Biology subjects

Edwards, S. W.

Publications and source records attributed to Edwards, S. W..

3 recordsLinked to original sources

bioRxiv2021

An improved culture protocol for the differentiation and survival of human promyelocytic leukaemia PLB-985 cell-line into mature neutrophil-like granulocytes

Circulating blood neutrophils are short-lived, lack proliferation capacity and cannot be transfected in vitro to express exogenous genes or proteins. These properties have made the ex vivo genetic manipulation of neutrophils challenging and hindered biochemical and molecular studies investigating the function of specific genes and proteins. Improved methodology for differentiating cell lines into mature neutrophil-like phenotypes, with similar morphological and functional properties to blood neutrophils would, therefore, be an important tool to probe the molecular properties of mature cells. The PLB-985 cell line was cultured in RPMI-1640 medium supplemented foetal calf serum (FCS) and penicillin/streptomycin. For induction of differentiation into neutrophil-like cells, the medium was supplemented with sodium pyruvate, N, N-dimethylformamide (DMF) and all-trans retinoic acid (ATRA), FCS and penicillin/streptomycin. The cytokines G-CSF and GM-CSF were used to enhance differentiation, prolong viability and delay the progression of the differentiated cells into apoptosis. The modified culture protocol and conditions induced PLB-985 cells to differentiate into mature, neutrophil-like granulocytes that resembled the morphology of mature blood neutrophils as evident by acquisition of a multi-lobed nucleus and granulated cytoplasm. These modified culture conditions resulted in enhanced differentiation into neutrophil-like cells and the apoptosis of these differentiated cells was delayed by supplementation with cytokines. This experimental system should be useful for studies probing the function of specific genes and proteins in human neutrophils.

immunology↗

bioRxiv2021

A structured reflective process supports student awareness of employability skills development in a science placement module

Placements are often an extra-curricular activity of a science degree. This study reports on the outcomes of a final year credit-bearing 6-week placement module that was specifically designed to develop and enhance students employability skills. A key element of this module was that the student placements were not just evaluated from a science perspective, but with an emphasis on meaningful reflection and evaluation of employability skills development. Students recorded their levels of confidence in skills before, during and after the placement via an Online Reflective Log, as part of a modules summative assessment. First, results showed that taking part in the placement and conducting their own independent research helped students to make connections between their scientific knowledge, otherwise constrained within the walls of the undergraduate science lab, and the wider impact of their research on society. Another theme that emerged concerned career choices and aspirations, and the placement experience either confirmed prior choices or opened new horizons. The Online Reflective Log helped students to feel supported by their university supervisor who were at a distance, while feedback on their tasks challenged them to reflect on the scientific and personal skills as they were engaged in scientific activities during placement. Students agreed that they had further developed their employability skills during the placement. Students acknowledged it was challenging to have to acquire evidence of skills development but appreciated the usefulness of this reflection in relation to their future careers.

scientific communication and education↗

bioRxiv2020

Type I Interferon regulates cytokine-delayed neutrophil apoptosis, reactive oxygen species production and chemokine expression via activation of p38 MAPK

ABSTRACTInterferons (IFNs) are key regulators of a number of inflammatory conditions in which neutrophils play an important role in pathology, such as rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE), where Type I IFNs are implicated in disease pathology. However, IFNs are usually generated in vivo together with other cytokines that also have immunoregulatory functions but such interactions are poorly-defined experimentally. We measured the effects of Type-I IFN (IFNα), elevated in both RA and SLE, on the functions of healthy neutrophils incubated in vitro in the absence and presence of pro-inflammatory cytokines typically elevated in inflammatory diseases (TNFα, GM-CSF). IFNα alone had no effect on neutrophil apoptosis, however it did abrogate the anti-apoptotic effect of GM-CSF (18h, p< 0.01). The enhanced stabilty of the anti-apoptotic protein Mcl-1 and delayed activation of caspase activation normally regulated by GM-CSF were blocked by IFNα: this effect was mediated, in part, by activation of p38 MAPK, increased turnover of the anti-apoptotic protein Mcl-1 and cleavage of caspases. IFNα alone also primed ROS production alone and maintained the transient priming effect of TNF for up to 4h: it also down-regulated GM-CSF and TNFα-activated expression of CXCL1, CXCL2, CXCL3, CXCL8, CCL3 and CCL4, but in contrast increased the expression of CXCL10. These novel data identify complex regulatory signalling networks in which Type I IFNs profoundly alter the response of neutrophils to inflammatory cytokines. This is likely to have important consequences in vivo and may explain the complexity and heterogeneity of inflammatory diseases such as RA, in which multiple cytokine cascades have been activated.Competing Interest StatementThe authors have declared no competing interest.View Full Text

immunology↗