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At least 1,243 records · Page 69

Visualization of in-situ chemical flow through sand using neutron radiography

Chemical movement through soil is an important process in agriculture and ecology. Observing the spatial and temporal dynamics of these processes using conventional chemical ecology methods requires techniques that are destructive and/or lack resolution. Neutron radiography has the capability to allow chemical motion through sand/soil to be tracked with high spatial and temporal resolution, and we show that it allows for the motion of hydrophobic and hydrophilic chemicals to be distinguished. This technique can have an important impact on introducing neutron radiography to a wider community and into our understanding of chemical communication dynamics between plants and movement of applied chemicals in agricultural soils.

MORRIS, KATHRYN [Xavier University]↗

Visualization of the human CD4{sup +} T-cell response in humanized HLA-DR4-expressing NOD/Shi-scid/γc{sup null} (NOG) mice by retrogenic expression of the human TCR gene

Highlights: • β-Lactoglobulin (BLG) specific TCR genes were introduced to human HSC by retrovirus. • Human HSC with BLG-specific TCR were transplanted into NOG-HLA-DR4 I-A{sup −/−} mice. • BLG-specific TCR induced positive selection of thymocytes. • BLG-specific TCR positive CD4{sup +} T cells mediated immune responses in humanized mice. - Abstract: The development of severe immunodeficient mouse strains containing various human genes, including cytokines or HLA, has enabled the reconstitution of functional human immune systems after transplantation of human hematopoietic stem cells (HSC). Accumulating evidence has suggested that HLA-restricted antigen-specific human T-cell responses can be generated in these humanized mice. To directly monitor immune responses of human CD4{sup +} T cells, we introduced β-lactoglobulin (BLG)-specific T cell receptor (TCR) genes derived from CD4{sup +} T-cell clones of cow-milk allergy patients into HSCs, and subsequently transplanted them into NOG-HLA-DR4 transgenic/I-Aβ deficient mice (NOG-DR4/I-A{sup o}). In the thymus, thymocytes with BLG-specific TCR preferentially differentiated into CD4{sup +}CD8{sup −} single-positive cells. Adoptive transfer of mature CD4{sup +} T cells expressing the TCR into recipient NOG-DR4/I-A{sup o} mice demonstrated that human CD4{sup +} T cells proliferated in response to antigenic stimulation and produced IFN-γ in vivo, suggesting that functional T-cell reactions (especially Th1-skewed responses) were induced in humanized mice.

60 APPLIED LIFE SCIENCES↗

Revelation of the dynamic progression of hypoxia-reoxygenation injury by visualization of the lysosomal hydrogen peroxide

Hydrogen peroxide (H{sub 2}O{sub 2}) plays an important role in pathological conditions, such as cerebral ischemia-reperfusion (I-R) injury. Fluorescent probes may serve as valuable tools to detect the amount, temporal and spatial distribution of H{sub 2}O{sub 2} in living cells. To investigate the role of lysosomal H{sub 2}O{sub 2} involved in cerebral I-R injury, we designed and synthesized a lysosome-targetable two-photon fluorescent probe ztl-4, through expansion and substitution of the original pyridazinone scaffold, conjugation of electronic-donating aromatic ring and precise terminal modification of the alkyl linker. The probe ztl-4 exhibited fast, sensitive and highly selective response toward H{sub 2}O{sub 2}. ztl-4 could image exogenous H{sub 2}O{sub 2} in SH-SY5Y cells and brain slices. In addition, ztl-4 was located in lysosomes with high colocalization coefficient compared with LysoTracker. ztl-4 was further applied for detecting the endogenous generation of H{sub 2}O{sub 2} in SH-SY5Y cells subjected to oxygen and glucose deprivation (OGD) or OGD/reoxygenation (OGD/R) injury. Both OGD- and OGD/R-induced cell injury caused a time-dependent increase of H{sub 2}O{sub 2} production within lysosomes. Moreover, OGD/R-treated cells showed much more amount of H{sub 2}O{sub 2} than OGD-treated cells, indicating that reoxygenation will promote H{sub 2}O{sub 2} accumulation in lysosomes of post-hypoxia cells. Therefore, the probe is suitable for monitoring the dynamic changes of lysosomal H{sub 2}O{sub 2} in cells. - Highlights: • New fluorescent probe displays high selectivity for H{sub 2}O{sub 2}. • The probe is lysosome-targetable. • The probe can real-time monitor hypoxia/reoxygenation injury-induced generation of H{sub 2}O{sub 2} in lysosomes of cells.

60 APPLIED LIFE SCIENCES↗

Visualization of specific collagen-producing cells by Col1-GFP transgenic mice revealed novel type I collagen-producing cells other than fibroblasts in systemic organs/tissues

Highlights: • A new reporter mouse was generated to label type I collagen-producing cells. • Type I collagen-producing cells were specifically labeled in this reporter mouse. • Podocytes and Leydig cells showed the ability to produce type I collagen. Type I collagen is one of the most abundant proteins in mammals and plays important roles in maintaining the integrity of many tissues. Although fibroblasts are the main source of type I collagen, other cells also produce it; however, these cells are not well-defined owing to the lack of a specific marker. A transgenic (Tg) mouse line has been generated in which type I collagen-producing cells are labeled with enhanced green fluorescent protein (EGFP), which enables the monitoring of these cells without requiring an additional cell marker. This Tg mouse line has since been widely used to study type I collagen-producing cells and fibrosis; one study revealed that podocytes, which were not previously considered to produce type I collagen, expressed EGFP. This raises a question regarding the specificity of EGFP expression in this Tg mouse line. To exclude the possibility of non-specific EGFP expression in the existing Tg mouse line and specifically monitor type I collagen-producing cells, we generated a new Tg mouse line and histologically confirmed the specificity of EGFP expression throughout the body. Moreover, we explored type I collagen-producing cells other than fibroblasts and revealed for the first time that Leydig cells have the ability to produce type I collagen. Because of its highly specific and physiologically accurate expression, our new Tg mouse line will help to accurately elucidate not only type I collagen-producing cells in normal tissues but also the potential cells in fibrotic tissues, providing new insights into the pathology of fibrosis.

60 APPLIED LIFE SCIENCES↗