Bone marrow-derived dendritic cell progenitors (NLDC 145+, MHC class II+, B7-1dim, B7-2-) induce alloantigen-specific hyporesponsiveness in murine T lymphocytes.

The functional maturation of dendritic cells (DC) and other antigen-presenting cells is believed to reflect the upregulation of cell surface major histocompatibility complex (MHC) class II and other T cell co-stimulatory molecules, especially the CD28 ligands B7-1 (CD80) and B7-2 (CD86). In this study, we propagated cells exhibiting characteristics of DC precursors from the bone marrow (BM) of B10 mice (H-2b; I-A+) in response to granulocyte-macrophage colony stimulating factor (GM-CSF). The methods used were similar to those employed previously to propagate DC progenitors from normal mouse liver. Cells expressing DC lineage markers (NLDC 145+, 33D1+, N418+) harvested from 8-10-day GM-CSF stimulated BM cell cultures were CD45+, heat-stable antigen+, CD54+, CD44+, MHC class II+, B7-1dim but B7-2- (costimulatory molecule-deficient). Supplementation of cultures with interleukin-4 (IL-4) in addition to GM-CSF however, resulted in marked upregulation of MHC class II and B7-2 expression. These latter cells exhibited potent allostimulatory activity in primary mixed leukocyte cultures. In contrast, the cells stimulated with GM-CSF alone were relatively weak stimulators and induced alloantigen-specific hyporesponsiveness in allogeneic T cells (C3H; H-2k; I-E+) detected upon restimulation in secondary MLR. This was associated with blockade of IL-2 production. Reactivity to third-party stimulators was intact. The hyporesponsiveness induced by the GM-CSF stimulated, costimulatory molecule-deficient cells was prevented by incorporation of anti-CD28 monoclonal antibody in the primary MLR and was reversed by addition of IL-2 to restimulated T cells. The findings show that MHC class II+ B7-2- cells with a DC precursor phenotype can induce alloantigen-specific hyporesponsiveness in vitro. Under the appropriate conditions, such costimulatory molecule-deficient cells could contribute to the induction of donor-specific unresponsiveness in vivo

Growth of donor-derived dendritic cells from the bone marrow of murine liver auograft recipients in response to granulocyte/macrophage colony-stimulating factor

Allografts of the liver, which has a comparatively heavy leukocyte content compared with other vascularized organs, are accepted permanently across major histocompatibility complex barriers in many murine strain combinations without immunosuppressive therapy. It has been postulated that this inherent tolerogenicity of the liver may be a consequence of the migration and perpetuation within host lymphoid tissues of potentially tolerogenic donor-derived ("chimeric") leukocytes, in particular, the precursors of chimeric dendritic cells (DC). In this study, we have used granulocyte/macrophage colony-stimulating factor to induce the propagation of progenitors that give rise to DC (CD45+, CDllc+, 33D1+, nonlymphoid dendritic cell 145 +, major histocompatibility complex class II+, B7-1+) in li-tuid cultures of murine bone marrow cells. Using this technique, together with immunocytochemical and molecular methods, we show that, in addition to cells expressing female host (C3H) phenotype (H-2Kk+; I-E+; Y chromosome-), a minor population of male donor (B10)-derived cells (H-2Kb+; I-A+; Y chromosome+) can also be grown in 10-d DC cultures from the bone marrow of liver allograft recipients 14 d after transplant. Highly purified nonlymphoid dendritic cell 145+ DC sorted from these bone marrow-derived cell cultures were shown to comprise ~1-10% cells of donor origin (Y chromosome +) by polymerase chain reaction analysis. In addition, sorted DC stimulated naive, recipient strain T lymphocytes in primary mixed leukocyte cultures. Evidence was also obtained for the growth of donor-derived cells from the spleen but not the thymus. In contrast, donor ceils could not be propagated from the bone marrow or other lymphoid tissues of nonimmunosuppressed C3H mice rejecting cardiac allografrs from the same donor strain (B10). These findings provide a basis for the establishment and perpetuation of cell chimerism after organ transplantation. © 1995, Rockefeller University Press., All rights reserved

CRISPR/Cas9-based editing of a sensitive transcriptional regulatory element to achieve cell type-specific knockdown of the NEMO scaffold protein

The use of alternative promoters for the cell type-specific expression of a given mRNA/protein is a common cell strategy. NEMO is a scaffold protein required for canonical NF-κB signaling. Transcription of the NEMO gene is primarily controlled by two promoters: one (promoter B) drives NEMO transcription in most cell types and the second (promoter D) is largely responsible for NEMO transcription in liver cells. Herein, we have used a CRISPR/Cas9-based approach to disrupt a core sequence element of promoter B, and this genetic editing essentially eliminates expression of NEMO mRNA and protein in 293T human kidney cells. By cell subcloning, we have isolated targeted 293T cell lines that express no detectable NEMO protein, have defined genomic alterations at promoter B, and do not support activation of canonical NF-κB signaling in response to treatment with tumor necrosis factor. Nevertheless, noncanonical NF-κB signaling is intact in these NEMO-deficient cells. Expression of ectopic wildtype NEMO, but not certain human NEMO disease mutants, in the edited cells restores downstream NF-κB signaling in response to tumor necrosis factor. Targeting of the promoter B element does not substantially reduce NEMO expression (from promoter D) in the human SNU423 liver cancer cell line. Thus, we have created a strategy for selectively eliminating cell typespecific expression from an alternative promoter and have generated 293T cell lines with a functional knockout of NEMO. The implications of these findings for further studies and for therapeutic approaches to target canonical NF-κB signaling are discussed.Published versio

Ames collaborative study of cosmic ray neutrons

The results of a collaborative study to define both the neutron flux and the spectrum more precisely and to develop a dosimetry package that can be flown quickly to altitude for solar flare events are described. Instrumentation and analysis techniques were used which were developed to measure accelerator-produced radiation. The instruments were flown in the Ames Research Center high altitude aircraft. Neutron instrumentation consisted of Bonner spheres with both active and passive detector elements, threshold detectors of both prompt-counter and activation-element types, a liquid scintillation spectrometer based on pulse-shape discrimination, and a moderated BF3 counter neutron monitor. In addition, charged particles were measured with a Reuter-Stokes ionization chamber system and dose equivalent with another instrument. Preliminary results from the first series of flights at 12.5 km (41,000 ft) are presented, including estimates of total neutron flux intensity and spectral shape and of the variation of intensity with altitude and geomagnetic latitude

Waste in China

With a world full of inescapable garbage, how much is the emerging country of China producing and should the globe be concerned? As the most populous country in the world, China accounted for over 15% of the earth’s waste volume in 2020. China has the biggest share of waste in the world, but is it enough for other countries to intervene? Higher levels of greenhouse gas in the earth’s atmosphere change its climate and some of these gases are linked to solid waste. These changes can result in more frequent, dangerous natural disasters (floods, hurricanes, mass species extinction, disease, etc.) and could damage communities all over.https://digitalscholarship.unlv.edu/durep_posters/1051/thumbnail.jp

Long‐Distance Natal Dispersal Is Relatively Frequent and Correlated with Environmental Factors in a Widespread Raptor

Dispersal is a critical process influencing population dynamics and responses to global change. Long‐distance dispersal (LDD) can be especially important for gene flow and adaptability, although little is known about the factors influencing LDD because studying large‐scale movements is challenging and LDD tends to be observed less frequently than shorter‐distance dispersal (SDD). We sought to understand patterns of natal dispersal at a large scale, specifically aiming to understand the relative frequency of LDD compared to SDD and correlates of dispersal distances. We used bird banding and encounter data for American kestrels (Falco sparverius) to investigate the effects of sex, migration strategy, population density, weather, year and agricultural land cover on LDD frequency, LDD distance and SDD distance in North America from 1961 to 2015. Nearly half of all natal dispersal (48.9%) was LDD (classified as \u3e30 km), and the likelihood of LDD was positively associated with the proportion of agricultural land cover around natal sites. Correlates of distance differed between LDD and SDD movements. LDD distance was positively correlated with latitude, a proxy for migration strategy, suggesting that migratory individuals disperse farther than residents. Distance of LDD in males was positively associated with maximum summer temperature. We did not find sex‐bias or an effect of population density in LDD distance or frequency. Within SDD, females tended to disperse farther than males, and distance was positively correlated with density. Sampling affected all responses, likely because local studies more frequently capture SDD within study areas. Our findings that LDD occurs at a relatively high frequency and is related to different proximate factors from SDD, including a lack of sex‐bias in LDD, suggest that LDD may be more common than previously reported, and LDD and SDD may be distinct processes rather than two outcomes originating from a single dispersal distribution. To our knowledge, this is the first evidence that LDD and SDD may be separate processes in an avian species, and suggests that environmental change may have different outcomes on the two processes

Strontium potently inhibits mineralisation in bone-forming primary rat osteoblast cultures and reduces numbers of osteoclasts in mouse marrow cultures

The basic mechanisms by which strontium ranelate acts on bone are still unclear. We show that an important action of strontium salts is to block calcification in cultures of osteoblasts, the bone-forming cells. These results suggest that strontium treatment could have previously overlooked effects on bone