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The syndrome of pregnancy- and lactation-associated osteoporosis is a rare disorder whose precise etiology and treatment are largely unknown. We herein report two such cases occurring in the early postpartum period that led to multiple fragility compression fractures. Combination therapy of vitamin D and vitamin K enabled a marked gradual increase in bone mineral density. [ABSTRACT FROM AUTHOR]

The present study measured changes in plasma pentosidine and bone turnover markers in elderly patients with osteoporosis treated using bisphosphonate. The relationship between pentosidine and bone turnover markers and bone mineral density (BMD) was investigated. This study consisted of post-menopausal osteoporotic women who could be treated using bisphosphonate for 3 years were included in the present analysis. The study population consisted of 58 cases, all women, ranging in age from 53 to 86 years (mean, 67.1 years). Bisphosphonate treatment significantly increased BMD of the lumbar spine to 0.914 ± 0.141 g/cm2 and BMD of the femoral neck to 0.708 ± 0.086 g/cm2 after 3 years ( p < 0.001 versus baseline). The mean BAP level was 27.3 ± 8.3 U/L in patients at baseline. After bisphosphonate treatment, BAP significantly decreased to 18.1 ± 7.2 U/L at 3 years ( p < 0.001). Urinary NTX also decreased after bisphosphonate treatment. After 3 years of treatment, urinary NTX significantly decreased from 50.0 ± 19.0 nmol BCE/mmol Cr to 24.6 ± 10.2 nmol BCE/mmol Cr at 3 years ( p < 0.001). Serum pentosidine levels were 0.0413 ± 0.0094 μg/mL at baseline and 0.0413 ± 0.0122 μg/mL after 3 years. They were not significantly changed by bisphosphonate treatment. Serum pentosidine levels were not changed by treatment with bisphosphonates. Thus, serum pentosidine may not be suitable as a marker of bone quality after 3 years of bisphosphonate treatment. [ABSTRACT FROM AUTHOR]

Bisphosphonates are effective in increasing bone mineral density (BMD), but fragility fractures can still occur despite bisphosphonate treatment. The purpose of this study was to determine if long-term bisphosphonate users have characteristic findings in the musculoskeletal system, which could put them at risk of developing typical or atypical femoral fractures. We recruited 40 female patients who had taken bisphosphonates for more than 3 years. The control group included 60 volunteers who were matched by age, body mass index, and dual-energy X-ray absorptiometry-derived BMDs. We measured the skeletal muscle cross sectional area around the proximal thigh and buckling ratio of the femoral neck using quantitative computed tomography (qCT) and several biochemical markers of bone metabolism. Those parameters were compared between the groups. While no significant differences of buckling ratio derived from qCT were detected, the skeletal muscle cross sectional area was significantly smaller in the long-term bisphosphonate users than in the controls. Furthermore, the serum pentosidine level was significantly higher in the bisphosphonate users than in the controls. To determine if those differences were attributable to bisphosphonate treatment, we further compared those parameters between before and after 3 years of bisphosphonate treatment in 32 patients. After 3 years of bisphosphonate treatment, the BMD of the femoral neck and serum pentosidine level increased but not the skeletal muscle cross sectional area. In the present study, the skeletal muscle mass did not match the bone mass in long-term bisphosphonate users, thus suggesting that increases in BMD by bisphosphonates are unlikely to have secondary positive effects on the surrounding skeletal muscles. Also, serum pentosidine levels were greater in the long-term bisphosphonate users. Further study is necessary to test if such patients are prone to develop typical or atypical femoral fractures. [ABSTRACT FROM AUTHOR]

Aim Human induced pluripotent stem (hi PS) cells are an alternative cell source of regenerative medicine for liver disease. Because variations in hepatic differentiation efficacy among hi PS cells exist, it is important to select a hi PS cell line with hepatic differentiation propensity. In addition, nuclear receptors ( NR) regulate essential biological processes including differentiation and development. In this study, we identified the hi PS cell line with hepatic differentiation propensity and examined expression levels of 48 NR during this process. Methods We screened 28 hi PS cell lines, which are established from various tissues of healthy persons with various reprogramming methods, using a three-step differentiation method, and examined expression levels of 48 NR by quantitative real-time polymerase chain reaction during the differentiation process in the selected cells. Results hi PS- RIKEN- 2B and hi PS- RIKEN- 2F cells have hepatic differentiation propensity. Differentiation propensity towards endoderm was affected by donor origin but not by reprogramming methods or cell type of origins. Expression levels of NR were closely associated with those of hepatic differentiation markers. Furthermore, expression patterns of NR were categorized as five patterns. In particular, seven NR such as chicken ovalbumin upstream promoter transcription factor 1, retinoic acid receptor α, peroxisome proliferator-activated receptor-γ, progesterone receptor, photoreceptor cell-specific nuclear receptor, tailless homolog orphan receptor and glucocorticoid receptor were identified as the genes of which expression gradually goes up with differentiation. Conclusion These findings will be useful for not only elucidating mechanisms of hepatic differentiation of hi PS cells but also cell-based therapy for liver diseases. [ABSTRACT FROM AUTHOR]

Congenital sideroblastic anemia (CSA) is a hereditary disorder characterized by microcytic anemia and bone marrow sideroblasts. The most common form of CSA is attributed to mutations in the X-linked gene 5-aminolevulinic acid synthase 2 ( ALAS2 ). ALAS2 is a mitochondrial enzyme, which utilizes glycine and succinyl-CoA to form 5-aminolevulinic acid (ALA), a crucial precursor in heme synthesis. Therefore, ALA supplementation could be an effective therapeutic strategy to restore heme synthesis in CSA caused by ALAS2 defects. In a preclinical study, we examined the effects of ALA in human erythroid cells, including K562 cells and human induced pluripotent stem cell-derived erythroid progenitor (HiDEP) cells. ALA treatment resulted in significant dose-dependent accumulation of heme in the K562 cell line. Concomitantly, the treatment substantially induced erythroid differentiation as assessed using benzidine staining. Quantitative reverse transcription polymerase chain reaction (RT-PCR) analysis confirmed significant upregulation of heme-regulated genes, such as the globin genes [hemoglobin alpha ( HBA ) and hemoglobin gamma ( HBG )] and the heme oxygenase 1 ( HMOX1 ) gene, in K562 cells. Next, to investigate the mechanism by which ALA is transported into erythroid cells, quantitative RT-PCR analysis was performed on previously identified ALA transporters, including solute carrier family 15 (oligopeptide transporter) , member (SLC15A) 1 , SLC15A2 , solute carrier family 36 (proton/amino acid symporter) , member (SLC36A1) , and solute carrier family 6 (neurotransmitter transporter) , member 13 (SLC6A13) . Our analysis revealed that SLC36A1 was abundantly expressed in erythroid cells. Thus, gamma-aminobutyric acid (GABA) was added to K562 cells to competitively inhibit SLC36A1-mediated transport. GABA treatment significantly impeded the ALA-mediated increase in the number of hemoglobinized cells as well as the induction of HBG , HBA , and HMOX1 . Finally, small-interfering RNA-mediated knockdown of ALAS2 in HiDEP cells considerably decreased the expression of HBA , HBG , and HMOX1 , and these expression levels were rescued with ALA treatment. In summary, ALA appears to be transported into erythroid cells mainly by SLC36A1 and is utilized to generate heme. ALA may represent a novel therapeutic option for CSA treatment, particularly for cases harboring ALAS2 mutations. [ABSTRACT FROM AUTHOR]

The amnion membrane is developed from embryo-derived cells, and amniotic cells have been shown to exhibit multidifferentiation potential. These cells represent a desirable source for stem cells for a variety of reasons. However, to date very few molecular analyses of amnion-derived cells have been reported, and efficient markers for isolating the stem cells remain unclear. This paper assesses the characterization of amnion-derived cells as stem cells by examining stemness marker expressions for amnion-derived epithelial cells and mesenchymal cells by flow cytometry, immunocytochemistry, and quantitative PCR. Flow cytometry revealed that amnion epithelial cells expressed CD133, CD 271, and TRA-1-60, whereas mecenchymal cells expressed CD44, CD73, CD90, and CD105. Immunohistochemistry showed that both cells expressed the stemness markers Oct3/4, Sox2, Klf4, and SSEA4. Stemness genes' expression in amnion epithelial cells, mesenchymal cells, fibroblast, bone marrow-derived mesenchymal stem cells (MSCs), and induced pluripotent stem cells (iPSCs) was compared by quantitative reverse-transcription polymerase chain reaction (RT-PCR). Amnion-derived epithelial cells and mesenchymal cells expressed Oct3/4, Nanog, and Klf4 more than bone marrow-derived MSCs. The sorted TRA1-60-positive cells expressed Oct3/4, Nanog, and Klf4 more than unsorted cells or TRA1-60-negative cells. TRA1-60 can be a marker for isolating amnion epithelial stem cells. [ABSTRACT FROM AUTHOR]

Beta-hemoglobinopathies are caused by mutations in the β-globin gene. One strategy to cure this disease relies on re-activating the γ-globin expression. BCL11A is an important transcription factor that suppresses the γ-globin expression, which makes it one of the most promising therapeutic targets in β-hemoglobinopathies. Here, we performed single-gene editing and multiplex gene editing via CRISPR/Cas9 technology to edit BCL11A erythroid-specific enhancer and BCL11A binding site on γ-globin gene promoter in HUDEP-2 cells and adult human CD34+ cells. Multiplex gene editing led to higher γ-globin expression than single-gene editing without inhibiting erythroid differentiation. By further optimizing the on-target DNA editing efficiency of multiplex gene editing, the percentage of F-cells exceeded 50% in HUDEP-2 cells. Amplicon deep sequencing and whole genome sequencing were used to detect the editing frequency of on- and potential off-target sites in CD34+ cells. No off-target mutations were detected, suggesting its accuracy in HSPCs. In summary, our study provides a new approach which can be used for the treatment of β-hemoglobinopathies in the future. [ABSTRACT FROM AUTHOR]

Molecular therapies and functional studies greatly benefit from spatial and temporal precision of genetic intervention. We therefore conceived and explored tag-activated microRNA (miRNA)-mediated endogene deactivation (TAMED) as a research tool and potential lineage-specific therapy. For proof of principle, we aimed to deactivate γ-globin repressor BCL11A in erythroid cells by tagging the 3′ untranslated region (UTR) of BCL11A with miRNA recognition sites (MRSs) for the abundant erythromiR miR-451a. To this end, we employed nucleofection of CRISPR/Cas9 ribonucleoprotein (RNP) particles alongside double- or single-stranded oligodeoxynucleotides for, respectively, non-homologous-end-joining (NHEJ)- or homology-directed-repair (HDR)-mediated MRS insertion. NHEJ-based tagging was imprecise and inefficient (≤6%) and uniformly produced knock-in- and indel-containing MRS tags, whereas HDR-based tagging was more efficient (≤18%), but toxic for longer donors encoding concatenated and thus potentially more efficient MRS tags. Isolation of clones for robust HEK293T cells tagged with a homozygous quadruple MRS resulted in 25% spontaneous reduction in BCL11A and up to 36% reduction after transfection with an miR-451a mimic. Isolation of clones for human umbilical cord blood-derived erythroid progenitor-2 (HUDEP-2) cells tagged with single or double MRS allowed detection of albeit weak γ-globin induction. Our study demonstrates suitability of TAMED for physiologically relevant modulation of gene expression and its unsuitability for therapeutic application in its current form. [ABSTRACT FROM AUTHOR]

MicroRNAs (miRNAs) are small non-coding RNAs, which play an important role in various cellular and developmental processes. The study of miRNAs in erythropoiesis is crucial to uncover the cellular pathways that are modulated during the different stages of erythroid differentiation. Using erythroid cells derived from human CD34+ hematopoietic stem and progenitor cells (HSPCs)and small RNA sequencing, our study unravels the various miRNAs involved in critical cellular pathways in erythroid maturation. We analyzed the occupancy of erythroid transcription factors and chromatin accessibility in the promoter and enhancer regions of the differentially expressed miRNAs to integrate miRNAs in the transcriptional circuitry of erythropoiesis. Analysis of the targets of the differentially expressed miRNAs revealed novel pathways in erythroid differentiation. Finally, we described the application of Clustered regularly interspaced short palindromic repeats-Cas9 (CRISPR-Cas9) based editing of miRNAs to study their function in human erythropoiesis. [ABSTRACT FROM AUTHOR]

Background: Although poor liver function is associated with a high morbidity rate and poor prognosis in hepatocellular carcinoma (HCC) patients, the exact effects of liver pathology on the surgical outcomes of HCC patients are poorly understood. The purpose of this study was to assess how the liver pathology of HCC patients affects their prognosis and complications rate after liver resection. Methods: Between January 2006 and November 2010, 149 consecutive hepatocellular carcinoma patients, including 79 noncirrhosis patients and 70 cirrhosis patients, were enrolled in this study. Results: Among the noncirrhotic patients, operative time, fresh frozen plasma (FFP) transfusion requirement, tumor size, and serum retinol binding protein (RBP) levels were significantly higher in the complications group than in the complications-free groups. On the other hand, in the cirrhotic patients the prothrombin time (PT) and indocyanine green retention value at 15 min (ICGR) of the complications group were significantly lower and higher, respectively, than those of the complications-free group. In the noncirrhotic patients, recurrence-free survival and overall survival did not differ between the complications and complications-free groups. On the other hand, in the cirrhotic patients, the recurrence-free survival and overall survival of the complications-free group were significantly longer than those of the complications group. Conclusions: In the noncirrhotic patients, surgical complications had no prognostic effect, whereas they had a significant survival impact in the cirrhotic patients. The surgical strategy for HCC should be based on the patient's pathological background. [ABSTRACT FROM AUTHOR]

Transfusion of red blood cells (RBCs) is a standard and indispensable therapy in current clinical practice. In vitro production of RBCs offers a potential means to overcome a shortage of transfusable RBCs in some clinical situations and also to provide a source of cells free from possible infection or contamination by microorganisms. Thus, in vitro production of RBCs may become a standard procedure in the future. We previously reported the successful establishment of immortalized mouse erythroid progenitor cell lines that were able to produce mature RBCs very efficiently. Here, we have developed a reliable protocol for establishing immortalized human erythroid progenitor cell lines that are able to produce enucleated RBCs. These immortalized cell lines produce functional hemoglobin and express erythroid-specific markers, and these markers are upregulated following induction of differentiation in vitro. Most importantly, these immortalized cell lines all produce enucleated RBCs after induction of differentiation in vitro, although the efficiency of producing enucleated RBCs remains to be improved further. To the best of our knowledge, this is the first demonstration of the feasibility of using immortalized human erythroid progenitor cell lines as an ex vivo source for production of enucleated RBCs. [ABSTRACT FROM AUTHOR]

Although many of the frequently used pluripotency biomarkers are glycoconjugates, a glycoconjugate-based exploration of novel cellular biomarkers has proven difficult due to technical difficulties. This study reports a unique approach for the systematic overview of all major classes of oligosaccharides in the cellular glycome. The proposed method enabled mass spectrometry-baseci structurally intensive analyses, both qualitatively and quantitatively, of cellular N- and 0-linked glycans derived from glycoproteins, glyco- saminoglycans, and glycosphingolipids, as well as free oligosacchar- ides of human embryonic stem cells (hESCs), induced pluripotent stem cells (hiPSCs), and various human cells derived from normal and carcinoma cells. Cellular total glycomes were found to be highly cell specific, demonstrating their utility as unique cellular descriptors. Structures of glycans of all classes specifically observed in hESCs and hiPSCs tended to be immature in general, suggesting the presence of stem cell-specific glycosylation spectra. The current analysis revealed the high similarity of the total cellular glycome between hESCs and hiPSCs, although it was suggested that hESCs are more homogeneous than hiPSCs from a glycomic standpoint. Notably, this study enabled a priori identification of known pluripotency biomarkers such as SSEA-3, -4, and -5 and Tra-1-6O/81, as well as a panel of glycans specifically expressed by hESCs and hiPSCs. [ABSTRACT FROM AUTHOR]

Abstract: In the present study, we investigated the possible tick vectors that can transmit Theileria orientalis in eastern Hokkaido, Japan. Questing ticks collected from three different districts, Taiki, Otofuke, and Shin-Hidaka, of Hokkaido included Ixodes persulcatus, Haemaphysalis megaspinosa, Haemaphysalis douglasi, and Ixodes ovatus, while all the ticks collected from Yonaguni island of Okinawa were identified as Haemaphysalis longicornis. When the ticks were screened by polymerase chain reaction (PCR) for T. orientalis, the parasite was commonly detected among all tick species. Genotype-specific PCR assays revealed that all tick species in Hokkaido were predominantly detected with type 2, while ticks collected from Okinawa (H. longicornis) were predominantly detected with type 1. Consistent with the genetic diversity of T. orientalis in ticks, genotyping PCR assays from cattle grazed in the same Hokkaido sampling locations identified type 2 as the most prevalent genotype. This study provides the first identification of I. persulcatus, H. megaspinosa, H. douglasi, and I. ovatus as possible tick vectors of T. orientalis, and finds that the variety of vectors apparently capable of transmitting T. orientalis is wider in Japan than expected. The authors suggest that tick control strategies should be modified in Hokkaido based on the seasonal activities of ticks identified in the present study. [Copyright &y& Elsevier]

Induced pluripotent stem (iPS) cells can be established from somatic cells. However, there is currently no established strategy to generate corneal epithelial cells from iPS cells. In this study, we investigated whether corneal epithelial cells could be differentiated from iPS cells. We tested 2 distinct sources: human adult dermal fibroblast (HDF)-derived iPS cells (253G1) and human adult corneal limbal epithelial cells (HLEC)-derived iPS cells (L1B41). We first established iPS cells from HLEC by introducing the Yamanaka 4 factors. Corneal epithelial cells were successfully induced from the iPS cells by the stromal cell-derived inducing activity (SDIA) differentiation method, as Pax+/K12+ corneal epithelial colonies were observed after prolonged differentiation culture (12 weeks or later) in both the L1B41 and 253G1 iPS cells following retinal pigment epithelial and lens cell induction. Interestingly, the corneal epithelial differentiation efficiency was higher in L1B41 than in 253G1. DNA methylation analysis revealed that a small proportion of differentially methylated regions still existed between L1B41 and 253G1 iPS cells even though no significant difference in methylation status was detected in the specific corneal epithelium-related genes such as K12, K3, and Pax6. The present study is the first to demonstrate a strategy for corneal epithelial cell differentiation from human iPS cells, and further suggests that the epigenomic status is associated with the propensity of iPS cells to differentiate into corneal epithelial cells. [ABSTRACT FROM AUTHOR]

The current study was conducted to improve fissure sealing by pre-treatment with Er:YAG laser irradiation in order to remove organic debris. The surface morphology, surface roughness of fissure cavities, and the degree of microleakage after laser treatment were compared with those after bristle brush treatment in vitro. Sixty extracted human teeth were used in this study. The teeth were randomly divided into two groups of 30 each. Artificial fissures were prepared in all teeth into which artificial organic debris was placed. The debris in 30 teeth of one group was removed by means of Er:YAG laser system and the remaining 30 teeth were cleaned using a bristle brush with prophylaxis paste. Surface morphology and surface roughness of were analyzed in ten samples from each group by color laser three-dimensional (3D) microscopy and by scanning electron microscopic examination. The remaining samples were then filled with sealant and subjected to a microleakage test under thermocycling. Statistical analysis was performed using the Mann-Whitney U test; a value of p < 0.05 was considered significant. Morphologically, most of the debris was removed by Er:YAG laser treatment, whereas some fissures were not cleaned by bristle brush. However, microleakage test of both laser and etched brush methods showed similar results. Laser technique might facilitate good adaptation of resin sealant to enamel, because of an increase in surface roughness and favorable surface characteristics. [ABSTRACT FROM AUTHOR]

The generation of induced pluripotent stem cells (iPSCs) by introducing reprogramming factors into somatic cells is a promising method for stem cell therapy in regenerative medicine. Therefore, it is desirable to develop a minimally invasive simple method to create iPSCs. In this study, we generated human nasal epithelial cells (HNECs)-derived iPSCs by gene transduction with Sendai virus (SeV) vectors. HNECs can be obtained from subjects in a noninvasive manner, without anesthesia or biopsy. In addition, SeV carries no risk of altering the host genome, which provides an additional level of safety during generation of human iPSCs. The multiplicity of SeV infection ranged from 3 to 4, and the reprogramming efficiency of HNECs was 0.08--0.10%. iPSCs derived from HNECs had global gene expression profiles and epigenetic states consistent with those of human embryonic stem cells. The ease with which HNECs can be obtained, together with their robust reprogramming characteristics, will provide opportunities to investigate disease pathogenesis and molecular mechanisms in vitro, using cells with particular genotypes. [ABSTRACT FROM AUTHOR]

Non-steroidal anti-inflammatory drugs (NSAIDs) often cause gastrointestinal complications such as gastric ulcers and erosions. Recent studies on the pathogenesis have revealed that NSAIDs induce lipid peroxidation in gastric epithelial cells by generating superoxide in mitochondria, independently with cyclooxygenase inhibition and the subsequent prostaglandin deficiency. More recently, gastric hydrochloric acid (HCl) has been regarded as an inciting factor of gastric mucosal injuries, and reportedly induced cellular lipid peroxidation in vitro. We hypothesized that gastric acid and NSAID treatment synergistically induce cellular injury in gastric epithelial cells. We treated gastric epithelial RGM1 cells with acidic solutions and NSAIDs, and examined cellular injury, lipid peroxidation, mitochondrial transmenbrane potential and mitochondrial superoxide. We pretreated RGM1 cells with the acidic solutions for 0.5 h and after that treated them with each NSAID for 15 h and found that the exposure to acid and NSAIDs indeed induced cellular injury. We hypothesized that gastric acid and NSAID treatment synergistically induce mitochondrial superoxide production, which induces gastric cellular injury. Copyright © 2012 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Background: Gastric hydrochloric acid (HCl) has been regarded as an inciting factor in gastric mucosal injuries and has been reported to induce lipid peroxidation in vitro. However, because HCl is not an oxidant per se, the exact mechanism by which the acid induces lipid peroxidation is unknown. We hypothesized that gastric acid may disrupt mitochondrial transmembrane potential and induce the production of superoxide in mitochondria, which subsequently may induce lipid peroxidation and apoptosis in gastric mucosal cells. Methods: Firstly we treated gastric epithelial RGM1 cells with solutions containing various concentrations of HCl (i.e., of varying pH), and examined cellular injury, lipid peroxidation, and apoptosis with specific fluorescent dyes. Secondly, we performed electron paramagnetic resonance (EPR) spectroscopy of isolated, acid-exposed mitochondria from the cells, using a spin-trapping reagent for superoxide, 5-(2,2-dimethyl-1,3-propoxy cyclophosphoryl)-5-methyl-1-pyrroline N-oxide (CYPMPO). Finally, we established novel RGM1 cells that overexpressed manganese superoxide dismutase (MnSOD), which removes superoxide from mitochondria, and examined the effect of acid treatment on cellular membrane lipid peroxidation. Results: The results indicated that the exposure to acid indeed induced cellular injury, cellular lipid peroxidation, apoptosis, and the demonstration of the exact superoxide spectra on EPR spectroscopy in gastric epithelial cells, and that overexpression of MnSOD decreased superoxide production and prevented cellular lipid peroxidation. Conclusion: These results suggested that gastric acid, like nonsteroidal anti-inflammatory drugs (NSAIDs), induces mitochondrial superoxide production, which induces gastric cellular injury by triggering cellular lipid peroxidation and apoptosis. [ABSTRACT FROM AUTHOR]

Background: Mesothelioma is a highly malignant tumor that is primarily caused by occupational or environmental exposure to asbestos fibers. Despite worldwide restrictions on asbestos usage, further cases are expected as diagnosis is typically 20- 40 years after exposure. Once diagnosed there is a very poor prognosis with a median survival rate of 9 months. Considering this the development of early pre clinical diagnostic markers may help improve clinical outcomes. Methodology: Microarray expression arrays on mesothelium and other tissues dissected from mice were used to identify candidate mesothelial lineage markers. Candidates were further tested by qRTPCR and in-situ hybridization across a mouse tissue panel. Two candidate biomarkers with the potential for secretion, uroplakin 3B (UPK3B), and leucine rich repeat neuronal 4 (LRRN4) and one commercialized mesothelioma marker, mesothelin (MSLN) were then chosen for validation across a panel of normal human primary cells, 16 established mesothelioma cell lines, 10 lung cancer lines, and a further set of 8 unrelated cancer cell lines. Conclusions: Within the primary cell panel, LRRN4 was only detected in primary mesothelial cells, but MSLN and UPK3B were also detected in other cell types. MSLN was detected in bronchial epithelial cells and alveolar epithelial cells and UPK3B was detected in retinal pigment epithelial cells and urothelial cells. Testing the cell line panel, MSLN was detected in 15 of the 16 mesothelioma cells lines, whereas LRRN4 was only detected in 8 and UPK3B in 6. Interestingly MSLN levels appear to be upregulated in the mesothelioma lines compared to the primary mesothelial cells, while LRRN4 and UPK3B, are either lost or down-regulated. Despite the higher fraction of mesothelioma lines positive for MSLN, it was also detected at high levels in 2 lung cancer lines and 3 other unrelated cancer lines derived from papillotubular adenocarcinoma, signet ring carcinoma and transitional cell carcinoma. [ABSTRACT FROM AUTHOR]

Introduction: The branched-chain amino acid/tyrosine ratio (BTR) reflects the amino acid balance and the severity of liver disease. The aim of the present study was to determine the relationship between BTR and liver function in patients with hepatocellular carcinoma (HCC). Furthermore, we evaluated the clinical usefulness of BTR as a prognostic indicator of disease-free and overall patient survival after initial hepatectomy.Methods: Between January 2004 and December 2008, 105 consecutive HCC patients who underwent initial hepatectomy were enrolled in this study. The correlation between BTR and preoperative liver functional indicators was evaluated. The cutoff levels of BTR for 2-year survival prediction were evaluated using a dot blot diagram. The patients were divided into high BTR (4.5 or higher) and low BTR (4.4 or lower) groups and these were compared in terms of clinical variables such as liver functional indicators, operative variables, and tumor characteristics.Results: The preoperative BTR level decreased according to the severity of liver disease. BTR was correlated with the albumin, bilirubin, and prealbumin levels, as well as the prothrombin time. Although the preoperative liver function was significantly different between the high BTR and low BTR groups, the operative variables and tumor-related variables were not found to be significantly different. Postoperative complications in the high BTR group were significantly less frequent than in the low BTR group (p = 0.003). Disease-free and overall patient survival in the high BTR group were significantly longer than in the low BTR group (p < 0.001 and p = 0.021, respectively).Conclusions: BTR reflected the pathological liver background with a high correlation to the other liver functional indicators. BTR is thus considered to be a useful marker to predict postoperative complications, disease-free survival, and overall survival of HCC patients after initial hepatectomy. It is, therefore, a useful indicator of liver function and a predictor for the risk of cancer recurrence and overall survival in HCC patients. [ABSTRACT FROM AUTHOR]

The Sonoda–Tajima Cell Collection includes cell samples obtained from a range of ethnic minority groups across the world but in particular from South America. The collection is made all the more valuable by the fact that some of these ethnic populations have since died out, and thus it will be impossible to prepare a similar cell collection again. The collection was donated to our institute, a public cell bank in Japan, by Drs Sonoda and Tajima to make it available to researchers throughout the world. The original cell collection was composed of cryopreserved peripheral blood samples that would obviously have been rapidly exhausted if used directly. We, therefore, immortalized some samples with the Epstein–Barr virus and established B-lymphoblastoid cell lines (B-LCLs). As there is continuing controversy over whether the B-LCL genome is stably maintained, we performed an array comparative genomic hybridization (CGH) analysis to confirm the genomic stability of the cell lines. The array CGH analysis of the B-LCL lines and their parental B cells demonstrated that genomic stability was maintained in the long-term cell cultures. The B-LCLs of the Sonoda–Tajima Collection will therefore be made available to interested scientists around the world. At present, 512 B-LCLs have been developed, and we are willing to increase the number if there is sufficient demand. [ABSTRACT FROM PUBLISHER]

Purpose: A previous meta-analysis study demonstrated that bleeding and the duration of the hospital stay following laparoscopic hepatectomy (Lap) were significantly smaller and shorter, respectively, than for patients undergoing an open approach (Op). The aim of the present study was to re-evaluate perioperative variables and adverse outcomes in patients undergoing Lap versus (vs) Op after 2000. Methods: A PubMed and Ovid Medline search identified clinical studies that compared the outcomes of Lap vs Op patients after 2000. A meta-analysis and power analysis were performed. Results: Operative time was not significantly different between the two approaches (95% confidence interval [CI]: −0.063 to 0.992). Patient bleeding in the Lap group was significantly lower than in the Op group (95% CI: −1.027 to −0.390). Complications with Lap patients were significantly less frequent (95% CI: 0.231-0.642), and the duration of the hospital stay for Lap patients was significantly shorter (95% CI: −0.950 to −0.530) than for Op patients. Only one paper presented 80% power with 0.05 α-errors in all four outcomes, whereas four studies did not have sufficient statistical power. Conclusions: The clinical benefits of Lap include a smaller incidence of complications and a shorter duration of hospital stay at the current time. Several studies had too few cases to sufficiently evaluate these factors, although other studies were appropriately analyzed. [ABSTRACT FROM AUTHOR]

Misidentification of cultured cell lines results in the generation of erroneous scientific data. Hence, it is very important to identify and eliminate cell lines with a different origin from that being claimed. Various methods, such as karyotyping and isozyme analysis, can be used to detect inter-species misidentification. However, these methods have proved of little value for identifying intra-species misidentification, and it will only be through the development and application of molecular biological approaches that this will become practical. Recently, the profiling of microsatellite variants has been validated as a means of detecting gene polymorphisms and has proved to be a simple and reliable method for identifying individual cell lines. Currently, the human cell lines provided by cell banks around the world are routinely authenticated by microsatellite polymorphism profiling. Unfortunately, this practice has not been widely adopted for mouse cells lines. Here we show that the profiling of microsatellite variants can be also applied to distinguish the commonly used mouse inbred strains and to determine the strain of origin of cultured cell lines. We found that approximately 4.2% of mouse cell lines have been misidentified; this is a similar rate of misidentification as detected in human cell lines. Although this approach cannot detect intra-strain misidentification, the profiling of microsatellite variants should be routinely carried out for all mouse cell lines to eliminate inter-strain misidentification. [ABSTRACT FROM AUTHOR]

Abstract: Introduction: The aim of this study was to evaluate N-benzyl-N-[11C]methyl-2-(7-methyl-8-oxo-2-phenyl-7,8-dihydro-9H-purin-9-yl)acetamide ([11C]DAC) as a novel peripheral-type benzodiazepine receptor (PBR) ligand for tumor imaging. Methods: [11C]DAC was synthesized by the reaction of a desmethyl precursor with [11C]CH3I. In vitro uptake of [11C]DAC was examined in PBR-expressing C6 glioma and intact murine fibrosarcoma (NFSa) cells. In vivo distribution of [11C]DAC was determined using NFSa-bearing mice and small-animal positron emission tomography (PET). Results: [11C]DAC showed specific binding to PBR in C6 glioma cells, a standard cell line with high PBR expression. Specific binding of [11C]DAC was also confirmed in NFSa cells, a target tumor cell line in this study. Results of PET experiments using NFSa-bearing mice, showed that [11C]DAC was taken up specifically into the tumor, and pretreatment with PK11195 abolished the uptake. Conclusions: [11C]DAC was taken up into PBR-expressing NFSa. [11C]DAC is a promising PET ligand that can be used for imaging PBR in tumor-bearing mice. [Copyright &y& Elsevier]

We previously reported that long-lasting in vitro hematopoiesis could be achieved using the cells differentiated from primate embryonic stem (ES) cells. Thus, we speculated that hematopoietic stem cells differentiated from ES cells could sustain long-lasting in vitro hematopoiesis. To test this hypothesis, we investigated whether human hematopoietic stem cells could similarly sustain long-lasting in vitro hematopoiesis in the same culture system. Although the results varied between experiments, presumably due to differences in the quality of each hematopoietic stem cell sample, long-lasting in vitro hematopoiesis was observed to last up to nine months. Furthermore, an in vivo analysis in which cultured cells were transplanted into immunodeficient mice indicated that even after several months of culture, hematopoietic stem cells were still present in the cultured cells. To the best of our knowledge, this is the first report to show that human hematopoietic stem cells can survive in vitro for several months. [ABSTRACT FROM AUTHOR]

The differential roles of circulating and intrarenal renin-angiotensin system (RAS) in glomerulonephntis have not been elucidated. In this study, we investigated the levels of circulating and intrarenal RAS activity and urinary angiotensinogen (AGT) excretion in anti-thymocyte serum (ATS) nephritis induced by an ATS injection (ATS group). The effect of olmesartan, an angiotensin II (ANG II) type 1 receptor blocker (ARB), on the development of nephritis was also examined (ATS+ARB group). In addition, the rats received a saline injection instead of ATS (control group). Mesangial proliferation with transient proteinuria, which peaked at day 7, was significantly increased in the ATS group compared with the control group. The levels of glomerular AGT mRNA, intrarenal ANG II, and urinary AGT excretion in the ATS group were increased significantly at day 7 compared with the control group. Administration of olmesartan (ATS+ARB group) significantly decreased the levels of renal lesions, proteinuria, and intrarenal RAS activity compared with the ATS group. In addition, the levels of urinary AGT excretion correlated with the levels of glomerular damage, urinary protein excretion, and immunoreactivity for AGT and ANG II in kidney. On the other hand, plasma renin activity was significantly lower in the ATS group compared with the control group and significantly higher in the ATS+ARB group than in the ATS group. These data suggest that an increase in kidney-specific RAS activity, which parallels urinary AGT excretion, plays an important role in the development of ATS nephritis. [ABSTRACT FROM AUTHOR]

Lipocalin 2 (LCN2), a secreted protein of the lipocalin family, induces apoptosis in some types of cells and inhibits bacterial growth by sequestration of the iron-laden bacterial siderophore. We have recently reported that LCN2 inhibits the production of red blood cells in the mouse. Here we analyzed the role of LCN2 in human hematopoiesis. Expression of LCN2 was observed not only in mature cells such as those of the granulocyte/macrophage and erythroid lineages but also in hematopoietic stem/progenitor cells. We also examined expression of two candidate receptors for LCN2, brain type organic cation transporter (BOCT) and megalin, in various cell types. BOCT showed relatively high levels of expression in erythroid and hematopoietic stem/progenitor cells but lower levels in granulocyte/macrophage and T lymphoid cells. Megalin was expressed at high levels in T lymphoid and erythroid cells but at lower levels in granulocyte/macrophage lineage cells. LCN2 suppressed the growth of erythroid and monocyte/macrophage lineages in vitro, but did not have this effect on cells of other lineages. In addition, immature hematopoietic stem/progenitor cells were not sensitive to LCN2. These results demonstrate a lineage-specific role for LCN2 in human hematopoiesis that is reminiscent of its effects upon mouse hematopoiesis and strongly suggest an important in vivo function of LCN2 in the regulation of human hematopoiesis. J. Cell. Physiol. 215: 526–537, 2008. © 2007 Wiley-Liss, Inc. [ABSTRACT FROM AUTHOR]

Abstract: Borealin/DasraB is a member of the chromosomal passenger protein complex (CPC) required for proper segregation of chromosomes during mitosis. In Drosophila melanogaster, inactivation of Borealin/DasraB results in polyploidy, delayed mitosis and abnormal tissue development, indicating its critical role for cell proliferation. However, the in vivo role of mammalian Borealin/DasraB remains unclear. Here, we analyzed the expression of Borealin/DasraB and found that borealin is widely expressed in embryonic tissues and later restricted to adult tissues which relies on rapid cell proliferation. To determine the role of borealin during mouse development, we generated borealin-null mice through targeted disruption. While heterozygous mice developed normally, disruption of both borealin alleles resulted in early embryonic lethality by 5.5dpc (days postcoitus) due to mitotic defects and apoptosis in blastocyst cells that showed microtubule disorganization and no CPC enrichment. At 5.5dpc, borealin-null embryos exhibited excessive apoptosis and elevated expression of p53. However, loss of p53 did not abrogate or delay embryonic lethality, revealing that Borealin/DasraB inactivation triggered impaired mitosis and apoptosis though p53-independent mechanisms. Our data show that Borealin/DasraB is essential for cell proliferation during early embryonic development, and its early embryonic lethality cannot be rescued by the loss of p53. [Copyright &y& Elsevier]

While normal epithelial cells are anchorage-dependent, cancer cells are anchorage-independent. To elucidate the mechanism underlying the anchorage-independence of cancer cells, we cultured detached cells in medium containing elastase. Detached human epidermal cancer cells (DJM-1) survived for at least 3 weeks and focal adhesion kinase was still phosphorylated. In contrast, most detached keratinocytes underwent rapid apoptosis and focal adhesion kinase was not phosphorylated while the cells were alive. Thus, discontinuation of the phosphorylation of focal adhesion kinase preceded cell death. Immunostaining showed laminin-5 expression on the surface of detached DJM-1 cells, but not on detached keratinocytes. Receptors for laminin-5 (i.e. integrins) were detected on both detached DJM-1 cells and keratinocytes. Laminins are secreted proteins, so we speculated that laminin-5 adhered to the surface of secreting DJM-1 cells via integrins and evoked activation of focal adhesion kinase, with the resultant signalling cascade promoting cellular survival. If this hypothesis is correct, cell surface expression of laminin-5 may be used to explain the characteristics of cancer, immortality, tumour formation, metastasis and angiogenesis. [ABSTRACT FROM AUTHOR]

Background. The supply of transfusable red blood cells (RBCs) is not sufficient in many countries. If erythroid cell lines able to produce transfusable RBCs in vitro were established, they would be valuable resources. However, such cell lines have not been established. To evaluate the feasibility of establishing useful erythroid cell lines, we attempted to establish such cell lines from mouse embryonic stem (ES) cells. Methodology/Principal Findings. We developed a robust method to obtain differentiated cell lines following the induction of hematopoietic differentiation of mouse ES cells and established five independent hematopoietic cell lines using the method. Three of these lines exhibited characteristics of erythroid cells. Although their precise characteristics varied, each of these lines could differentiate in vitro into more mature erythroid cells, including enucleated RBCs. Following transplantation of these erythroid cells into mice suffering from acute anemia, the cells proliferated transiently, subsequently differentiated into functional RBCs, and significantly ameliorated the acute anemia. In addition, we did not observe formation of any tumors following transplantation of these cells. Conclusion/Significance. To the best of our knowledge, this is the first report to show the feasibility of establishing erythroid cell lines able to produce mature RBCs. Considering the number of human ES cell lines that have been established so far, the intensive testing of a number of these lines for erythroid potential may allow the establishment of human erythroid cell lines similar to the mouse erythroid cell lines described here. In addition, our results strongly suggest the possibility of establishing useful cell lines committed to specific lineages other than hematopoietic progenitors from human ES cells. [ABSTRACT FROM AUTHOR]

Abstract: Objective: The objective of the present study was to improve fissure sealing by pre-treatment with Carisolv in order to remove organic debris. The surface morphology and roughness of fissure cavities and the degree of microleakage after Carisolv application were compared with those after bristle brush treatment in vitro. Methods: Fifty extracted human teeth were used in this study. The teeth were randomly divided into two groups of 25 each. Artificial fissures were prepared in all teeth into which artificial organic debris was placed. The debris in 25 teeth of one group was removed using Carisolv applied for 30s and excavation was performed with a dental explorer until the gel was clear. The remaining 25 teeth were then cleaned using bristle brush with prophylaxis paste. Surface roughness was analyzed in five samples from each group by color laser three-dimensional (3D) microscopy and by scanning electron microscopic examination. The remaining samples were filled with sealant and subjected to a microleakage test under thermocycling. Statistical analysis was performed using the Mann–Whitney U-test; a value of p <0.05 was considered significant. Results: Morphologically, most of the debris in the fissures was removed by Carisolv treatment, whereas some fissures were not cleaned by bristle brush. Carisolv-treatment with acid-etching resulted in removal of debris-like smear layer leaving enamel prisms open, and 3D laser microscopy demonstrated that the roughness values increased after acid-etching. However, microleakage test of both Carisolv and brush methods showed similar results. Conclusions: Carisolv-treated surfaces especially when subjected to acid-etching might facilitate good adaptation of sealant to enamel, because of an increase in surface roughness and favorable surface characteristics. [Copyright &y& Elsevier]

The article reports on the results of a study of the coexistence of Ser84 in renin and His13 in angiotensinogen which brings a pH profile of two separate peaks to the reaction of human renin and sheep angiotensionogen. A description of the experimental set-up and measurement methods is presented. The study indicated that sheep angiotensinogen has a factor other than His13 to enhance the activity of human renin at basic pH.

Abstract: Clinical application of human embryonic stem (ES) cells will require the establishment of methods for their culture, either in the presence or absence of human-derived feeder cells. We have tested the ability of non-immortalized cultured cells derived from human umbilical cord (HUC cells) to support ES cell culture. A primate ES cell line that had been established and maintained with mouse embryonic fibroblasts was cultured on HUC cells for >3months (HUC-maintained ES cells). These cells retained their expression of alkaline phosphatase, SSEA-4, Oct-3/4, and to a lesser extent Nanog, but did not express Rex-1. Nevertheless, HUC-maintained ES cells could produce ectoderm-, mesoderm- and endoderm-derived cells in teratomata that they formed in immunodeficient mice. We show that HUC-maintained ES cells could give rise to hematopoietic cells, although this ability of HUC cells varied among HUC cell populations derived from different neonates. HUC cells are promising as human material with which to maintain ES cells in a state that retains their ability to produce mature cells, including hematopoietic cells. [Copyright &y& Elsevier]

The article investigates the pH dependence of the reaction of various renins using sheep angiotensinogen as a substrate. The pH profile of the human renin reaction showed two separate peaks against sheep angiotensinogen. The article indicates that Ser84 of human renin contributes to the biphasic pH dependence of the renin-angiotensinogen reaction.

The amino acid sequence His-Pro-Phe as N-terminal residues 6–8 of the natural renin substrate, angiotensinogen, is conserved among species. We investigated whether this His-Pro-Phe motif functions as the determinant of the substrate specificity of renin. Mutant angiotensinogens in which the Ile-His-Pro-Phe-His-Leu sequence at positions 5–10 of wild-type angiotensinogen was replaced by either His-Pro-Phe-His-Leu-Leu or Ala-Ile-His-Pro-Phe-His were cleaved by renin at the C-terminal side of residues 9 and 11, respectively, while wild-type angiotensinogen was cleaved at residue 10. A triple Ala substitution for the His-Pro-Phe motif of angiotensinogen prevented its cleavage by renin. In contrast, triple Ala substitution for residues 9–11, including the natural site of cleavage by renin, allowed cleavage between the two Ala residues at positions 10 and 11. Furthermore, the 33-residue C-terminal peptide of human megsin, which carries a naturally occurring His-Pro-Phe sequence, was cleaved by renin at the C-terminal side of the His-Pro-Phe-Leu-Phe sequence. These results indicate that the His-Pro-Phe motif of angiotensinogen is a crucial determinant of the substrate specificity of renin. By binding to a corresponding pocket on renin, the His-Pro-Phe motif may act as a molecular anchor to recruit the scissile peptide bond to a favorable site for catalysis. [ABSTRACT FROM AUTHOR]

The article focuses on the production of human prorenin attached by a decahistidine tag at the C-terminus in Chinese hamster ovary cells. The tagged protein secreted into the culture medium was activated to mature renin by proteolytic removal of its prosegment by trypsin in the same manners a native prorenin. The tagged protein was also in the inactive prorenin form.

Objective: Induction of hematopoietic cells from human embryonic stem (ES) cells has been reported recently. However, before cells derived from human ES cells can be used in the clinic, preclinical studies using these cells in experimental primates will be necessary. Therefore, we attempted to establish a method to induce hematopoietic cells robustly and abundantly from primate ES cells. Methods: A primate ES cell line, CMK-6, derived from the cynomolgus monkey was used in this study. We adapted a method to induce hematopoiesis from CMK-6 cells on feeder cells, and tested the effectiveness of three kinds of feeder cell lines (OP9, C2C12, and C3H10T1/2). In addition, we tested the effect of vascular endothelial growth factor (VEGF) and insulin-like growth factor-II (IGF-II) on hematopoiesis induction from CMK-6 cells. Results: VEGF and IGF-II showed an extremely strong synergistic effect to induce hematopoiesis from CMK-6 cells. C3H10T1/2 cells proved to be very useful for the induction of hematopoiesis from CMK-6 cells, and the production of blood cells on C3H10T1/2 cells has been maintained as long as 5 months. During this long period, ES cell derivatives continuously produced mature blood cells, including terminally differentiated cells. Conclusion: We have developed an original method to produce enriched blood cells abundantly from primate ES cells for an extremely long period. This method may represent a good in vitro model for studying primate hematopoiesis and related diseases. Furthermore, our method may be useful for preclinical studies of transfusion therapy using blood cells derived from ES cells in experimental primate systems. [Copyright &y& Elsevier]

Abstract: Angiotensin III (Ang III) as well as angiotensin II (Ang II) suppressed body weight loss of the clam worm Perinereis sp. under a hyper-osmotic condition, and enhanced body weight gain under a hypo-osmotic condition. Under a drying condition where the water inflow from outside the body was eliminated, Ang II suppressed body weight loss, but Ang III did not. Under these conditions, angiotensins I, IV, and (1–7) had no effect, and saralasin blocked the effects of Ang II and Ang III. It is concluded that Ang II and Ang III upregulate body fluid volume of the clam worm via Ang II receptors in different ways. [Copyright &y& Elsevier]

Examines the role of angiotensins II and III in regulating water flow through aquaporins in clam worms of Perinereis. Treatment of the clam worms with tetrachloroaurate (III) before and after angiotensin treatment; Function of tetrachloroaurate as blocker of aquaporins; Role of the angiotensins in increasing body weight in clam worms under a hypo-osmotic condition and decreasing body weight under a hyper-osmotic condition.

Background. Tissue hypoxia stimulates the induction of the angiogenic substances vascular endothelial growth factor and erythropoietin in the locus of the tissue. We have previously demonstrated that erythropoietin promotes angiogenesis by binding to its receptor in the endothelial cells of uterine and ovarian malignancies. In the present study, we examined whether malignant uterine cervix tissue showed hypoxia and whether hypoxia correlated with high vascular density through vascular endothelial growth factor. Methods. To detect tissue hypoxia, we estimated the content of ATP in squamous cell carcinoma of the uterine cervix and in the normal cervix, using liquid chromatography columns. Surgically resected samples were fixed in Zamboni solution and processed for immunohistochemical microscopy to identify the endothelial cells and the location of vascular endothelial growth factor, with the use of antifactor VIII and anti-vascular endothelial growth factor antibody, respectively. The microvessels in a definite area were counted in sections of each specimen. Results. Significantly lower ATP levels and significantly higher vascular density were seen in squamous cell carcinoma than in the controls (P _ 0.05). The microvessel number in relation to ATP content was significantly higher in squamous cell carcinoma than in the controls (P < 0.001). Moreover vascular endothelial growth factor, the hyperplastic epithelium of the squamous cell carcinoma contained the immunoreactivity, with characteristic histopathological features suggesting retention of tissue fluid. Conclusion. Squamous cell carcinoma of the uterine cervix showed hypoxia which correlated with abundant vascularity. Vascular endothelial growth factor expressed in the hyperplastic epithelium appears to promote angiogenesis in squamous cell carcinoma of the uterine cervix. [ABSTRACT FROM AUTHOR]

Dynamical behavior of optical gain formation has been assessed at room temperature in the In[sub x]Ga[sub 1-x]N multi-quantum-well (MQW) based laser diodes (LDs) by employing pump and probe spectroscopy with a pulse width of 150 fs. The LDs are composed of (a) In[sub 0.1]Ga[sub 0.9]N-In[sub 0.02]Ga[sub 0.98]N MQW and (b) In[sub 0.3]Ga[sub 0.7]N-In[sub 0.05]Ga[sub 0.95]N MQW, whose stimulated emissions correspond to near ultraviolet (390 nm) and blue (440 nm), respectively. The optical gain was contributed from the nearly delocalized states [the lowest-quantized MQW levels (LQL)] in the sample (a), while it was from highly localized levels with respect to the LQL by 500 meV for the sample (b). It was found that the photogenerated carriers rapidly (less than 1 ps) transferred to the LQL, and then relaxed to the localized tail within the time scale of about 5 ps, giving rise to the optical gain. Such gain spectra were saturated and other bands appeared in the vicinity of the LQL under higher photoexcitation. © 2000 American Institute of Physics. [ABSTRACT FROM AUTHOR]

Surveys of the estrogenic activity in coastal areas of Japan were performed by using two serum vitellogenins (Vg-530 and Vg-320) of male Japanese common goby Acanthogobius flavimanus as biomarkers. To evaluate the relationship between the Vg concentrations in the serum of male fish and the concentrations of environmental estrogens in seawater, 3 week exposure experiments were performed using estradiol (E2), nonylphenol (NP) and bisphenol A (BPA). From the results of the E2 exposure, mean concentrations of the Vg and the ratio of Vg-positive fish increased from actual concentration at 10.5 ng/L E2 treatment. Exposure of NP and BPA showed that the 19.0 µg/L NP treatment induced the Vg but 133.7 µg/L BPA did not. As for the survey of the estrogenic activity in coastal areas, the mean Vg concentrations and ratio of Vg-positive fish in some urban areas were similar to values of the 10.5 ng/L E2 treatment. Although E2 and NP were detected in the seawater from urban areas, the concentrations were concluded as not sufficient to independently induce the Vg. Considering these results, it is concluded that the total estrogenic activity in some urban areas is dependent not only on E2 but synergistically with other environmental estrogens. [ABSTRACT FROM AUTHOR]

Investigates the mechanism for non-proteolytic activation of human prorenin using five kinds of antibodies. Use of the tertiary structure of predicted prorenin to design each of the antigens; Blunting re-inactivation of acid-activated prorenin by specific antibodies.

Because blood vitellogenin (Vg) has been considered a biomarker for environmental estrogens, the basal levels of Vg and 17β-estradiol (E2 ) were determined in male Japanese whiting reared under natural conditions. Serum levels of Vg and E2 were measured and gonadal development was assessed by gonadosomatic index (GSI) and histological observation in 8–10 male fish at monthly intervals throughout the annual reproductive cycle. Serum E2 was <60 pg/mL throughout the study period. In contrast, serum Vg exhibited seasonal changes: serum levels of Vg gradually increased from April to May (mean 63 ± 13 ng/mL and 124 ± 48 ng/mL in April and May, respectively), and then reached a peak value (mean 352 ± 68 ng/mL) in June. Thereafter, serum Vg gradually decreased, reaching undetectable levels (<50 ng/mL) in October. Serum levels of Vg tended to increase in the male fish in which the GSI was >1%. Histological observation revealed that testes in such male fish were in active spermatogenesis and then all of the testes of male fish in which serum Vg decreased to ND levels were regressed. These results suggest that Vg productive potency (sensitivity to estrogens) may increase in the spermatogenic stage, resulting in production of Vg in response to very low levels of natural or xenobiotic estrogens. [ABSTRACT FROM AUTHOR]

Two vitellogenins (Vgs) were detected in serum from estradiol-17β (E2)-injected Japanese common goby (Acanthogobius flavimanus). Vitellogenins with molecular masses of 530 kDa (Vg-530) and 320 kDa (Vg-320) were purified, and used to raise specific antisera in rabbits. Sandwich enzyme-linked immunosorbent assays (ELISAs) for Vg-530 and Vg-320 were developed using the antisera and the isolated Vgs. The sensitivity ranges of these ELISAs were 1.25–160 ng/ml for Vg-530 and 0.26–66 ng/ml for Vg-320, and very low cross-reactivity was found with the alternate Vg in each assay. Treatment of male gobys with E2 by injection and immersion induced both Vgs in sera in a dose-dependent manner. The mean concentrations of the Vgs increased from 10 ng/L E2 exposure for three weeks. Serum concentrations of the two Vgs in field-collected maturing females increased in accordance with increment of E2 level and ovarian development, and the mean concentrations of Vg-530 were higher than those of Vg-320 in maturing female. These results indicate that the sandwich ELISAs for Vg-530 and Vg-320 developed in the present study is useful as an assay system for surveys of estrogenic activity in coastal areas of Japan. [Copyright &y& Elsevier]

The incisors of 21 Wistar rats were transected, pulp tissue was extirpated for 10 mm from the level of the gingival margin and each canal was prepared with files. The fibre tip of a pulsed neodymium:yttrium–aluminium–garnet laser was inserted into the root canal for 10 mm and laser irradiation delivered at 2 W and 20 pulses/s for 10 s. After 6 weeks the mandibles were removed and sectioned. Sections were stained either with haematoxylin and eosin or immunohistochemically using polyclonal antibodies against keratin/cytokeratin, amelogenin and type I collagen. The inner epithelial cells on the labial side differentiated into ameloblasts in animals where eruption had recovered. The pulp cells differentiated into odontoblast-like cells and staining for type I collagen was evident in pulp cells, odontoblast-like cells and inside dentinal tubules. In animals where eruption had ceased, the inner epithelial cells on the labial side did not differentiate into ameloblasts. Staining for type I collagen was observed in the mineralized nodules and tubules of dentine-like hard tissues in the pulp cavity. These results suggest that differentiation of epithelial cells on the labial side into ameloblasts is involved in the re-eruption process. [Copyright &y& Elsevier]

Examines the cryogenic stability and mechanical behavior of superconducting magnet helical coils used for Large Hadron Collider. Magnetic diffusion in an aluminum stabilizer; Balance voltage of the helical coils; Pulse height analysis.

ABSTRACT: To determine the thermal limit for normal spawning of Japanese whiting and to evaluate thermal effects on spawning in the normal temperature range, two experiments were conducted. In Experiment 1, the temperature of three experimental groups (Groups A, B and C) was increased from 25 to 28, 31 or 33°C, respectively. Subsequently, temperatures in all groups were decreased to 28°C. During the experimental period, the number of eggs, egg size, and hatching rate were monitored. According to the results, the upper thermal limit of normal spawning was considered to be 28–29°C, because the number of eggs and the hatching rate decreased at temperatures over 28°C. However, egg size tended to be reduced at 28°C, and was found to be strongly dependent on temperature. In Experiment 2, the number of eggs, egg size and time of spawning were compared between fish held at a constant temperature of 22°C (Group CT) and those held at increasing temperatures from 22 to 28°C (Group IT). Group IT spawned more in number and had smaller-sized eggs than did Group CT, while spawning was delayed in group IT. To identify the thermal effects on spawning of Japanese whiting, the mechanisms governing egg size must be determined. [ABSTRACT FROM AUTHOR]

The Rel/nuclear factor (NF)-κB family of transcription factors have been implicated in the regulation of a wide variety of genes, in particular those encoding proteins crucial to the function of the immune system. Through the use of mutant mice that lack one or more of these proteins, we have begun to examine the individual and combined roles of Rel, RelA and NF-κB1 and B-cell development and function. Here we outline and discuss how these transcription factors operate as differentiation stage-specific regulators of B-cell development, survival, division and immunoglobulin expression, emphasizing those Rel/NF-κB-regulated genes that mediate these functions. [ABSTRACT FROM AUTHOR]