High efficiency Hall effect micro-biosensor platform for detection of magnetically labeled biomolecules

Detection of magnetically labeled biomolecules using micro-Hall biosensors is a promising method for monitoring biomolecular recognition processes. The measurement efficiency of standard systems is limited by the time taken for magnetic beads to reach the sensing area of the Hall devices. Here, micro-current lines were integrated with Hall effect structures to manipulate the position of magnetic beads via field gradients generated by localized currents flowing in the current lines. Beads were accumulated onto the sensor surface within seconds of passing currents through the current lines. Real-time detection of magnetic beads using current lines integrated with Hall biosensors was achieved. These results are promising in establishing Hall biosensor platforms as efficient and inexpensive means of monitoring biomolecular reactions for medical applications.     

System A amino acid transporter SNAT2 shows subtype-specific affinity for betaine and hyperosmotic inducibility in placental trophoblasts

Betaine uptake is induced by hypertonic stress in a placental trophoblast cell line, and involvement of amino acid transport system A was proposed. Here, we aimed to identify the subtype(s) of system A that mediates hypertonicity-induced betaine uptake. Measurement of [¹⁴C]betaine uptake by HEK293 cells transiently transfected with human or rat sodium-coupled neutral amino acid transporters (SNATs), SNAT1, SNAT2 and SNAT4 revealed that only human and rat SNAT2 have betaine uptake activity. The Michaelis constants (K_m) of betaine uptake by human and rat SNAT2 were estimated to be 5.3 mM and 4.6 mM, respectively. Betaine exclusively inhibited the uptake activity of SNAT2 among the rat system A subtypes. We found that rat SNAT1, SNAT2 and SNAT4 were expressed at the mRNA level under isotonic conditions, while expression of SNAT2 and SNAT4 was induced by hypertonicity in TR-TBT 18d-1 cells. Western blot analyses revealed that SNAT2 expression on plasma membrane of TR-TBT 18d-1 cells was more potently induced by hypertonicity than that in total cell lysate. Immunocytochemistry confirmed the induction of SNAT2 expression in TR-TBT 18d-1 cells exposed to hypertonic conditions and indicated that SNAT2 was localized on the plasma membrane in these cells. Our results indicate that SNAT2 transports betaine, and that tonicity-sensitive SNAT2 expression may be involved in regulation of betaine concentration in placental trophoblasts.     

Strain field in actin filament network in lamellipodia of migrating cells: Implication for network reorganization

Cell motility is spatiotemporally regulated by interactions among mechanical and biochemical factors involved in the regulation of cytoskeletal actin structure reorganization. Although the molecular mechanisms underlying cell motility have been well investigated, the contributions of mechanical factors such as strain in the network reorganization remain unclear. In this study, we have quantitatively evaluated the strain field in the actin filament network forming the lamellipodia of migrating fish keratocytes to elucidate the mechanism by which actin filament network reorganization is regulated by biomechanical factors. The results highlight the existence of a negative (compressive) strain in the lamellipodia whose direction is parallel to that of cell movement. A close correlation was found between the distributions of the strain and the actin filament density in the lamellipodia, suggesting that negative strain may be involved in filament depolymerization. Based on this result, we propose a selective depolymerization model which suggests that negative strain may couple with biomechanical factors such as ADF/cofilin to promote selective depolymerization of filaments oriented in the direction of the deformation because such filaments experience relatively higher levels of the deformation. This model, in conjunction with others, may explain the observed reduction in filament density and the reorganization of actin filament network at the back of the lamellipodia of migrating fish keratocytes. Thus, we suggest that by coupling with biochemical factors, mechanical factors are involved in the regulation of actin filament depolymerization, thereby contributing to the regulation of cell motility.     

Culture of human mast cells from peripheral blood progenitors

This protocol details a method for obtaining a considerable number of human mast cells from peripheral blood cells from normal donors without using stem cell mobilization treatment. By using the magnetic cell sorting system, 10(4)-10(5) cells are retrieved in the CD34+ fraction when 100 ml of blood is drawn from a healthy donor. When these cells are cultured using methylcellulose medium supplemented with stem cell factor and interleukin 6, 10(3)-10(4) mast cell colonies are formed in 6 weeks. The total mast cell number at 6 weeks of culture will be 10(6)-10(7).     

Four TFL1/CEN-Like Genes on Distinct Linkage Groups Show Different Expression Patterns to Regulate Vegetative and Reproductive Development in Apple (Malusxdomestica Borkh.)

Recent molecular analyses in several plant species revealedthat TERMINAL FLOWER1 (TFL1) and CENTRORADIALIS (CEN) homologsare involved in regulating the flowering time and/or maintainingthe inflorescence meristem. In apple (Malusxdomestica Borkh.),four TFL1/CEN-like genes, MdTFL1, MdTFL1a, MdCENa and MdCENb,were found and mapped by a similar position on putatively homoeologouslinkage groups. Apple TFL1/CEN-like genes functioned equivalentlyto TFL1 when expressed constitutively in transgenic Arabidopsisplants, suggesting that they have a potential to complementthe TFL1 function. Because MdTFL1 and MdTFL1a were expressedin the vegetative tissues in both the adult and juvenile phases,they could function redundantly as a flowering repressor anda regulator of vegetative meristem identity. On the other hand,MdCENa was mainly expressed in fruit receptacles, cultured tissuesand roots, suggesting that it is involved in the developmentof proliferating tissues but not in the control of the transitionfrom the juvenile to the adult phase. In contrast, MdCENb wassilenced in most organs probably due to gene duplication bythe polyploid origin of apple. The expression patterns of MdTFL1and MdCENa in apple were also supported by the heterologousexpression of β-glucuronidase fused with their promoterregions in transgenic Arabidopsis. Our results suggest thatfunctional divergence of the roles in the regulation of vegetativemeristem identity may have occurred among four TFL1/CEN-likegenes during evolution in apple.     

Mast cell death induced by 24(S),25-epoxycholesterol

Mast cell is one of the central effectors in inflammatory responses. Recent studies suggest that a promising therapeutic approach for various inflammatory immune diseases, including rheumatoid arthritis, multiple sclerosis, and type I allergies, is to inhibit mast cell growth and/or survival. Studies also indicate that a balanced lipid metabolism is crucial for regulating the life span of cells. Oxysterol is a well-known regulator of lipid metabolism and has diverse functions, such as inhibition of the mevalonate isoprenoid pathway, efflux of free cholesterols, and synthesis of cholesterol esters. Here, we show that 24(S),25-epoxycholesterol, a representative endogenous oxysterol, induces apoptosis in bone marrow-derived murine mast cells. Furthermore, we have revealed, for the first time, that the accumulation of neutral lipids catalyzed by acyl-CoA:cholesterol acyltransferase in the cells was involved in induction of mast cell apoptosis. Our present findings confer new insights into the roles of lipid metabolism during oxysterol-mediated mast cell apoptosis.     

中国的肉座菌科Ⅲ.肉座菌属的菌生种

对肉座菌属的系统分类史进行了回顾,并对3个菌生种进行了报道;其中硫磺肉座菌(新拟)Hypocrea sulphurea(Schwein.)Sacc.为中国新记录种,拟层孔肉座菌H.fomitopsisP.G.Liuet     

Photosynthesis and water-relation traits of the summer annual C4 grasses, Eleusine indica and Digitaria adscendens, with contrasting trampling tolerance

Two summer annual C₄ grasses with different trampling susceptibilities were grown as potted plants, and diurnal leaf gas exchange and leaf water potential in each grass were compared. The maximum net photosynthetic rate, leaf conductance and transpiration rate were higher in the trampling-tolerant Eleusine indica (L.) Gaertn. than in trampling sensitive Digitaria adscendens (H. B. K.) Henr. Leaf water potential was much lower in E. indica than in D. adscendens. There were no differences in soil-to-leaf hydraulic conductance and leaf osmotic potential at full turgor as obtained by pressure–volume analysis. However, the bulk modulus of elasticity in cell walls was higher in E. indica leaves than in D. adscendens leaves. This shows that the leaves of E. indica are less elastic. Therefore, the rigid cell walls of E. indica leaves reduced leaf water potential rapidly by decreasing the leaf water content, supporting a high transpiration rate with high leaf conductance. In trampled habitats, such lowering of leaf water potential in E. indica might play a role in water absorption from the compacted soil. In contrast, the ability of D. adscendens to colonize dry habitats such as coastal sand dunes appears to be due to its lower transpiration rate and its higher leaf water potential which is not strongly affected by decreasing leaf water content.     

Angiogenesis after administration of basic fibroblast growth factor induces proliferation and differentiation of mesenchymal stem cells in elastic perichondrium in an in vivo model: mini review of three sequential republication-abridged reports

To date, studies on mesenchymal tissue stem cells (MSCs) in the perichondrium have focused on in vitro analysis, and the dynamics of cartilage regeneration from the perichondrium in vivo remain largely unknown. We have attempted to apply cell and tissue engineering methodology for ear reconstruction using cultured chondrocytes. We hypothesized that by inducing angiogenesis with basic fibroblast growth factor (bFGF), MSCs or cartilage precursor cells would proliferate and differentiate into cartilage in vivo and that the regenerated cartilage would maintain its morphology over an extended period. As a result of a single administration of bFGF to the perichondrium, cartilage tissue formed and proliferated while maintaining its morphology for at least 3 months. By day 3 post bFGF treatment, inflammatory cells, primarily comprising mononuclear cells, migrated to the perichondrial region, and the proliferation of matrix metalloproteinase 1 positive cells peaked. During week 1, the perichondrium thickened and proliferation of vascular endothelial cells was noted, along with an increase in the number of CD44-positive and CD90-positive cartilage MSCs/progenitor cells. Neocartilage was formed after 2 weeks, and hypertrophied mature cartilage was formed and maintained after 3 months. Proliferation of the perichondrium and cartilage was bFGF concentration-dependent and was inhibited by neutralizing antibodies. Angiogenesis induction by bFGF was blocked by the administration of an angiogenesis inhibitor, preventing perichondrium proliferation and neocartilage formation. These results suggested that angiogenesis may be important for the induction and differentiation of MSCs/cartilage precursor cells in vivo, and that morphological changes, once occurring, are maintained.