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.     

Characterisation of stroma-dependent blast colony-forming cells in human marrow

Human bone marrow contains a population of haemopoietic progenitor cells that can be distinguished by their ability to adhere to preformed stromal layers (cultured in the presence of methylprednisolone [MP+] and form blast cell colonies. The stromal layers function in the colony assay after they have been heavily irradiated but not after they have been passaged. The binding of the progenitor cells to the stromal cells is complete after 2 hours of coincubation, and stromal layers of 9.6 cm2 can provide adhesion sites for at least 2,000 blast colony-forming cells. The blast colony-forming cells were shown by micromanipulation to self-renew as well as to give rise to multipotential and lineage-committed colony-forming progenitor cells.     

Polymorphisms of DQ β genes in HLA-DR4 haplotypes from healthy and diabetic individuals

The restriction fragment length polymorphism (RFLP) of DQ was assessed in a panel of control and insulin-dependent diabetes (IDD) patients who were serologically typed as HLA-DR4 homozygotes or HLA-DR3, DR4 heterozygotes. Digestions of genomic DNA with Barn HI, Bg1 II, Pst I, Xba I, and Hind III revealed a total of 15 RFLPs in the panel of 71 HLA-DR4 chromosomes. These RFLPs were organized into six allelic groups on the basis of segregation analysis in families. Complete RFLP haplotypes for the 5 restriction enzymes could be constructed for 42 of the HLA-DR4 chromosomes. This analysis revealed 18 RFLP haplotypes of DQ associated with the DR4 chromosomes tested. Two of these haplotypes, designated DQ3.DR4.a and DQ3.DR4.b, accounted for over 50 % of the DR4 chromosomes analyzed. These two haplotypes were antithetical for the RFLPs detected by all five enzymes, indicating that they represent very distinct forms of DQ . The remaining 16 haplotypes were infrequent or unique and were closely related to either a DQ3.DR4.a or DQ3.DR4.b. Two of the RFLPs detected, a 5.8 kb Bg1 II fragment and a 10.5 kb Barn HI fragment, had increased frequencies in disease-associated chromosomes. However, none of the RFLPs we detected exhibited a statistically significant increase in IDD or control populations. In contrast, the DQ3.DR4.b DQ haplotype was significantly decreased in IDD-associated DR4 chromosomes. (P=0.04). These results suggest that the DQ3.DR4.b DQ allele may be protective for the development of IDD.     

Age at maturity in landlocked and anadromous Atlantic salmon parr from two Québec rivers

Synopsis Age at maturity in male Atlantic salmon parr from landlocked populations in the Watshishou and Musquaro Rivers is significantly greater than in anadromous populations from the same rivers. We conclude that high post-smolt mortality in anadromous stocks is conducive to male parr maturity at an early age. We also suggest that the lower proportion of maturing male parr in landlocked stocks may be related to competition among males for mates and the smaller ultimate size of spawning adult landlocked salmon.     

A miniaturised parallel-plate shearing apparatus for the measurement of cell adhesion

The design and construction of a miniaturised shearing apparatus is described. Cultures (24 h) of an established epithelial cell line (BEB) were exposed to flow conditions in the shearing chamber at 37 degrees C, and subsequently glass coverslip cultures were prepared for photography. The critical shear radius (CSR) was determined by densitometry from a negative film and the minimum distraction force (MDF) at the CSR calculated using predetermined viscosity values of the flow medium. The mean calculated MDF of BEB cells ranged from 1.04-1.36 Nm-2, and was independent of the culture inoculation density (9 to 37 X 10(4) cells cm-2) and the time (5-20 min) of exposure to shearing conditions. The MDF was increased to 3.3 Nm-2 by a 30 min exposure of cultures to concanavalin A (50 micrograms ml-1), and this effect was abolished by treating with mannose (0.05 M). The results demonstrated that the radial flow chamber principle is applicable to the measurement of cell to substratum adhesion of cultured mammalian cells.     

Flow cytometric analysis of protein thiol groups in relation to the cell cycle and the intracellular content of glutathione in rat hepatocytes

The relationship between protein thiols (PSH) and cell proliferation was examined in ethanol-fixed rat hepatocytes. A new protocol was developed for simultaneous measurement of protein thiol vs. DNA content by flow cytometry. The fluorescent dye o-phthalaldehyde (OPT) was used for flow cytometric measurements of protein thiol groups. The influence of nonprotein thiols was examined by monitoring the cell cycle of cells in which the glutathione content (GSH) was modified by treatment with buthionine sulphoximine (BSO). Three rat liver cell lines (IAR 20, IAR 6.1, IAR 6.1RT7) were used: these cell lines possess different growth characteristics and degrees of tumorigenicity, which made it possible to analyse changes in PSH during normal and deranged cell proliferation. The effects on the cell cycle of the changes in PSH due to the depletion of GSH were measured by 5-bromo-2'-deoxyuridine (BrdUrd) incorporation and flow cytometry. The data obtained can be summarised as follows: a) OPT fluorescence increases with increasing DNA content in all rat liver cell lines examined; b) the greatest variation in PSH content occurs in G1. There is a smaller variation in G2 + M, and PSH levels are relatively invariant throughout S-phase; c) a higher content of PSH is found in the tumorigenic cell lines; d) the amount and distribution of PSH is not affected by BSO treatment; e) kinetic studies indicate that BSO treatment has no effect on the ability of the IAR rat liver cell lines to progress through the cycle.