Burn Injury Reduces Neutrophil Directional Migration Speed in Microfluidic Devices

Thermal injury triggers a fulminant inflammatory cascade that heralds shock, end-organ failure, and ultimately sepsis and death. Emerging evidence points to a critical role for the innate immune system, and several studies had documented concurrent impairment in neutrophil chemotaxis with these post-burn inflammatory changes. While a few studies suggest that a link between neutrophil motility and patient mortality might exist, so far, cumbersome assays have prohibited exploration of the prognostic and diagnostic significance of chemotaxis after burn injury. To address this need, we developed a microfluidic device that is simple to operate and allows for precise and robust measurements of chemotaxis speed and persistence characteristics at single-cell resolution. Using this assay, we established a reference set of migration speed values for neutrophils from healthy subjects. Comparisons with samples from burn patients revealed impaired directional migration speed starting as early as 24 hours after burn injury, reaching a minimum at 72–120 hours, correlated to the size of the burn injury and potentially serving as an early indicator for concurrent infections. Further characterization of neutrophil chemotaxis using this new assay may have important diagnostic implications not only for burn patients but also for patients afflicted by other diseases that compromise neutrophil functions.     

The Invisibility of Disability: Using Dance to Shake from Bioethics the Idea of ‘Broken Bodies’

Complex social and ethical problems are often most effectively solved by engaging them at the messy and uncomfortable intersections of disciplines and practices, a notion that grounds the InVisible Difference project, which seeks to extend thinking and alter practice around the making, status, ownership, and value of work by contemporary dance choreographers by examining choreographic work through the lenses of law, bioethics, dance scholarship, and the practice of dance by differently‐abled dancers. This article offers a critical thesis on how bioethics has come to occupy a marginal and marginalizing role in questions about the differently‐abled body. In doing so, it has rendered the disabled community largely invisible to and in bioethics. It then defends the claim that bioethics – as a social undertaking pursued collaboratively by individuals from different disciplines – must take much better notice of the body and the embodied individual if it is to better achieve its ends, which include constructing a moral and just society. Finally, this article considers how the arts, and specifically dance (and here dance by differently‐abled dancers), provides us with rich evidence about the body and our ability to respond positively to normally ‘othered’ bodies. It concludes that greater attention to empirical evidence like that being generated in InVisible Difference will help to expand the reach and significance of bioethics, and thereby its relevance to (and consciousness of) important questions about the status of bodies and bodily differences, which must be considered as central to its ambitions.     

Aggregation of macrophages and fibroblasts is inhibited by a monoclonal antibody to the hyaluronate receptor

To examine the role of the hyaluronate receptor in cell to cell adhesion, we have employed the K-3 monoclonal antibody (MAb) which specifically binds to the hyaluronate receptor and blocks its ability to interact with hyaluronate. In the first set of experiments, we investigated the spontaneous aggregation of SV-3T3 cells, which involves two distinct mechanisms, one of which is dependent upon the presence of divalent cation and the other is independent. The divalent cation-independent aggregation was found to be completely inhibited by both intact and Fab fragments of the K-3 MAb. In contrast, the K-3 MAb had no effect on the divalent cation-dependent aggregation of cells. In a second set of experiments, we examined alveolar macrophages. The presence of hyaluronate receptors on alveolar macrophages was demonstrated by the fact that detergent extracts of these cells could bind [3H]hyaluronate, and this binding was blocked by the K-3 MAb. Immunoblot analysis of alveolar macrophages showed that the hyaluronate receptor had a Mr of 99,500, which is considerably larger than the 85,000 Mr for that on BHK cells. When hyaluronate was added to suspensions of alveolar macrophages, the cells were induced to aggregate. This effect was inhibited by the K-3 MAb, suggesting that the hyaluronate-induced aggregation was mediated by the receptor.     

The complete amino acid sequences of cytosolic and mitochondrial aspartate aminotransferases from horse heart,and inferences on evolution of the isoenzymes

Summary We report here the complete amino acid sequences of the cytosolic and mitochondrial aspartate aminotransferases from horse heart. The two sequences can be aligned so that 48.1% of the amino acid residues are identical. The sequences have been compared with those of the cytosolic isoenzymes from pig and chicken, the mitochondrial isoenzymes from pig, chicken, rat, and human, and the enzyme fromEscherichia coli. The results suggest that the mammalian cytosolic and mitochondrial isoenzymes have evolved at equal and constant rates whereas the isoenzymes from chicken may have evolved somewhat more slowly. Based on the rate of evolution of the mammalian isoenzymes, the geneduplication event that gave rise to cytosolic and mitochondrial aspartate aminotransferases is estimated to have occurred at least 10⁹ years ago. The cytosolic and mitochondrial isoenzymes are equally related to the enzyme fromE. coli; the prokaryotic and eukaryotic enzymes diverged from one another at least 1.3×10⁹ years ago.     

Mass characteristics of DNA obtained from chromosomes of a human carcinoma cell line

Sedimentation studies of DNA from chromosomes extracted from human mitotic cells showed that highmolecular-weight DNA can be obtained if cell hypotonic treatments and prolonged metaphase blocks are avoided. Two types of large double-stranded DNA were observed. One of these (M _r = 2.5×10⁸) appeared as a size class with characteristics reminiscent of the chromosomal DNA subunit hypothesis. However, this DNA is the decay product of larger molecules, whose minimum molecular weight is 6×10⁸.     

How do retroviral oncogenes induce transformation in avian erythroid cells?

The v-erb B oncogene, as well as other oncogenes of the src-gene family transform immature erythroid cells from chick bone marrow in vivo and in vitro. The erb B-transformed erythroid cells differ from normal late erythroid precursors (CFU-E) in that they have acquired the capacity to undergo self-renewal as well as to differentiate terminally. They also do not require the normal erythroid differentiation hormone, erythropoietin, for either process. Cooperation of v-erb B with a second oncogene, v-erb A, results in a differentiation arrest of the transformed cells, which now only use the self-renewal pathway. Studies with conditional and non-conditional mutants in both v-erb B and v-erb A will be presented to elucidate further how the transforming proteins encoded by these oncogenes, gp74erb B and gp75gag-erb A, affect the differentiation programme of the infected erythroid precursor with the outcome of hormone-independent leukaemic cells arrested at an early stage of erythroid differentiation.     

Temperature-sensitive mutants of MH2 avian leukemia virus that map in the v-mil and the v-myc oncogene respectively.

MH2 is an avian retrovirus that contains the v-mil and v-myc oncogenes. In vitro it transforms chick macrophages that are capable of proliferation in the absence of growth factor. Earlier work showed that v-myc induces macrophage transformation and that v-mil induces the production of chicken myelomonocytic growth factor (cMGF), thus generating an autocrine system. We describe the isolation of temperature-sensitive (ts) mutants of MH2 virus. As suggested by marker rescue experiments, one mutant bears a ts lesion in v-mil, whereas the other carries a mutation in v-myc. Ts v-mil MH2-transformed macrophages become factor-dependent at the non-permissive temperature (42 degrees C), while ts-v-myc MH2-transformed macrophages cease growing and acquire a more normal macrophage phenotype at 42 degrees C irrespective of the presence of cMGF. Both phenotypes can be reversed by backshift to the permissive temperature. These results suggest that the gene products of v-mil and v-myc function independently of each other and that v-mil is necessary for the maintenance of autocrine growth, whereas v-myc is required to maintain the transformed phenotype.     

Human HepG2 and Rat Fao Hepatic-Derived Cell Lines Show Different Responses to Ciprofibrate, a Peroxisome Proliferator: Analysis by Flow Cytometry

Peroxisome proliferators, and especially hypolipidemic drugs such as ciprofibrate, are known to be hepatocarcinogens in rodents, but their effect in humans is controversial. In an attempt to investigate the effects of ciprofibrate at a cellular level, the analysis of individual whole cells was performed by flow cytometry on samples from two hepatic-derived cell lines: the rat Fao cell line and the human HepG2 cell line. The increase of light scatter signals in rat Fao cells treated for 3 days with ciprofibrate at 250 μMwas related to modifications of intrinsic cellular parameters, such as size and cytoplasmic granularity. Conversely, no variations appeared in human HepG2-treated cells. Moreover, the study of the cell cycle distribution of asynchronously growing cells showed an increase in the percentage of proliferative cells in Fao-treated cells, but not in HepG2-treated cells. In order to give a simultaneous assessment of changes in cellular parameters and cell metabolism, these flow cytometric experiments were completed with the measurements of the palmitoyl–CoA oxidase activity, used as a marker of peroxisome proliferation. The cellular modifications in the rat Fao cell line were accompanied by a great increase in this enzymatic activity, whereas the human HepG2 cell line, which failed to exhibit changes of cytometric data, presented no, or weak, increase in this oxidase activity. The cellular modifications observed in the rat Fao cell line may be related to the well-known hepatocarcinogenicity of ciprofibrate in rodents, whereas the absence of response of HepG2 cells is in favor of the noncarcinogenicity of this drug in humans. This report validates another methodological approach for the investigation of the safety of peroxisome proliferators in humans.