Differential radiation sensitivity to morphological, functional and molecular changes of human thyroid tissues and bone marrow cells maintained in SCID mice

Morphology and function of human organs and tissues are well maintained in the improved SCID (severe combined immunodeficient) mice for a long period (approximately 3 years). To study the radiation-induced damage on human thyroid gland, human thyroid tissues transplanted to SCID mice were consecutively exposed to X-rays or 137Cs gamma-rays at high and low dose rates for approximately 2 years. Consecutive irradiation resulted in the disappearance of follicles and significant decrease of thyroid hormone secretion. Mutations in p53 and c-kit genes were induced significantly in human thyroid tissues from old head and neck cancer patients (av. 56.8 years, 4 males) and a Graves' disease patient (20 years, male) over the dose of 24 Gy (44.7+/-5.9 Gy, mean+/-S.E) and 11 Gy (20.2+/-7.8 Gy), respectively, while mutations were not detected at lower doses nor in unexposed matched controls (p < 0.01). There were significant differences in mutation frequency in the transplanted human thyroid tissues (31 years, female) between high dose rate (1.19 Gy/min; 8 in 20 tissues) and low dose rate (0.00023 Gy/min; 0 in 14 tissues) exposures (p < 0.01). Mutations were not detected in RET, K-ras and beta-catenin genes. Expression analysis by GeneChip indicated that gene expression was also well maintained in the transplanted human thyroid tissues. However, lower doses (1 or 3 Gy) of 137Cs gamma-rays can induce changes in gene expression in the transplanted human thyroid tissues. Furthermore, fatally irradiated SCID mice could survive with human bone marrow cell transplantation. When about half of mouse bone marrows were replaced by human bone marrow cells, the human bone marrow cells showed high sensitivity to gamma-irradiation; 28.0% and 0.45% survival after 0.5 and 2.0 Gy exposures, respectively.     

Therapeutic potential and challenges of targeting receptor tyrosine kinase ROR1 with monoclonal antibodies in B-cell malignancies

Background

Based on its selective cell surface expression in chronic lymphocytic leukemia (CLL) and mantle cell lymphoma (MCL), receptor tyrosine kinase ROR1 has recently emerged as a promising target for therapeutic monoclonal antibodies (mAbs). To further assess the suitability of ROR1 for targeted therapy of CLL and MCL, a panel of mAbs was generated and its therapeutic utility was investigated.

Methodology and Principal Findings

A chimeric rabbit/human Fab library was generated from immunized rabbits and selected by phage display. Chimeric rabbit/human Fab and IgG1 were investigated for their capability to bind to human and mouse ROR1, to mediate antibody-dependent cellular cytotoxicity (ADCC), complement-dependent cytotoxicity (CDC), and internalization, and to agonize or antagonize apoptosis using primary CLL cells from untreated patients as well as MCL cell lines. A panel of mAbs demonstrated high affinity and specificity for a diverse set of epitopes that involve all three extracellular domains of ROR1, are accessible on the cell surface, and mediate internalization. The mAb with the highest affinity and slowest rate of internalization was found to be the only mAb that mediated significant, albeit weak, ADCC. None of the mAbs mediated CDC. Alone, they did not enhance or inhibit apoptosis.

Conclusions and Significance

Owing to its relatively low cell surface density, ROR1 may be a preferred target for armed rather than naked mAbs. Provided is a panel of fully sequenced and thoroughly characterized anti-ROR1 mAbs suitable for conversion to antibody-drug conjugates, immunotoxins, chimeric antigen receptors, and other armed mAb entities for preclinical and clinical studies.     

Genome-wide association analysis of seedling root development in maize (Zea mays L.)

Background

Plants rely on the root system for anchorage to the ground and the acquisition and absorption of nutrients critical to sustaining productivity. A genome wide association analysis enables one to analyze allelic diversity of complex traits and identify superior alleles. 384 inbred lines from the Ames panel were genotyped with 681,257 single nucleotide polymorphism markers using Genotyping-by-Sequencing technology and 22 seedling root architecture traits were phenotyped.

Results

Utilizing both a general linear model and mixed linear model, a GWAS study was conducted identifying 268 marker trait associations (p ≤ 5.3×10^-7). Analysis of significant SNP markers for multiple traits showed that several were located within gene models with some SNP markers localized within regions of previously identified root quantitative trait loci. Gene model GRMZM2G153722 located on chromosome 4 contained nine significant markers. This predicted gene is expressed in roots and shoots.

Conclusion

This study identifies putatively associated SNP markers associated with root traits at the seedling stage. Some SNPs were located within or near (<1 kb) gene models. These gene models identify possible candidate genes involved in root development at the seedling stage. These and respective linked or functional markers could be targets for breeders for marker assisted selection of seedling root traits.

Electronic supplementary material

The online version of this article (doi:10.1186/s12864-015-1226-9) contains supplementary material, which is available to authorized users.     

Development and validation of a highly sensitive LC-MS/MS method for organic cation transporter (OCT) substrate tetraethylammonium (TEA) in rabbits

Tetraethylammonium is widely used as a probe in organic cation transporters studies. A simple, highly sensitive, and specific method using direct protein precipitation was developed using Hydrophilic Interaction Liquid Chromatography coupled with positive electrospray ionization tandem mass spectrometry for the determination of tetraethylammonium (TEA) in rabbit plasma. Isocratic separation was achieved using a ZIC-HILIC column with acetonitrile and 5mM ammonium acetate in the ratio of 8:2 containing 0.1% formic acid. Acquisition was performed in multiple reaction monitoring mode with the transitions: m/z 130→100 and 130→86 for TEA and m/z 276.1→142.2 for internal standard (homatropine). This method was validated to determine selectivity, linearity, sensitivity, precision, accuracy, recovery and stability. A good linearity was found within a range of 1.53-784.6 ng/mL. The above method has been demonstrated for its capability to estimate the plasma levels of TEA after its topical instillation in rabbit eyes. This method provides an accurate, precise and sensitive tool for determining TEA levels for transporter studies.     

Bioefficacy of Aristolochia tagala Cham. against Spodoptera litura Fab. (Lepidoptera: Noctuidae)

Bioefficacy of leaf and root extracts of Aristolochia tagala Cham. at different concentrations was evaluated at room temperature against Spodoptera litura Fab. Effects on feeding, larvicidal and pupicidal activities and larval–pupal duration were studied. Higher antifeedant activity (56.06%), lethal concentration for feeding inhibition (3.69%), larvicidal (40.66%), pupicidal (28%), total mortality (68.66%) and prolonged larval–pupal duration (12.04–13.08 days) were observed in ethyl acetate leaf extract at 5.0% concentration. Dose dependant effect of test extracts was observed. This plant could be used to isolate active principles and to develop a new botanical formulation in pest management programmes.     

Inhibition of histone deacetylases is the major pathway mediated by astaxanthin to antagonize LPS‐induced inflammatory responses in mammary epithelial cells

Mastitis is a major inflammatory response of the mammary gland due to various pathogenic invasions and is a serious disease that affects the production yield and health status of cows. Astaxanthin (AST), a xanthophyll carotenoid, is a secondary metabolite synthesized by microalgae and yeasts that has been reported to suppress various inflammatory responses. However, the protective effect of AST on lipopolysaccharide (LPS)‐induced mammary epithelial cells has not yet been reported. The present study results indicated that AST treatment markedly attenuated the oxidative stress markers and nitric oxide (NO) while improving the anti‐oxidant enzymes in LPS exposed cells. On the other hand, LPS‐exposed cells showed nuclear translocation of nuclear factor‐κB (NF‐κB) with the activation of inflammatory cytokines such as monocyte chemoattractant protein‐1, tumor necrosis factor‐α, interferon‐γ, and interleukin‐6 (IL‐6). In addition, mRNA expression analysis revealed that the histone deacetylase (HDAC) ‐1, ‐2, ‐3, ‐6, ‐7 and pentraxin 3 (PTX3) expressions were increased in the LPS group. Furthermore, the activity of HDAC was increased to 2‐fold with a significant reduction in the histone acetyltransferase activity in cells exposed to LPS. However, AST was able to inhibit the nuclear translocation of NF‐κB with attenuated HDAC activity. Intriguingly, HDAC inhibition studies demonstrated that the cytokines such as IL‐4, IL‐8, granulocyte‐mcrophage colony stimulating factor, C‐reactive protein, IL‐17A, and IL‐22 were significantly suppressed which were upregulated in LPS treatment; while AST was found acting by improving the anti‐inflammatory cytokine IL‐10, and thioredoxin reductase levels. Collectively, these findings provide novel insights into the role of HDACs in regulating cellular processes involved in the pathogenesis of LPS‐induced mastitis as well as the potential use of AST as a therapeutic in treatment for controlling disease progression.     

Murine cytomegalovirus infection of mouse testes.

With the aim of illustrating a mechanism of cytomegalovirus (CMV) venereal transmission, we induced murine CMV infection in the mouse testes of immunologically competent mice. Using in situ cytohybridization, we were able to show that murine CMV-specific DNA was associated with spermatocytes and mature sperm. Electron microscopy studies also supported sperm infection. The virus could be reisolated from infected epididymal sperm by cocultivation with mouse embryo fibroblasts. We found no difference in either the sexual performance or the fertilization efficiency of the sperm between infected and uninfected males.