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Microalgae, Botryococcus braunii in particular, have received increasing interest owing to their potential as biofuel sources. Although the fertilizer components present in wastewater are useful in reducing the cost of commercial production of microalgae, the ammonium nitrogen (NH4+-N) acidifies the medium and may inhibit the growth of B. braunii. In this study, we aimed to investigate the effectiveness of calcium carbonate in preventing the growth inhibition of B. braunii by suppressing pH decrease caused by NH4+-N. Four types of modified Chu13 media were prepared: a control, one with NH4+-N, one with calcium carbonate, and one with calcium carbonate and NH4+-N. Then, the pH, NH4+-N concentration, B. braunii growth, and hydrocarbon content were measured. We found that in the NH4+-N treatment without calcium carbonate, the pH decreased to approximately 4, and there was almost no algal growth even after 20 days. In contrast, there were no differences in the growth rate and medium pH (7–8) between the treatment with calcium carbonate and the control medium. In addition, calcium ions were leached into the medium with calcium carbonate as the culture progressed, confirming the effectiveness of calcium carbonate in maintaining a constant pH and thus preventing growth inhibition. As such, the addition of calcium carbonate effectively prevents B. braunii growth inhibition by suppressing pH decrease and is more cost-effective than aggressive pH adjustment with alkali or acid or by adding buffer solutions into the medium. This simple and inexpensive method can be used for the removal of ammonia from wastewater.

The intracellular trafficking pathway of albumin in podocytes remains controversial. We therefore analysed albumin endocytosis through caveolae, subsequent transcytosis, and exocytosis. In Western blot and immunofluorescence analysis in vitro, methyl-beta-cyclodextrin (MBCD) treatment significantly decreased the expression of caveolin-1 and albumin in cultured human podocytes after incubation with albumin; additionally, MBCD interfered with albumin endocytosis through caveolae in the experiment using Transwell plates. In the immunofluorescence analysis, albumin was incubated with cultured human podocytes, and colocalisation analysis with organelles and cytoskeletons in the podocytes showed that albumin particles colocalised with caveolin-1 and Fc-receptor but not clathrin in endocytosis, colocalised with actin cytoskeleton but not microtubules in transcytosis, and colocalised with early endosomes and lysosomes but not proteasome, endoplasmic reticulum, or Golgi apparatus. In the electron microscopic analysis of podocytes in nephrotic syndrome model mice, gold-labelled albumin was shown as endocytosis, transcytosis, and exocytosis with caveolae. These results indicate the intracellular trafficking of albumin through podocytes. Albumin enters through caveolae with the Fc-receptor, moves along actin, and reaches the early endosome, where some of them are sorted for lysosomal degradation, and others are directly transported outside the cells through exocytosis. This intracellular pathway may be a new aetiological hypothesis for albuminuria.

Introduction: Previously, we reported the caveolae-mediated intracellular trafficking pathway of albumin through glomerular endothelial cells (GEnCs) as a new etiological hypothesis of urinary albumin excretion. The selective serotonin reuptake inhibitor, sertraline (Ser), inhibits dynamin, which plays a pivotal role in the fission of caveolae from the cell membrane during caveolae endocytosis. Objective: In this study, we evaluated whether Ser reduces albuminuria levels by interfering with albumin endocytosis through caveolae into GEnCs and podocytes as a novel treatment for glomerulonephritis. Methods: After treating the cells with Ser, albumin and caveolin-1 (Cav-1) expression levels were evaluated by immunofluorescence (IF) and western blot (WB) analyses. The albuminuria level was determined by histology in a puromycin aminonucleoside (PAN)-induced nephrotic syndrome mouse model (PAN mice) treated with or without Ser. Results: IF and WB analyses showed that the albumin expression level was significantly decreased by Ser treatment; however, Cav-1 expression was not decreased in GEnCs or podocytes based on the IF results. In PAN mice treated with or without Ser, Cav-1 expression increased, and the foot process effacement of podocytes and swelling of GEnCs were observed. However, proteinuria levels were not increased in PAN mice treated with Ser relative to that in normal control mice (p = 0.17), and a significant increase was observed in PAN mice without Ser treatment (p = 0.0027). Conclusions: Ser interfered with albumin internalization through the caveolae into GEnCs and podocytes and reduced albuminuria. Dynamin inhibitors may serve as a novel therapeutic option for reducing albuminuria in glomerulonephritis.

The green colonial microalga Botryococcus braunii produces large amounts of hydrocarbons and has attracted attention as a potential source of biofuel. When this freshwater microalga is cultured in a brackish medium, the hydrocarbon recovery rate increases; furthermore, the colony size becomes large. In this study, the effects of such changes on the energy balance of harvesting and hydrocarbon recovery were studied via filtrate experiments on an inclined separator and extraction from a concentrated slurry. The inclined separator was effective for harvesting large-colony-forming algae. The water content on the wire screen of slit sizes larger than 150 µm was 85% could be achieved. The input energy of the harvesting using the brackish medium with this separator was ≈44% of that using the freshwater medium with vacuum filtration, while the input energy of the hydrocarbon recovery using the brackish medium was ≈88% of that using the freshwater medium with pre-heating before n-hexane extraction. Furthermore, the energy profit ratio of the process in the brackish medium was 2.92, which was ≈1.2 times higher than that in the freshwater medium. This study demonstrated that filtration techniques and hydrocarbon recovery from B. braunii with a low energy input through culture in a brackish medium are viable.

The expression of ATP‐binding cassette subfamily G member 2 (ABCG2) is related to tumorigenic cancer stem cells (CSC) in several cancers. However, the effects of ABCG2 on CSC‐related malignant characteristics in pancreatic ductal adenocarcinoma (PDAC) are not well elucidated. In this study, we compared the characteristics of low (ABCG2−) and high (ABCG2+)‐ABCG2‐expressing PDAC cells after cell sorting. In adherent culture condition, human PDAC cells, PANC‐1, contained approximately 10% ABCG2+ cell populations, and ABCG2+ cells displayed more and longer microvilli compared with ABCG2− cells. Unexpectedly, ABCG2+ cells did not show significant drug resistance against fluorouracil, gemcitabine and vincristine, and ABCG2− cells exhibited higher sphere formation ability and stemness marker expression than those of ABCG2+ cells. Cell growth and motility was greater in ABCG2− cells compared with ABCG2+ cells. In contrast, epithelial‐mesenchymal transition ability between ABCG2− and ABCG2+ cells was comparable. In 3D culture conditions, spheres derived from ABCG2− cells generated a large number of ABCG2+ cells, and the expression levels of stemness markers in these spheres were higher than spheres from ABCG2+ cells. Furthermore, spheres containing large populations of ABCG2+ cells exhibited high resistance against anti‐cancer drugs presumably depending on ABCG2. ABCG2+ cells in PDAC in adherent culture are not correlated with stemness and malignant behaviors, but ABCG2+ cells derived from ABCG2− cells after sphere formation have stemness characteristics and anti‐cancer drug resistance. These findings suggest that ABCG2− cells generate ABCG2+ cells and the malignant potential of ABCG2+ cells in PDAC varies depending on their environments.

In pancreatic cancer, morphologically and functionally heterogeneous cancer cells reside within the same patient. The heterogeneity is believed to promote metastasis and resistance to chemoradiotherapy. MIA PaCa-2, an established human pancreatic ductal adenocarcinoma (PDAC) cell line, contains round and spindle-shaped adherent cells, as well as, round floating cells. In this study, we aimed to assess if the floating cells might have greater metastatic potential and/or be more resistant to drug-induced apoptosis compared to adherent cells. Time-lapse analysis revealed that the two types of adherent cells transformed bilaterally, and some of the adherent, round cells converted to floating cells. Flow cytometry and electron microscopy showed that approximately 90% of the floating cells were viable. qRT-PCR analysis revealed that floating cells expressed lower levels of integrins and ATP-binding cassette (ABC) transporters than adherent cells. In contrast, except for vimentin, floating cells expressed more epithelial to mesenchymal transition markers than adherent cells. Floating cells included a larger population of G2/M-phase cells, and migration assays revealed a decreased migration ability by floating cells relative to adherent cells. A cell aggregation assay showed that the aggregative properties of the floating cells were lower than those of the adherent cells. In 3D culture, spheres derived from floating cells were more sensitive to anti-cancer drugs, including gemcitabine, 5-FU, and abraxane, than those derived from adherent cells. Expression levels of stemness markers in the spheres derived from floating cells were lower than those derived from adherent cells. Morphological characterization of human PDAC cell lines may help to clarify the series of alterations cancer cells undergo during the metastatic process and may contribute to the development of new PDAC diagnostics and more patient-specific treatments for those with PDAC.

Gangliosides, a group of glycosphingolipids, are known to be cell surface markers and functional factors in several cancers. However, the association between gangliosides and pancreatic ductal adenocarcinoma (PDAC) has not been well elucidated. In this study, we examined the expression and roles of ganglioside GM2 in PDAC. GM2+ cells showed a higher growth rate than GM2− cells in the adherent condition. When GM2– and GM2+ cells were cultured three-dimensionally, almost all cells in the spheres expressed GM2, including cancer stem cell (CSC)-like cells. A glycolipid synthesis inhibitor reduced GM2 expression and TGF-β1 signaling in these CSC-like cells, presumably by inhibiting the interaction between GM2 and TGFβ RII and suppressing invasion. Furthermore, suppression of GM2 expression by MAPK inhibition also reduced TGF-β1 signaling and suppressed invasion. GM2+ cells formed larger subcutaneous tumors at a high incidence in nude mice than did GM2– cells. In PDAC cases, GM2 expression was significantly associated with younger age, larger tumor size, advanced stage and higher histological grade. These findings suggest that GM2 could be used as a novel diagnostic and therapeutic target for PDAC.

Pancreatic cancer, composed of heterogeneous cancer cells, alters epithelial to mesenchymal features during growth and metastasis. In this study, we aimed to characterize pancreatic ductal adenocarcinoma (PDAC) cells showing epithelial or mesenchymal features in 3D culture. In 3D culture, PK-1 cells had high E-cadherin and low vimentin expression and exhibited a round-like appearance encircled by flat cells. PANC-1 cells had high vimentin and low E-cadherin expression and formed grape-like spheres. PK-1 cells had secretary granules and many microvilli, desmosomes, and adherens junctions, while PANC-1 cells had few microvilli, adherens junction, and no desmosomes. Cytokeratin 7, trypsin, CA19-9, and E-cadherin were highly expressed in PK-1 cells but not in PANC-1 cells. Ki-67 was diffusely expressed in PANC-1 spheres but was restricted to the peripheral flat cells of PK-1 spheres. PANC-1 and PK-1 cells were positive for transforming growth factor (TGF) β receptor II and phosphorylated smad2/3, but PK-1 cells were smad4 negative. Taken together, 3D culture enhanced morphofunctional differences of PDAC cells showing epithelial or mesenchymal characteristics, and epithelial phenotype maintenance may be due to the ineffectiveness of the TGF- β pathway. Clarification of heterogeneity using 3D culture may be useful for development of individualized diagnostic and therapeutic methods in patients with PDAC.

Pancreatic ductal adenocarcinoma (PDAC) is a major histological type of pancreatic cancer and remains one of the most lethal cancers with a high mortality rate owing to its aggressive growth, high metastatic rate, and recurrence. Recent studies on cancer stem cells (CSCs) have suggested that the aggressive growth, high metastatic rate, and recurrence might be caused by the ability of CSCs to self-renew, differentiate, and drive tumorigenesis. Thus, CSCs are expected to be a therapeutic target for PDAC. Sphere forming assay of cancer cells, including PDAC cells, is commonly performed using epidermal growth factor and fibroblast growth factor-2 containing serum-free medium to identify and isolate the enriched CSCs. Recently, we observed that PDAC spheres cultured in fetal bovine serum containing medium are morphologically similar to spheres cultured in the growth factor containing medium. In this study, we cultured two PDAC cell lines, PANC-1 and PK-1, in growth factor containing serum-free medium or serum containing medium, and compared the morphology of the spheres formed in detail by electron microscopy and examined the expression of major CSC marker genes. Both the PDAC cells formed larger spheres in the serum containing medium than in the growth factor containing medium. PK-1 cells formed more morphologically differentiated spheres, with peripheral flat lining cells, in the serum containing medium. Expression levels of most of the CSC markers were higher in the spheres of the two PDAC cells in both the culture mediums than in the cells cultured under adherent conditions. The expression levels of CSC markers in PDAC spheres cultured in the growth factor containing medium were not necessarily higher than that in the spheres cultured in the serum containing medium. These findings suggest that sphere forming assay using serum containing medium, by which large PDAC spheres with enriched CSCs are formed, may be useful for deciphering the characteristics of CSCs and for developing anti-CSC therapies for PDAC.

Here, we investigated the effects of culture medium salinity on the hydrocarbon extractability, growth and morphology of Botryococcus braunii strain Showa. At all tested salinities, the hydrocarbon recovery rate increased during the first or second cell division, which occurred within 7 days. Algal growth in modified Chu13 medium did not differ significantly from that in medium with 0.3% seawater salinity (0.3%SM) to which seawater (3.6%, salinity was 36 g L−1) was diluted. In contrast, culturing in 0.3% seawater salinity improved hydrocarbon extractability and shortened the time required for hydrocarbon extraction. This change supposed to be due to the decrease in the production of polysaccharides which prevent non-polar solvent from contacting hydrocarbons. Additionally, there was an increase in the colony sizes and flotation of the alga when cultured in 0.3% seawater salinity. The results of our experiments suggest that culturing B. braunii in a medium with the appropriate salinity can reduce the input energy and the costs incurred from harvest to hydrocarbon extraction without decreasing biomass production.

The green colonial microalga Botryococcus braunii (race B) has attracted considerable attention because it is known to produce many hydrocarbons. The hydrocarbons are present in the extracellular matrix that connects the cells located in the interior of a B. braunii colony; however, the surface of each colony is covered with a retaining wall and a fibrillar colony sheath. This unique colony structure is thought to be associated with hydrocarbon storage and known to affect the extractability of hydrocarbons by using an organic solvent. In this study, we investigated the relation between hydrocarbon extractability and changes in the colony surface structure after two distinct treatments: culture of the algae in brackish medium or thermal pretreatment of the algae before solvent extraction. Both the treatments improved hydrocarbon extractability when non-polar solvents such as n -hexane were used. Thermal pretreatment inhibited the formation of fibrils on the surface of algal colonies, as revealed by rapid-freezing and freeze-substitution electron microscopy. However, when the alga was cultured in brackish medium, the fibrils on the surface shortened and became less dense. Thus, the fibrillar colony sheath that mainly consisted of saccharic components prevented the entry of organic solvents into the colony interior, and thus hydrocarbon extraction.

Jatropha curcas L. is considered a promising next-generation biomass fuel source. Unfortunately, Jatropha seeds contain toxic phorbol esters (PEs) that must be removed from the extracted oil cake for safe use. We used Jatropha oil cake, a byproduct of biodiesel production, and heat-press equipment to evaluate optimum parameters for PE removal while producing a solid fuel with a high energy density. The PE concentration was measured by high-performance liquid chromatography (HPLC), and results showed that the heat-press treatment was effective for PE removal from Jatropha oil cake. Treatment for 15 min at 195 °C under 19.1 MPa pressure produced a solid fuel with PE concentration below the lower detection limit. Our results suggest both treatment temperature and pressure are important factors for PE removal.

(a) Purpose: Gangliosides are molecules composed of glycosphingolipids linked with one or more sialic acids, and are known to be key components of lipid rafts, which act as platforms for signal transduction. Changes in ganglioside levels affect the expression of raft-associated proteins on the cell surface and lead to reduced membrane fluidity, thus, resulting in cellular dysfunction, such as impaired signal transduction. The ganglioside GM2 is one of the major series of gangliosides that has several biological functions, such as in cell adhesion and signal transduction. GM2 is highly expressed in several types of human malignant tumors, such as melanomas, gliomas, and neuroblastomas, but is absent or weakly expressed in normal tissues. However, the association between gangliosides and pancreatic ductal adenocarcinoma (PDAC) has not been elucidated yet. Thus, in this study, we aimed to clarify the expression and role of ganglioside GM2 in PDAC. (b) Experimental procedures: GM2 expression in eight human PDAC cell lines was examined by fluorescence-activated cell sorter (FACS) analysis. The morphology, growth, and stemness of GM2- and GM2+ sorted-cells were compared by transmission electron microscopy, growth assays, real-time PCR analysis of the markers of stemness, and sphere forming assays. Cell motility was evaluated by invasion assay. For comparing tumorigenicity between the GM2- and GM2+ sorted-cells, heterotopic implantations were performed. The expression levels and clinico-pathological roles of GM2 in patients with PDAC were examined by immunohistochemical analysis of 117 pancreatic tissue samples. (c) Results: The fraction of GM2+ cells was the highest (21.4%) in MIA PaCa-2 from among the eight cell lines, as revealed by FACS analysis. GM2-expressing MIA PaCa-2 cells had higher growth rates under adherent growth conditions. In 3D-culture, which enriches cancer stem cells (CSCs), most MIA PaCa-2 cells expressed GM2 and the cells responded to TGF-β1 treatment to promote invasion. Transplantation of the GM2-expressing cells into nude mice resulted in development of larger tumors as compared to that for control cells that did not express GM2. In the PDAC cases, GM2 expression was significantly associated with younger age, larger tumor size, advanced stage, and higher histological grade. (d) Conclusions: We show that a PDAC cell line overexpressing GM2 exhibits high growth rate and high tumor initiation in 2D-culture. Further, pancreatic CSC-like cells expressing GM2 exhibit responsiveness against TGF-β1, resulting in enhanced invasion in 3D-culture. Furthermore, GM2 expression is associated with the growth and advanced stage of human PDAC. Further studies will be required for the development of an early detection method for GM2-overexpressing pancreatic cancers and to develop novel therapeutic strategies targeting GM2. Citation Format: Norihiko Sasaki, Kenichi Hirabayashi, Masaki Michishita, Kimimasa Takahashi, Fumio Hasegawa, Fujiya Gomi, Yoko Itakura, Naoya Nakamura, Masashi Toyoda, Toshiyuki Ishiwata. Increased expression of ganglioside GM2 correlates with aggressiveness of human pancreatic ductal adenocarcinoma [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 135.

Cancer stem cells (CSCs), which are pluripotent and self-renewable, contribute to the initiation and metastasis of cancer, and are responsible for resistance to chemotherapy and radiation. Pancreatic ductal adenocarcinoma (PDAC) is an aggressive type of cancer that is associated with a high incidence of distant metastasis and recurrence. Sphere formation reveals cell proliferation under nonadherent conditions and is commonly used to identify CSCs; measurements of the number, area and volume of the spheres are used to estimate stemness of PDAC cells. However, detailed morphological analysis of such spheres has not been performed. The aim of the present study was to examine the morphology of spheres isolated from PANC-1 human pancreatic cancer cells via scanning electron microscopy (SEM) and transmission electron microscopy (TEM). PANC-1 cells formed round to irregular oblong spheres within 1 week following seeding in ultra-low-attachment plates. These spheres exhibited higher levels of expression of CSC markers, including nestin, sex determining region Y-box 2, and CD44 containing variant exon 9, compared with adherent cells. SEM analysis revealed that the spheres exhibited a grape-like appearance, harboring cancer cells with smooth or rough surfaces. Similarly, TEM analysis detected cancer cells with varying surface types within the spheres: Those with smooth surfaces, irregular large protrusions, protrusions and a small number of microvilli, and those with many microvilli throughout the entire cell surface. These morphological differences among cancer cells may be indicative of different stages in the differentiation process, from CSCs to non-CSCs, within the spheres.

Water-soluble polymers are released from the extracellular matrix of Botryococcus braunii by thermal pretreatment prior to hydrocarbon extraction. In this study, hydrocarbon recovery was reduced by adding polymers to algal slurries from which they were previously washed. The water-soluble polymers are amphiphilic and emulsify the water–organic solvent systems used in hydrocarbon extraction. To obtain > 90% hydrocarbon yields from high-concentration slurry, the original 10% water-soluble polymer content of dry algal cells had to be reduced to less than 0.5%. The water-soluble polymers were polysaccharides with a molecular weight greater than 2 × 10 6 and mainly comprised of galactose, arabinose, and uronic acid. We suggest that high-molecular-weight water-soluble polymers are desirable as industrial emulsifiers and thickeners.

This study aimed to document the material balance and evaluate the energy profit ratios (EPRs) of two processes for recovering hydrocarbon from the green microalgae Botryococcus braunii, which produces hydrocarbon intercellularly, releases it extracellularly, and accumulates it in the extracellular matrix in colony-form. The first is a wet process that elutes an extraction inhibitor through heat, eliminates it through solid–liquid separation, and extracts a hydrocarbon using hexane. The second is a dry process that extracts hydrocarbon by hexane after drying. The EPR of the wet process (3.5) was approximately twice that of the dry process (1.8). Therefore, hydrocarbon can be efficiently recovered from B. braunii without drying.

We simulated III–V nitride heterojunction solar cells with a Mg-doped GaN/intrinsic InGaN/Si-doped GaN structure. We investigated the threshold dislocation density, the carrier lifetime, and the strain in the InGaN active layer. The conversion efficiency was sensitive to carrier lifetime limited by Shockely–Read recombination rather than the dislocation density. The electric field induced by the piezoelectric effect in the InGaN layer on +c polarity GaN was opposite to that in the depletion region. The recombination of photo-generated carriers in the p-GaN/i-InGaN/ n-GaN solar cell structure took place dominantly at the i/n interface even at 0 V, and the performance was remarkably degraded. It was found that this negative influence of the piezoelectric effect was reduced by a heavy doping of more than 3 × 1019 cm−3 into the n+-GaN layer at the interface.

Hydrocarbons are easily extracted by organic sol- vents such as n-decane from wet samples of Botryococcus braunii by thermal pretreatment at 90 °C even after being cooled to room temperature. However, hydrocarbon recover- ies are not as readily achieved at room temperature from samples pretreated at temperatures lower than 80 °C. This suggests that there is the point of no return for pretreatment temperature that enables effective solvent extraction of hydro- carbons at room temperature from wet algal samples of B. braunii. To elucidate the mechanism of hydrocarbon re- covery from B. braunii following thermal pretreatments, we investigated the thermophysical properties of the water phase separated from heated algal slurry. Differential scanning cal- orimetry (DSC) measurements revealed sol-gel transitions in the water phase of algal slurry after protein denaturation at 64 °C in samples that was pretreated at 70 or 80 °C but not in those pretreated at 90 °C. Furthermore, the pretreated >70 °C water-soluble polymers revealed polysaccharides composed of galactose, arabinose, and uronic acid. These results suggest that the transition from sol state to gel state of water-soluble polysaccharides in algal slurry prevented hydrocarbon recov- ery with organic solvents since hydrocarbons were easily recovered from sol state samples pretreated at 70 or 80 °C when the extraction temperature was kept the same as the pretreatment temperature. These results reveal that the pres- ence of water-soluble polymers with gelation ability in the water phase and removal of these polymers in sol state enable effective recovery of hydrocarbons at room temperature after thermal pretreatments.

The hyper-modular printer control project at PARC has proven that a tightly integrated model-based planning and control framework can effectively control a complex physical system. Recently, we have successfully applied this framework to another application: planning for the Material Control System (MCS) of Liquid Crystal Display (LCD) manufacturing plant in a joint project between the Embedded Reasoning Area at PARC and the Products Development Center at the IHI Corporation. The model-based planner created at PARC was able to successfully solve a diverse set of test scenarios provided by IHI, including those that were deemed very difficult by the IHI experts. The short projecttime (2 months) proved that model-based planning is a flexible framework that can adapt quickly to novel applications. In this paper, we will introduce this complex domain and describe the adaptation process of the Plantrol online planner. The main contributions are: (1) introducing a successful application of general-purpose planning; (2) outline the timeline-based online temporal planner; and (3) description of a complex warehouse management problem that can serve as an attractive benchmark domain for planning.

ATP-binding cassette subfamily G member 2 (ABCG2), an important multidrug resistance transporter, can mediate the efflux of various chemotherapy drugs and contribute to drug resistance in cancer cells. The correlation between ABCG2 expression and cancer stem cell (CSC) phenotypes has been examined in hepatocellular carcinoma, as well as in glioma, breast, prostate, and colon cancer; however, the results remain controversial. In this study, we compared the characteristics of low- (ABCG2-) and high-expressing (ABCG2+) pancreatic ductal adenocarcinoma (PDAC) cells using a human pancreatic cancerous cell line (PANC-1) because the role of ABCG2 in CSC-related malignant characteristics in PDAC is not well elucidated. ABCG2- and ABCG2+ PDAC cells were separated using flow cytometric cell sorting. In adherent cell culture conditions, 10% of all PANC-1 cells were ABCG2+. Using transmission electron microscopy, we found that ABCG2+ cells showed more abundant and longer microvilli on the cell surface than ABCG2- cells. Unexpectedly, ABCG2+ cells did not demonstrate significantly greater drug resistance against 5-FU, gemcitabine, and vincristine than ABCG2- cells, as assessed using a WST-8 assay. Furthermore, ABCG2- cells exhibited better sphere formation ability and higher stemness marker expression, including that of Sox2, Oct4, ALDH1, CD44v9, and Nestin, than ABCG2+ cells, as observed using qRT-PCR. Cell growth rates and motilities, examined using the Boyden chamber and scratch assays, were higher in ABCG2- cells than in ABCG2+ cells. However, the epithelial-mesenchymal transition (EMT) ability, assessed by examining the alteration of E-cadherin, N-cadherin, Snail, and Vimentin expression after TGFβ addition, was comparable between ABCG2- and ABCG2+ cells. In 3D-culture conditions using ultra low-attachment plates, ABCG2- cells formed spheres containing a large number of ABCG2+ cells, and expression of stemness markers in these spheres was higher than that of spheres derived from ABCG2+ cells. Furthermore, spheres derived from ABCG2- cells included large populations of ABCG2+ cells and exhibited high resistance against anti-cancer drugs, presumably depending on ABCG2 expression. We found that in adherent culture conditions, ABCG2+ PDAC cells do not exhibit stemness and malignant behaviors, but ABCG2+ cells derived from ABCG2- cells after sphere formation have high stemness and anti-cancer drug resistance. This suggests that ABCG2- cells have the capacity to generate ABCG2+ cells, and the malignant potential of ABCG2+ cells in PDAC depends on the environment. The 3D-culture system was expected to mimic in vivo environments. Therefore, ABCG2+ cells in pancreatic cancer tumors may exhibit high stemness and anti-cancer drug resistance. Thus, ABCG2+ cells should be considered as novel therapeutic targets for pancreatic cancer. Citation Format: Norihiko Sasaki, Fumio Hasegawa, Masaki Michishita, Yoko Matsuda, Tomio Arai, Naoshi Ishikawa, Yoko Itakura, Junko Aida, Kaiyo Takubo, Masashi Toyoda, Toshiyuki Ishiwata. ABCG2-positive cells derived from ABCG2-negative pancreatic cancer cells in 3D-culture conditions show high stemness and anti-cancer drug resistance [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3057.

The crystal quality of AlGaN layers was improved using alternate-source-feeding epitaxy (ASFE) in low-pressure (LP) metal organic vapor phase epitaxy (MOVPE). The green emission of photoluminescence (PL) from the AlGaN clad layer was reduced, indicating that point defects such as Ga or Al vacancy were reduced by ASFE. On the other hand, the full width at half maximum (FWHM) of the X-ray rocking curve was not improved at all and the dislocation density observed by transmission electron microscopy (TEM) was not reduced by ASFE. Furthermore, the yellow emission from GaN/AlGaN multi-quantum well (MQW) grown by conventional-simultaneous source-feeding epitaxy (SSFE) on the ASFE AlGaN clad layer was also reduced. This indicates that the improvement of the crystal quality of the AlGaN clad layer influences the crystal quality of the MQW grown on it.

AlGaN/AlN layers were grown on SiC substrates with an (AlN/GaN) multibuffer-layer structure (MBLS). The tensile strain in the a-axis of the grown layer was controlled by changing the structure of the (AlN/GaN) MBLS. It was found that the crystal quality of the grown layers was improved by increasing the strain. It was also confirmed by transmission electron microscopy (TEM) observation that the dislocation density of the AlN layer decreased with increase of the tensile strain in the a-axis. The screw dislocations decreased more rapidly with the distance from the substrate for the same tensile strain than the edge dislocations did.

AlN and AlGaN epitaxial layers with Al composition of 0.6–0.8 were grown by low-pressure metal organic vapor phase epitaxy on a (0 0 0 1) 6H–SiC substrate using the (AlN/GaN) multi-buffer layer structure (MBLS). Strain of the grown layer could be controlled by structure of the inserted (AlN/GaN) MBLS. It was found that the crystal quality of the grown layer could be improved with increase of the tensile strain in a -axis (compressive strain in c -axis): full-width at half-maximum (FWHM) of X-ray rocking curves (XRC) of both (0 0 0 2) plane ( ω scan) and (1 0 1¯ 2) plane ( ϕ scan) were decreased from 1530 arcsec to 79 arcsec and 3962 arcsec to 853 arcsec for the AlN template, respectively. FWHM of the ϕ scan XRC was several times larger than that of the ω scan, though the former was roughly proportional to the latter.

We recently established a mouse model of peritoneal dissemination of human gastric carcinoma, including the formation of ascites, by orthotopic transplantation of cultured gastric carcinoma cells. To clarify the processes of expansion of the tumors in this model, nude mice were sacrificed and autopsied at different points of time after the orthotopic transplantation of the cancer cells for macroscopic and histopathologic examination of the tumors. The cancer cells grew actively in the gastric submucosa and invaded the deeper layers to reach the serosal plane. The tumor cells then underwent exfoliation and became free followed by the formation of metastatic lesions initially in the greater omentum and subsequent colonization and proliferation of the tumors on the peritoneum. Although this model allowed the detection of even minute metastases, it was not satisfactory from the viewpoint of quantitative and objective evaluation. To resolve these problems, we introduced a luciferase gene into this tumor cell line with a high metastasizing potential and carried out in vivo photon counting analysis. This photon counting technique was found to allow objective and quantitative evaluation of the progression of peritoneal dissemination on a real-time basis. This animal metastatic model is useful for monitoring the responses of tumors to anticancer agents. (Cancer Res 2006; 66(15): 7532-9)

Electroluminescence (EL) properties of Si-based light emitting diodes with β-FeSi 2 particles active region grown by reactive deposition epitaxy are investigated. EL intensity of β-FeSi 2 particles versus excitation current densities has different behaviors at 8, 77 K and room temperature, respectively. The EL peak energy shifted from 0.81 to 0.83 eV at 77 K with the increase of current density from 1 to 70 A/cm 2 . Temperature dependence of the peak energy can be well fitted by semi-empirical Varshni's law with the parameters of α=4.34 e–4 eV/K and β=110 K. These results indicate that the EL emission originates from the band-to-band transition with the band gap energy of 0.824 eV at 0 K.

Optical and electrical properties of β-FeSi2 particles embedded in silicon matrix were improved by growing an un-doped silicon buffer layer at high temperature. With silicon buffer layer, the formation defects were prevented in the silicon matrix during the growth of β-FeSi2 and the subsequent annealing, which results in the narrow photoluminescence spectra. The diodes with silicon buffer layer showed smaller leakage current than those samples without silicon buffer layer. The thermal quenching of electroluminescence from such samples with silicon buffer layer occurred at higher temperature and the intrinsic band gap energy shift between silicon and β-FeSi2 was measured at about 0.23 eV.

Cr-doped GaN films with Cr concentration up to 16% were epitaxially grown on Si(111) substrates by metal organic molecular beam epitaxy (MOMBE) using ammonia or di-methyl-hydrazine (DMHy) as nitrogen source. It was found from a superconducting quantum interference device (SQUID) magnetometer that Cr-doped GaN films with Cr concentration ranging from 1 to 3% showed ferromagnetic behavior. However, no such ferromagnetic behavior was detected by magnetic circular dichroism (MCD) measurements. This result indicates that observed ferromagnetism is attributed to unidentified ferromagnetic precipitates included in the grown layers. (© 2005 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)

Background & Aims: Enhanced motility of cancer cells by remodeling of the actin cytoskeleton seems crucial in the process of cancer invasion and metastasis. We previously identified an actin-binding protein, actinin-4, as a new biomarker of cancer invasion and an indicator of prognosis for patients with breast cancer. However, its involvement in the mechanisms of cancer invasion and metastasis remains undetermined. The current study tested the role of actinin-4 in the motility and metastatic potential of colorectal cancer cells. Methods & Results: Quantitative immunofluorescence histochemistry showed that the expression level of the actinin-4 protein was increased in 73.1% (19/26) of the cases of colorectal cancer over the corresponding normal intestinal epithelium. The increased expression of actinin-4 was most significant in dedifferentiated cancer cells at the invasive front. A colorectal cancer cell clone capable of inducing actinin-4 using the tetracycline-regulatory system (designated DLD1 Tet-off ACTN-4) was established. Upon the induction of actinin-4, DLD1 Tet-off ACTN-4 cells spread filopodia and significantly increased their motility (P = .00027); actinin-4 protein was concentrated at the leading edges of these actin-rich podia. When injected into the mesocecum of severe combined immunodeficient mice, DLD1 Tet-off ACTN4 cells, but not the control cells, metastasized into regional mesenteric lymph nodes, resembling the behavior of clinical cancers. The expression of actinin-4 in focally dedifferentiated cancer cells at the invasive front was significantly correlated with the frequency of lymph node metastasis of colorectal cancer (P = .038). Conclusions: Actinin-4 actively increases cell motility and promotes lymph node metastasis of colorectal cancer.

In this paper, we review the detailed study of epitaxial growth of β-FeSi 2 films by reactive deposition epitaxy (RDE), multilayer technique and molecular beam epitaxy (MBE). The p - and n -type β-FeSi 2 was formed when it was grown under an Fe-rich and an Si-rich condition, respectively. The maximum electron and hole mobilities of the β-FeSi 2 epitaxial films reached 6900 and 13000 cm 2 /V·s for the n - and p -type β-FeSi 2 , respectively, at around 50 K. Room temperature (RT) 1.6 μm electroluminescence (EL) was realized by optimizing the growth conditions for p -Si/β-FeSi 2 particles/ n -Si structures prepared by RDE for β-FeSi 2 and by MBE for Si.

In this paper, we discuss the tendencies of telephone call frequencies in Japan. In our analysis, we developed a new data-visualization method using backward-calculated distances from gravity model. The tendencies of the phone traffic data seem to follow the gravity model as a whole, but if we analyze details, the tendencies are very complicated. The reason might be influences of big cities or regional geographical features. We do not develop a model that explains the general phone traffic structure, but we develop a new method for visualizing the regional structre of phone-traffic. We use the gravity model as a basis, and using the concept of backward-calculated distance. Finally, the multi-dimensional scaling method is applied for visualizing the data. The concept of backward calculated distance was developed by Plane (1984), in analyzing the migration space. Our method is unique because we apply the backward calculated distance into multi dimensional scaling method. We calculated distances from non-constrained gravity model, and it enables us to analyze regional structure in many aspects simply. We analyze Japanese phone traffic data which is between 567 MAs'.

c-axis-oriented Cr-doped hexagonal GaN films with Cr of 0.3, 1 and 5 at.% were epitaxially grown on GaAs(111)A substrates by metal organic molecular beam epitaxy (MOMBE) with metal Ga and dimethylhydrazine (DMHy) as a nitrogen source. Clear hysteresis loop was observed in magnetization versus magnetic filed (M–H) curves for Cr-doped GaN films, and the ferromagnetic behaviour was confirmed even at 350 K. The saturation magnetization per Cr atom is estimated to be approximately 1 Bohr magnetron at 300 K. (© 2003 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)

We developed a video archiving method using MPEG-7 description for both retrieval and editing. When using an MPEG-7 description, only one MPEG-7 instance can cover the area from retrieval to editing. A key frame and a headline can be used for retrieval. With editing, an original time code, taken at the time when recorded or registered, can be used to construct an edit decision list (EDL). This EDL can be used wherever the archived video is available. This paper describes the method and includes examples of an MPEG-7 description of a genre dictionary, a content ID, a key frame, a headline, and an original time code.

In the broadband era, video distribution is being accelerated, and this requires metadata technology for retrieving desired video materials This paper describes a study of a profile for video retrieval based on a layered structure that uses MPEG-7 descriptors. The profiles purpose is to facilitate efficient retrieval of video materials in an archive by means of the internet