B cells and their immunoglobulin products participate in allograft rejection of transplanted human kidneys in which an interesting feature is the presence of a germinal center like B-cell clusters in the allograft. We report here that the immunoglobulin repertoires of these infiltrating B cells are highly restricted and the B cells within a cluster are clonal. Antibody libraries made from the infiltrating B cells of individual patients unexpectedly revealed that each patient utilizes a particular set of dominant germ line genes as well as dominant complementarity determining region 3. Comparison of kidney and peripheral blood from the same patient showed that the immunoglobulin genes from both compartments had dominant clones, but they differed. The lymphocytes that infiltrate the kidneys express the immunoglobulin gene somatic recombination machinery usually restricted to highly activated lymphocytes in germinal centers and lymphomas. An analogy can be made between the inescapable antigenic drive in chronic infection versus that in an allograft, both of which may lead to emergence of dominant B-cell clones and even lymphoid malignancy.
Aims To develop a multiplex real‐time PCR assay using TaqMan probes for the simultaneous detection and quantification of Tobacco etch virus (TEV), Potato virus Y (PVY) and Tobacco vein banding mosaic virus (TVBMV). Methods and Results Specific primer and probe combinations for TEV and TVBMV were developed from the coat protein region of the viral genome. To detect PVY, a primer and probe combination PVY‐Univ F, PVY‐Univ R and PVY‐Univ P for amplifying the coat protein region of the virus genome was employed. The detection limit of multiplex real‐time PCR for these viruses was 10 copies μl−1 of the standard plasmid. The multiplex reaction was successful in the detection of these three pathogens, with no non‐specific amplification and cross‐reaction. Conclusions This multiplex real‐time PCR provides a rapid, effective, specific and sensitive method for the simultaneous detection and quantification of the three pathogens on infected tobacco plants. Significance and Impact of the Study This multiplex real‐time PCR will be useful not only for diagnostic, ecological, epidemiological and pathogenesis studies, but also for investigating host/virus or virus/virus interactions, in particular during mix infection.
The mechanism of chronic rejection of transplanted human kidneys is unknown. An understanding of this process is important because, chronic rejection ultimately leads to loss of the kidney allograft in most transplants. One feature of chronic rejection is the infiltration of ectopic B-cell clusters that are clonal into the transplanted kidney. We now show that the antibodies produced by these B-cells react strongly with the core carbohydrate region of LPS. Since LPS is a costimulatory immunogen that can react with both the B-cell receptor (BCR) and the Toll-like receptor 4 (TLR4), these results suggest a mechanism for the selective pressure that leads to clonality of these B-cell clusters and opens the possibility that infection and the attendant exposure to LPS plays a role in the chronic rejection of human kidney transplants. If confirmed by clinical studies, these results suggest that treating patients with signs of chronic rejection with antibiotics may improve kidney allograft survival.
Cytoplasmic membrane-associated DNA (cmDNA) is a species of DNA that attaches to the plasma membrane and has physical and chemical properties that differ from those of bulk chromosomal and mitochondrial DNAs. Here, we used deep sequencing to analyze cmDNA and showed that satellite DNAs consisting of both of simple (CCATT) N repeats from the pericentromere regions of the chromosomes and 171-bp α-satellite repeat sequences from centromeres were highly enriched. Importantly, we found there is a special cytoplasmic membrane-associated transcription system in which DNA-dependent RNA polymerase II, which colocalizes with template cmDNA at the plasma membrane, can transcribe the membrane-associated 171-bp α-satellite repeat sequences into RNA. Analysis of phosphorylation patterns indicated that the RNA polymerase II in the plasma membrane is in a different chemical state from its nuclear counterpart.nongenomic gene expression | extranuclear coding D NA in eukaryotic organisms generally is considered to be confined to the nuclei or mitochondria of cells. However, in the early 1970s we described a species of cytoplasmic, membrane-associated DNA (cmDNA) in continuously growing lymphocytes that was associated with the plasma membrane (1, 2). Electron microscopic studies of plasma membrane fractions showed that every membrane fragment in the preparation was associated with a linear DNA fragment that appeared to make an end-on attachment to the membrane (1). This DNA appeared to originate from the nucleus, but density-labeling studies suggested that its replication was not synchronous with the rest of the genome and that after replication it exited the nucleus and became attached to cytoplasmic membranes. A study of its reassociation kinetics using the then widely used Cot 1/2 curves showed that about 73% of the cytoplasmic DNA had a rapidly reassociating component with a Cot 1/2 of about 5.0 × 10 −3 and, in general, had a reannealing pattern that differed from bulk nuclear DNA (3-6). Since these early findings, there have been many studies of this DNA that can essentially be divided into three eras that reflect the thinking and available methodology of the times (7-10). After its initial discovery, many thought that cmDNA might be a simple contamination from nuclear DNA or some infectious agent such as a virus or mycoplasma, although none of these explanations was consistent with the analytical studies done at the time of its discovery. Then, in 1975, Saunders and coworkers (7) performed a key experiment in which they demonstrated by in situ hybridization that this DNA hybridized to the heterochromatic regions of chromosomes. These in situ studies were confirmed and extended much later by Kanda and co-workers (8), who demonstrated that cytoplasmic DNA contained α-satellite sequences that hybridized to the nucleus and the cytoplasm of cultured human cells as well as to freshly prepared bone marrow smears. This latter experiment was important because it showed that cytoplasmic DNA was present in fresh bone marrow that had not...
Tobacco viruses may cause a wide range of diseases that heavily reduce tobacco quality and yield worldwide. In order to detect viral diseases in tobacco fields, a one-step reverse transcription loop-mediated isothermal amplification (RT-LAMP) method was established. Nucleotide amplification could be observed clearly after adding SYBR Green I, within 60 min under isothermal conditions, at 63-65 °C with a set of primers targeting the viral coat protein (CP) genes of tobacco viruses including cucumber mosaic virus (CMV), potato virus Y (PVY), tobacco etch virus (TEV), tobacco mosaic virus (TMV) and tobacco vein banding mosaic virus (TVBMV). This method has high specificity and sensitivity. The sensitivity of the RT-LAMP was 10 to 100 times higher than that of the conventional RT-PCR method. The RT-LAMP assay was proven reliable for virus diagnosis of tobacco samples from the field.
A combination of bi-specific antibodies (BsAb), anti-tumorxanti-CD3 and anti-tumorxanti-CD28, is effective in vitro and in vivo, whereas production of two kinds of bi-specific antibodies is labor intensive and administration is complicated. Accordingly, we previously developed a new model of single chain tri-specific antibody (scTsAb), sTRI, which linked both the CD3 and CD28 signals for T-cell activation in one molecule, and demonstrated its capacity for triggering T-cells to kill ovary tumor cells. To improve the pharmacokinetics further and decrease the immunogenicity of scTsAb, we have now generated a new format of scTsAb, TR3H, whose molecular size is smaller than sTRI. Here we describe the construction, purification and characterization of TR3H. TR3H scTsAb bound to effector cells and tumor target cells specifically and induced redirected lyses of ovary tumor cells through freshly isolated, unstimulated human peripheral blood lymphocytes (PBLs). This new format of scTsAb possesses properties that support its potential as a new tumor immunotherapeutic agent.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.