which also contribute to lipid peroxidation, is unknown. To date, the study of free radicals in blood has been severely hampered by technical difficulties in their detection. We used an in vitro and ex vivo electron spin resonance (ESR) method employing 5,5-dimethyl-1-pyrroline-N-oxide as a spin trap to investigate whether edaravone can scavenge OR-. By mixing either methemoglobin or human blood with tert-butyl hydroperoxide, we found that this technique can detect OR- generated in vitro. We also found that generated OR- can be completely absorbed by administration of edaravone in vitro (400 microM). Analysis of jugular venous blood collected from 17 TBI patients immediately before and 20 minutes after the administration of edaravone (30 mg, i.v.) revealed higher OR- levels in the untreated patients blood than in normal control blood samples. However, treatment with edaravone suppressed these OR- levels by 24.6% (radical intensity = 71.1 +/- 5.2-53.6 +/- 5.2; p < 0.01). Thus, edaravone can scavenge OR- and significantly reduce levels of these radicals in TBI patients. The novel ex vivo ESR method described here provides a valuable clinical measure of oxidative stress.

症例:57歳の女性。発熱,下痢,嘔吐を主訴に近医を受診し,補液などの治療を受けたが症状が改善しないため,紹介にて当院救急外来を受診した。来院時意識障害,発熱,項部硬直を認め,両下腿に紫斑を認めた。原因不明の感染症として入院し,第3病日両手指・足趾・足底に壊死が出現した。入院時の血液検体中からA群β溶連菌が検出されたことにより,toxic shock like syndromeの診断が強く疑われ,救急医学科へ転科となった。壊死部を切開しても,筋膜壊死や感染徴候を認めないことから,敗血症により,播種性血管内凝固(DIC)を生じ,symmetrical peripheral gangrene (SPG)となっていることが考えられた。敗血症に対し,ABPC 8g/dayとγ-グログリン投与,抗DIC療法(低分子ヘパリン,AT製剤,FFP)を行った。第6病日にはDICは改善し,第94病日に両手指(左第1指を除く)と両下腿の切断術を施行し,義足にてリハビリテーション後退院となった。考察:SPGとはDICによって生じた微小塞栓が微小循環を閉塞させるなどして末梢組織の壊死を起こすもので,敗血症によるものが多い。SPGとなる例は非常にまれであり,その発生には,基礎疾患(Raynaud's syndrome,動脈硬化,糖尿病など)の存在の他にスーパー抗原の関与が示唆されている。本症例では,ステロイド投与中であったこと以外には基礎疾患はなく,感染源も不明であった。

The treatment of carotid artery stenosis in elderly patients over 75 years old who present with impending stroke such as progressing or fluctuating stroke are still controversial. To confirm the efficacy of surgical revascularization, we analyzed the treatment outcome of surgical treatment and conservative medical treatment. Seven patients (mean age 78 y/o) were treated by carotid endoarterectomy (CEA: 3 cases) or carotid artery stenting (CAS: 4 cases), and 8 patients (mean age 84 y/o) were treated by anticoagulant or antiplatelet therapy. The outcomes of all surgically treated patients were favorable. But in medical group, 6 patients died and 1 was m-RS Grade 5. The median time between onset and complete stroke was 7 days. Four patients had a complete internal carotid artery occlusion in the medical group. Even though elderly patients presenting with hemodynamic impending stroke have a treatment high-risk, the emergency revascularization by CEA or CAS after careful evaluation of whole body seems to be a safe and effective strategy.

A 47-year-old woman, suffering from von Recklinghausen's disease, visited us because the tumor grew rapidly on her right neck. Except for numerous pigmented spots and cutaneous neurofibromas, there was a 18×10cm tumor on the area from right lateral neck to the right supraclavicular fossa, in addition to an indurated tumor measuring approximately 5cm in diameter on the right breast. According to the operative findings of an emergency operation for the lesion on the right neck, it was suspected of malignant tumor.Histopathologic diagnosis from department of clinical pathology was malignant peripheral nerve sheath tumor complicated with von Recklinghausen's disease. However, since the lesion had intralesional small nests and scattered large clear cells, it was suspected of undifferentiated cancer. Immunohistochemistry for myoglobin, desmin, NSE, S-100, EMA etc. could not show definite result. On the other hand, a biopsy specimen from her right breast was histopathologically diagnosed as breast cancer. Based upon all the above-mentioned histopathologic findings of both lesions of the right neck and the right breast, the lesion of the right neck was finally diagnosed as metastatic lesion of the breast cancer.

We recently showed that tactile stimulation (tickling) accompanied by positive emotion altered the expression of many genes in the rat hypothalamus (Hori et al., 2009 [15]). In this study, the effect of repeated tickling on gene expressions of the rat salivary gland was examined. After 4-week stimulation, several genes of the kallikrein (Klk) family were remarkably up-regulated and the alpha-amylase (amylase) gene was down-regulated in DNA microarray analysis. In quantitative analysis using real-time PCR of the submandibular gland of the rats tickled for 2 weeks, mRNAs of Klk1, Klk2 (Klk1c2, Tonin), Klk7 (Klk1l), Klk1b3 (Nerve growth factor, gamma), Klk1c10, Klks3 (Klk1c9) and GK11 were significantly 2-5-fold increased among 18 members of the Klk gene family examined and the submandibular amylase was decreased compared with the lightly touched and untouched control rats. In immunoblot analysis the increase in Klk7 protein was observed in the whole cell lysate fraction of the submandibular gland. In immunohistochemical analysis with anti-Klk7 polyclonal antibody, the immunostain was increased in duct cells of the submandibular gland of the tickled rat when compared with the lightly touched and untouched control rats. These results suggest that tactile sensory processing in the central nervous system affects the gene expression in the peripheral tissue probably via hormonal and/or autonomic neural activities. Submandibular Klks may be biochemical markers indicating positive emotional states.

We studied three cases of genital lichen sclerosus et atrophicus (LSA) using histological, immunohistochemical and ultrastructural techniques to elucidate the characteristics of the collagen fibres, the elastic fibres and the interfilamentous matrix in the upper dermal homogeneous zone. In the early stages of LSA, the homogeneous zone caused elaunin fibres to push downwards and partially disappear, the collagen fibres were presumably phagocytosed by fibroblasts, and there were peculiar cells containing numerous vacuoles in the cytoplasm. In the advanced stages of LSA, the homogeneous zones showed newly formed collagen fibres and amorphous, dispersed, medium electron-dense substances that were probably composed of elastin or elastin-like substances and microfilaments originating from the collagen fibres. Immunohistochemically, there were numerous anti-aortic alpha-elastin-positive substances, but there was no positivity for anti-amyloid P component. These findings suggest that in advanced LSA a hybrid substance (elastocollagenous mass) between collagen fibres and elastic fibres may be formed in the homogeneous zone.

Although solar keratosis (SK) is commonly classified histologically into five types i.e., hypertrophic, atrophic, acantholytic, bowenoid and pigmented types, we added another type, spongiotic clefting type, which has spongiotic suprabasal cleft. Squamous cell carcinoma (SCC) derived from SK has been reported to have such histologic types as de novo, acantholytic and bowenoid SCCKs and small celled carcinoma. Of 109 cases of SK studied, hypertrophic type was 56.9%, atrophic type 5.5%, acantholytic type 4.6%, bowenoid type 17.4% and spongiotic clefting type 15.6%, respectively. Of 17 cases of SCC from SK, de novo type was 23.5%, acantholytic type 47.0%, bowenoid type 5.9%, and small celled carcinoma 23.5%, respectively. Acantholytic type of SCC having the highest rate of occurrence, possibly, derives from acantholytic type of SK. Thus, it is supposed that tumor cells of acantholytic type of SK have higher tumor invading rate. Using PCNA and AgNORs as index of cell proliferating rate, we examined all types of SK. However we could not have any significant difference among them.

Acute subdural hematoma (ASDH) is a critical condition following the onset of traumatic brain injury, and it is essential to immediately reduce elevated intracranial pressure (ICP). Single burr hole surgery/twist drill craniostomy is commonly performed in patients with ASDH as an emergency surgical intervention, usually preceding decompressive craniotomy. A novel method using a cerebrospinal fluid (CSF) drainage catheter kit for rapid drainage of ASDH is described. Percutaneous twist drill craniostomy using a CAMINO(®) micro ventricular bolt pressure-temperature monitoring kit was performed in the emergency room in 12 patients with severe ASDH. The kit contained a closed-system CSF drainage and pressure-temperature monitoring catheter, which allowed aspiration of the hematoma and monitoring of the ICP. The tip of the catheter was inserted into the hematoma from the forehead. The mean initial ICP was 61 mmHg, with a range of 31 to 120 mmHg. The liquid hematoma was aspirated, and the ICP was temporarily controlled to the normal range. Pupil dilation recovered immediately after aspiration of the hematoma in 3 patients. No complications occurred either during or after the operation. This new method for craniostomy is easy, safe, and effective to monitor and rapidly control ICP in the emergency room. This technique also offers the possibility of evaluating the patient's prognosis and determining indications for further decompressive craniectomy by the continuation of ICP control under ICP monitoring and evaluation of the reversibility of pupillary findings in ASDH patients.

Orexins are neuropeptides that have a wide range of physiological effects, and recent studies have suggested that the orexin system may be involved in traumatic brain injury. However, the expression and localization of orexin receptors have not been examined yet under brain injury conditions. In the present study, we used immunohistochemical techniques to investigate the expression of orexin-1 receptor (OX1R) and its time-dependent changes in the mouse brain after controlled cortical impact (CCI) injury. OX1R-like immunoreactivity was first detected 6 h after injury in the surrounding penumbra of the injury. The intensity of this immunoreactivity was increased at 12 h, peaked at day 1, and then decreased from day 2 to day 7. To identify the cellular localization of OX1R, we also performed double-immunohistochemical staining with OX1R and several cell marker antibodies. OX1R-like immunopositive cells were clearly co-localized with immunoreactivity for the neuronal marker NeuN at day 7. It was also expressed on the periphery of cells immunopositive for CD11b, a microglial cell marker, at days 1 and 7. These results suggest that orexin and its receptor may play roles in traumatic brain injury, and that OX1R is induced in neurons and microglial cells after traumatic brain injury.

PACAP is a pleiotropic peptide and is well known to suppress neuronal cell death after ischemic injury through PACAP specific receptor (PAC1R). However, the pathological role of PAC1R is not elicited well on traumatic injured brain. Therefore, the purpose of this study is to investigate the expression and localization of PAC1R after traumatic brain injury (TBI) with immunohistochemistry. The PAC1R-immunoreactions (ir) were detected in peri-contusional area 3h after TBI and gradually increased up to 7d. Double immunohistochemical studies were revealed that the PAC1R-ir were co-localized with a microglial marker, CD11b. At 7d after TBI, the PAC1R-ir were merged with CD11b and with GFAP, an astroglial marker. These results suggested that PAC1R expresses in microglia and astrocyte with different time points after TBI, and PACAP and its receptor might play an important role for brain injury as well as ischemia.

Introducing a C(4)-like pathway into C(3) plants is one of the proposed strategies for the enhancement of photosynthetic productivity. For this purpose it is necessary to provide each component enzyme that exerts strong activity in the targeted C(3) plants. Here, a maize C(4)-form phosphoenolpyruvate carboxylase (PEPC, EC 4.1.1.31) was engineered for its regulatory and catalytic properties so as to be functional in the cells of C(3) plants. Firstly, amino acid residues Lys-835 and Arg-894 of maize PEPC, which correspond to Lys-773 and Arg-832 of Escherichia coli PEPC, respectively, were replaced by Gly, since they had been shown to be involved in the binding of allosteric inhibitors, malate or aspartate, by our X-ray crystallographic analysis of E. coli PEPC. The resulting mutant enzymes were active but their sensitivities to the inhibitors were greatly diminished. Secondly, a Ser residue (S780) characteristically conserved in all C(4)-form PEPC was replaced by Ala conserved in C(3)- and root-form PEPCs to decrease the half-maximal concentration (S(0.5)) of PEP. The double mutant enzyme (S780A/K835G) showed diminished sensitivity to malate and decreased S(0.5)(PEP) with equal maximal catalytic activity (V(m)) to the wild-type PEPC, which will be quite useful as a component of the C(4)-like pathway to be introduced into C(3) plants.

Lipid peroxidation is induced by reactive oxygen species and is involved in acute neuronal damage. Thus, controlling this process may be a realistic therapeutic strategy for treating or preventing neuronal damage. However, the study of free radicals in body fluids is currently severely hampered by technical difficulties in their detection. We have developed an ex vivo electron spin resonance (ESR) method, that employs 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) as a spin trap, to measure the alkoxyl radical (OR) in human blood. We found that this method can detect OR produced by treating human blood ex vivo. Analysis of jugular venous and radial arterial blood from patients with acute neuronal damage revealed higher OR levels in the venous blood compared to arterial samples. In brain death patients, however, the jugular vein OR spectrum was similar to the radial artery OR level. The ratio between radial artery and jugular vein levels, expressed as the RI (radical intensity) ratio = aORI/jORI, was calculated. A rterial-jugular bulb RI ratio below 1, together with accepted clinical criteria (unresponsive coma with brainstem areflexia), may provide a non-invasive assessment of cerebral circulatory arrest that can help to predict brain death. This novel ex vivo ESR method may be very useful for measuring oxidative stress in patients with acute neuronal damage.

Oxygen free radicals play important roles in brain injury, and one of the most important effects of brain hypothermia treatment is to suppress the generation of free radicals. In the present study, alkoxyl radical was determined utilizing ex vivo electron spin resonance (ESR) spectroscopy. Brain hypothermia was induced in traumatic brain injury (TBI) patients (n = 12) and global ischemic (GI) patients after cardiac arrest (n = 3) for 72h. Whole blood was sampled from the jugular vein and the radial artery at each stage of the treatment. In TBI patients, the radical intensity of the jugular vein (RIJV) was significantly higher than that of the control group and decreased gradually (P < 0.05: Student’s t-test). Conversely, the radical intensity of the radial artery (RIRA) gradually increased throughout the treatment. In GI patients, high RIJV and RIRA were observed in comparison with those of TBI patients. These data reflect the oxidative stresses of patients under brain hypothermia, and the ex vivo ESR method is useful for determination of the kinetics of oxygen free radical generation.

A rudimentary meningocele, a variant of primary cutaneous meningioma, was seen on the scalp of a 9-month-old Japanese boy. Clinically, the lesion on the left parietal area was round, about 1.6 cm in diameter, alopecic, and slightly elevated. Histologically, the lesion, located from the dermis to the subcutis, consisted of scattered foci of meningothelial cells, an anastomosing network of empty spaces with psammoma bodies and collagen bodies, and small vessels. Immunohistochemically, the meningothelial cells were positive for vimentin and desmin. Ultrastructurally, they had elongated cytoplasmic processes, intermediate filaments in the cytoplasm, and desmosomal junctions.