Day case superficial parotidectomy-does it work?

PURPOSE: To establish if day case superficial parotidectomy is feasible, safe and does not result in excess readmissions. METHOD: A retrospective review was carried out of all patients listed for superficial parotidectomy with day case intent by a single surgeon between January 2016 and December 2019 inclusively. The reasons for failure of same day discharge were established. Postoperative complications and readmissions were recorded. Our approach for a superficial parotidectomy typically includes the use of a 10Fr suction drain which is removed at 4 h postoperatively if the output is less than 30 ml. RESULTS: Ninety-one consecutive superficial parotidectomies listed for day case surgery were eligible for inclusion. Seventeen patients failed to be discharged on the same day and were admitted giving a day case success rate of 81%. Most of these (n = 9) occurred in the first year of adopting day case surgery. The most common reason to admit patients was a late finish (n = 8, 47%). Six patients (25%) were admitted due to anaesthetic complications. One patient had a surgical complication requiring admission. CONCLUSION: Our series demonstrates that day case superficial parotidectomy using a surgical drain is feasible, safe and does not result in an unacceptable readmission rate. In our experience, surgical complications are an uncommon cause for day case failure. The most common cause for day case failure was a late finish. Postoperative complications including bleeding, seroma/salivary collection and facial nerve palsy were in keeping with or better than those quoted in the literature.

Preparing for patient partnership: A scoping review of patient partner engagement and evaluation in research.

BACKGROUND: Realizing patient partnership in research requires a shift from patient participation in ancillary roles to engagement as contributing members of research teams. While engaging patient partners is often discussed, impact is rarely measured. OBJECTIVE: Our primary aim was to conduct a scoping review of the impact of patient partnership on research outcomes. The secondary aim was to describe barriers and facilitators to realizing effective partnerships. SEARCH STRATEGY: A comprehensive bibliographic search was undertaken in EBSCO CINAHL, and Embase, MEDLINE and PsycINFO via Ovid. Reference lists of included articles were hand-searched. INCLUSION CRITERIA: Included studies were: (a) related to health care; (b) involved patients or proxies in the research process; and (c) reported results related to impact/evaluation of patient partnership on research outcomes. DATA EXTRACTION AND SYNTHESIS: Data were extracted from 14 studies meeting inclusion criteria using a narrative synthesis approach. MAIN RESULTS: Patient partners were involved in a range of research activities. Results highlight critical barriers and facilitators for researchers seeking to undertake patient partnerships to be aware of, such as power imbalances between patient partners and researchers, as well as valuing of patient partner roles. DISCUSSION: Addressing power dynamics in patient partner-researcher relationships and mitigating risks to patient partners through inclusive recruitment and training strategies may contribute towards effective engagement. Further guidance is needed to address evaluation strategies for patient partnerships across the continuum of patient partner involvement in research. CONCLUSIONS: Research teams can employ preparation strategies outlined in this review to support patient partnerships in their work.

Lasing within Live Cells Containing Intracellular Optical Microresonators for Barcode-Type Cell Tagging and Tracking.

We report on a laser that is fully embedded within a single live cell. By harnessing natural endocytosis of the cell, we introduce a fluorescent whispering gallery mode (WGM) microresonator into the cell cytoplasm. On pumping with nanojoule light pulses, green laser emission is generated inside the cells. Our approach can be applied to different cell types, and cells with microresonators remain viable for weeks under standard conditions. The characteristics of the lasing spectrum provide each cell with a barcode-type label which enables uniquely identifying and tracking individual migrating cells. Self-sustained lasing from cells paves the way to new forms of cell tracking, intracellular sensing, and adaptive imaging.

Monitoring the Rab27 associated exosome pathway using nanoparticle tracking analysis.

Exosomes are secreted by many cell types and display multiple biological functions. The ability to both rapidly detect and quantify exosomes in biological samples would assist in the screening of agents that interfere with their release, and which may therefore be of clinical relevance. Nanoparticle tracking analysis, which detects the size and concentration of exosomes, was used to monitor the inhibition of exosome secretion from MDA-MB-231 breast cancer cells expressing inhibitory RNA targeted for Rab27a, a known component of the exosome pathway. Inhibition of both Rab27a and Rab27b was observed, resulting in alterations to intracellular CD63+ compartments and the release of fewer exosomes into the culture medium, as determined by nanoparticle tracking analysis and confirmed by immunoblotting and protein quantification. These data show that nanoparticle tracking analysis can be used effectively and rapidly to monitor the disruption of exosome secretion.

Public sphere as assemblage: the cultural politics of roadside memorialization.

This paper investigates contemporary academic accounts of the public sphere. In particular, it takes stock of post-Habermasian public sphere scholarship, and acknowledges a lively and variegated debate concerning the multiple ways in which individuals engage in contemporary political affairs. A critical eye is cast over a range of key insights which have come to establish the parameters of what 'counts' as a/the public sphere, who can be involved, and where and how communicative networks are established. This opens up the conceptual space for re-imagining a/the public sphere as an assemblage. Making use of recent developments in Deleuzian-inspired assemblage theory - most especially drawn from DeLanda's (2006) 'new philosophy of society' - the paper sets out an alternative perspective on the notion of the public sphere, and regards it as a space of connectivity brought into being through a contingent and heterogeneous assemblage of discursive, visual and performative practices. This is mapped out with reference to the cultural politics of roadside memorialization. However, a/the public sphere as an assemblage is not simply a 'social construction' brought into being through a logic of connectivity, but is an emergent and ephemeral space which reflexively nurtures and assembles the cultural politics (and political cultures) of which it is an integral part. The discussion concludes, then, with a consideration of the contribution of assemblage theory to public sphere studies. (Also see Campbell 2009a).

The multi-faceted nature of HLA class I dimer molecules.

The canonical role of major histocompatibility complex class I (MHCI) molecules in antigen presentation involves the recognition of a short peptide of intracellular origin, bound to the upper surface of the class I molecule, by CD8(+) T lymphocytes. Assembly and loading of the MHCI is a highly regulated, chaperone-mediated process and only when the fully folded MHCI molecule is correctly loaded with peptide is it released from the endoplasmic reticulum for trafficking to the cell surface. Current models of the interactions of MHCI molecules with their cognate receptors visualize them functioning as monomeric entities. However, in recent years, new data have revealed MHCI molecules with the ability to form disulphide-linked dimeric structures, with several distinct dimer entities being elucidated. We describe here three types of MHCI dimers; HLA-B27 dimers formed predominantly through the possession of an unpaired cysteine within the peptide-binding groove; HLA-G dimers, which form through a cysteine on its external surface; and a novel population we term redox-induced dimers, which can form between cysteine residues in the cytoplasmic tail domains. The characteristics of these dimeric MHCI molecules and their role in both normal immune responses and in disease pathogenesis are reviewed in this article.

MHC class I dimer formation by alteration of the cellular redox environment and induction of apoptosis.

Many MHC class I molecules contain unpaired cysteine residues in their cytoplasmic tail domains, the function of which remains relatively uncharacterized. Recently, it has been shown that in the small secretory vesicles known as exosomes, fully folded MHC class I dimers can form through a disulphide bond between the cytoplasmic tail domain cysteines, induced by the low levels of glutathione in these extracellular vesicles. Here we address whether similar MHC class I dimers form in whole cells by alteration of the redox environment. Treatment of the HLA-B27-expressing Epstein-Barr virus-transformed B-cell line Jesthom, and the leukaemic T-cell line CEM transfected with HLA-B27 with the strong oxidant diamide, and the apoptosis-inducing and glutathione-depleting agents hydrogen peroxide and thimerosal, induced MHC class I dimers. Furthermore, induction of apoptosis by cross-linking FasR/CD95 on CEM cells with monoclonal antibody CH-11 also induced MHC class I dimers. As with exosomal MHC class I dimers, the formation of these structures on cells is controlled by the cysteine at position 325 in the cytoplasmic tail domain of HLA-B27. Therefore, the redox environment of cells intimately controls induction of MHC class I dimers, the formation of which may provide novel structures for recognition by the immune system.

Nanoparticle tracking analysis monitors microvesicle and exosome secretion from immune cells.

Nanoparticle tracking analysis permits the determination of both the size distribution and relative concentration of microvesicles, including exosomes, in the supernatants of cultured cells and biological fluids. We have studied the release of microvesicles from the human lymphoblastoid T-cell lines Jurkat and CEM. Unstimulated, both cell lines release microvesicles in the size range 70-90 nm, which can be depleted from the supernatant by ultracentrifugation at 100 000 g, and by anti-CD45 magnetic beads, and which by immunoblotting also contain the exosome-associated proteins Alix and Tsg101. Incubation with known potentiators of exosome release, the ionophores monensin and A23187, resulted in a significant increase in microvesicle release that was both time and concentration dependent. Mass spectrometric analysis of proteins isolated from ultracentrifuged supernatants of A23187-treated cells revealed the presence of exosome-associated proteins including heat-shock protein 90, tubulin, elongation factor α1, actin and glyceraldehyde 3-phosphate dehydrogenase. Additionally, treatment of peripheral blood monocyte-derived dendritic cells with bacterial lipopolysaccharide displayed an increase in secreted microvesicles. Consequently, nanoparticle tracking analysis can be effectively applied to monitor microvesicle release from cells of the immune system.

The Changing Face of Head and Neck Cancer; How the Head and Neck One-Stop Clinic Has Evolved Since Conception.

Purpose: The purpose of this study was to compare the patient journey through the head and neck clinic across 13 years of service improvement. We aimed to compare pick-up rates of cancer; number of patients receiving tissue diagnoses at first visit; and number of patients who were discharged on their first visit. Methods: In the one-stop head and neck cancer clinic, the demographic data, investigations and outcomes for 277 patients who attended in 2004 were compared to those of 205 patients who attended in 2017. The number of patients receiving ultrasonography and fine needle aspiration cytology was compared. Patient outcomes were analysed: specifically, the number discharged on first visit and the number of malignancies diagnosed. Results: The pick-up rate for malignancy from 2004 to 2017 has remained stable (17.3% vs 17.1%). The number of patients receiving ultrasound has remained stable from 264 (95%) in 2004 to 191 (93%) in 2017. The number undergoing FNA has decreased from 139 (50%) to 68 (33%) (p < 0.01). The number of patient's discharged on the first visit has significantly increased from 82 (30%) in 2004 to 89 (43%) in 2017 (p < 0.01). Conclusion: The one-stop clinic provides an effective and efficient means of head and neck lump assessment. Since inception of this service, the accuracy of diagnostic investigation has improved over time.

Expression of MHC class I dimers and ERAP1 in an ankylosing spondylitis patient cohort.

The human leucocyte antigen HLA-B27 is strongly associated with ankylosing spondylitis, a form of seronegative inflammatory arthritis. In this study aspects related to several hypothesized mechanisms of disease pathogenesis have been investigated. Blood monocyte-derived dendritic cells (DC) from a small patient cohort of 29 patients with ankylosing spondylitis and one with reactive arthritis, were compared with DC from 34 healthy control subjects, of whom four were found to be HLA-B27 positive. The ability of HLA-B27 to form heavy-chain dimers reactive with monoclonal antibody HC10 was tested, along with the induction of endoplasmic reticulum (ER) stress, assessed by splicing xbp1 mRNA and immunoblotting of Immunoglobulin Binding Protein (BiP). Additionally, the protein expression levels of the ER resident aminopeptidase gene ERAP1 in patients with ankylosing spondylitis was also determined, following its recent identification as a novel disease-associated gene. No significant difference was noted in the global levels of HC10-reactive MHC class I dimers formed in either the patient or control DC populations. Stress on the ER, as determined by xbp1 mRNA splicing, was not detected but lower levels of BiP were observed in the DC from patients. Of further potential interest, in this patient cohort the expression of ERAP1 appeared to be higher in a number of patient DC samples when compared with controls, suggesting over-expression of ERAP1 as a mechanism promoting ankylosing spondylitic pathogenesis.

Novel MHC class I structures on exosomes.

Exosomes are nanometer-sized vesicles released by a number of cell types including those of the immune system, and often contain numerous immune recognition molecules including MHC molecules. We demonstrate in this study that exosomes can display a significant proportion of their MHC class I (MHC I) content in the form of disulfide-linked MHC I dimers. These MHC I dimers can be detected after release from various cell lines, human monocyte-derived dendritic cells, and can also be found in human plasma. Exosome-associated dimers exhibit novel characteristics which include 1) being composed of folded MHC I, as detected by conformational-dependent Abs, and 2) dimers forming between two different MHC I alleles. We show that dimer formation is mediated through cysteine residues located in the cytoplasmic tail domains of many MHC I molecules, and is associated with a low level of glutathione in exosomes when compared with whole cell lysates. We propose these exosomal MHC I dimers as novel structures for recognition by immune receptors.

ERp57 interacts with conserved cysteine residues in the MHC class I peptide-binding groove.

The oxidoreductase ERp57 is a component of the major histocompatibility complex (MHC) class I peptide-loading complex. ERp57 can interact directly with MHC class I molecules, however, little is known about which of the cysteine residues within the MHC class I molecule are relevant to this interaction. MHC class I molecules possess conserved disulfide bonds between cysteines 101-164, and 203-259 in the peptide-binding and alpha3 domain, respectively. By studying a series of mutants of these conserved residues, we demonstrate that ERp57 predominantly associates with cysteine residues in the peptide-binding domain, thus indicating ERp57 has direct access to the peptide-binding groove of MHC class I molecules during assembly.

Polymorphisms in the F Pocket of HLA-B27 Subtypes Strongly Affect Assembly, Chaperone Interactions, and Heavy-Chain Misfolding.

OBJECTIVE: HLA-B27 is associated with the inflammatory spondyloarthritides (SpA), although subtypes HLA-B*27:06 and HLA-B*27:09 are not. These subtypes differ from the HLA-B*27:05 disease-associated allele primarily at residues 114 and 116 of the heavy chain, part of the F pocket of the antigen-binding groove. Dimerization of HLA-B27 during assembly has been implicated in disease onset. The purpose of this study was to investigate the factors that influence differences in dimerization between disease-associated and non-disease-associated HLA-B27 alleles. METHODS: HLA-B*27:05 and mutants resembling the HLA-B*27:06 and 09 subtypes were expressed in the rat C58 T cell line, the human CEM T cell line and its calnexin-deficient variant CEM.NKR. Immunoprecipitation, pulse-chase experiments, flow cytometry, and immunoblotting were performed to study the assembly kinetics, heavy-chain dimerization, and chaperone associations. RESULTS: By expressing HLA-B*27:05, 06-like, and 09 alleles on a restrictive rat transporter associated with antigen processing background, we demonstrate that a tyrosine expressed at p116, either alone or together with an aspartic acid residue at p114, inhibited HLA-B27 dimerization and increased the assembly rate. F-pocket residues altered the associations with chaperones of the early major histocompatibility complex class I folding pathway. Calnexin was demonstrated to participate in endoplasmic reticulum (ER) stress-mediated degradation of dimers, whereas the oxidoreductase ERp57 does not appear to influence dimerization. CONCLUSION: Residues within the F pocket of the peptide-binding groove, which differ between disease-associated and non-disease-associated HLA-B27 subtypes, can influence the assembly process and heavy-chain dimerization, events which have been linked to the initiation of disease pathogenesis.

Visualization of podocyte substructure with structured illumination microscopy (SIM): a new approach to nephrotic disease.

A detailed microscopic analysis of renal podocyte substructure is essential to understand and diagnose nephrotic kidney disease. Currently only time consuming electron microscopy (EM) can resolve this substructure. We used structured illumination microscopy (SIM) to examine frozen sections of renal biopsies stained with an immunofluorescence marker for podocin, a protein localized to the perimeter of the podocyte foot processes and compared them with EM in both normal and nephrotic disease biopsies. SIM images of normal glomeruli revealed curvilinear patterns of podocin densely covering capillary walls similar to podocyte foot processes seen by EM. Podocin staining of all nephrotic disease biopsies were significantly different than normal, corresponding to and better visualizing effaced foot processes seen by EM. The findings support the first potential use of SIM in the diagnosis of nephrotic disease.

Integrity and stability of the citrulline-arginine pathway in normal and tumour cell lines.

Arginine catabolizing enzymes have been used on cancers for over 60 years. In the last 5 years the ability of arginine catabolizing enzymes, not only to inhibit proliferation, but to kill tumour cells has been reinvestigated. Selectivity of action lies in the inability of many tumours to circumvent arginine deprivation by recycling precursors through the urea cycle. While this offers an immediate window of opportunity to treat, e.g. melanomas and hepatocellular carcinomas (HCC) that have poor citrulline converting ability, it is possible that the deprivation can be applied to many other types of cancer. The problem of deficiency of the urea cycle enzymes in a wider range of normal and malignant cell lines has been addressed, and shown to be variable throughout several different tumour types. We also need to know how fickle recycling enzyme activity can be in both normal and tumour cells, and found to be remarkable stable. Increasing interest is shown in the amino acid (arginine) deprivation protocol because it has already moved into the clinic. Initial findings on a named-patient basis have been encouraging, and the development of a new rational approach to the systemic treatment of melanomas, HCCs and leukemias seems imminent. This is the more attractive because arginine deprivation protocols can also 'stage' tumour cells for combination therapy in cases where they might not be killed outright by deprivation alone.

The use of wavelength modulated Raman spectroscopy in label-free identification of T lymphocyte subsets, natural killer cells and dendritic cells.

Determining the identity of cells of the immune system usually involves destructive fixation and chemical staining, or labeling with fluorescently labeled antibodies recognising specific cell surface markers. Completely label-free identification would be a significant advantage in conditions where untouched cells are a priority. We demonstrate here the use of Wavelength Modulated Raman Spectroscopy, to achieve label-free identification of purified, unfixed and untouched populations of major immune cell subsets isolated from healthy human donors. Using this technique we have been able to distinguish between CD4(+) T lymphocytes, CD8(+) T lymphocytes and CD56(+) Natural Killer cells at specificities of up to 96%. Additionally, we have been able to distinguish between CD303(+) plasmacytoid and CD1c(+) myeloid dendritic cell subsets, the key initiator and regulatory cells of many immune responses. This demonstrates the ability to identify unperturbed cells of the immune system, and opens novel opportunities to analyse immunological systems and to develop fully label-free diagnostic technologies.

Arginine catabolism, liver extracts and cancer.

Although it is self evident that cells will not grow in amino acid deficient medium, an observation less well appreciated is that malignant cells are particularly vulnerable to such deprivation, which can lead to their rapid demise. Indeed, the more flagrantly malignant the phenotype (anaplastic the tumor), the more susceptible the cells seem to be to deprivation. While some attempts to employ this strategy in cancer treatment have been made, the difference between normal and malignant cells should be more fully exploited as a means of selectively eliminating tumor cell populations. To be successful, information on differences between the normal and the deranged cell cycle engine and checkpoints, especially how these are affected by deprivation, is of crucial importance. Since it is only recently that the controls at restriction points have been elucidated, it is little surprise that earlier attempts to control tumor cell growth by limiting the availability of an essential amino acid have met with limited success. Studies have been sporadic and isolated, often with little more than anecdotal descriptions as far as clinical work was concerned. This review concentrates on what has been accomplished primarily in vitro and since about 1950 with regard to arginine catabolism, while recognising that other essential amino acids have also been the focus of attention by some investigators. Treatments have included medium and plasma manipulation, dietary control, enzymatic degradation, and the use of liver extracts. On some occasions, substitution of amino acid analogues has been explored. It is argued that current knowledge, combined with past experience, calls for a much closer examination of the full potential of amino acid (and specifically arginine) deprivation as a means of controlling tumor growth, with greater attention to protocols that might be used to treat human cancers.

Regulation of exosome release from mammary epithelial and breast cancer cells - a new regulatory pathway.

PURPOSE: Exosomes are small 50-100nm sized extracellular vesicles released from normal and tumour cells and are a source of a new intercellular communication pathway. Tumour exosomes promote tumour growth and progression. What regulates the release and homoeostatic levels of exosomes, in cancer, in body fluids remains undefined. METHODS: We utilised a human mammary epithelial cell line (HMEC B42) and a breast cancer cell line derived from it (B42 clone 16) to investigate exosome production and regulation. Exosome numbers were quantified using a Nanosight LM10 and measured in culture supernatants in the absence and presence of exosomes in the medium. Concentrated suspensions of exosomes from the normal mammary epithelial cells, the breast cancer cells and bladder cancer cells were used. The interaction of exosomes with tumour cells was also investigated using fluorescently labelled exosomes. RESULTS: Exosome release from normal human mammary epithelial cells and breast cancer cells is regulated by the presence of exosomes, derived from their own cells, in the extracellular environment of the cells. Exosomes from normal mammary epithelial cells also inhibit exosome secretion by breast cancer cells, which occurs in a tissue specific manner. Labelled exosomes from mammary epithelial cells are internalised into the tumour cells implicating a dynamic equilibrium and suggesting a mechanism for feedback control. CONCLUSIONS: These data suggest a previously unknown novel feedback regulatory mechanism for controlling exosome release, which may highlight a new therapeutic approach to controlling the deleterious effects of tumour exosomes. This regulatory mechanism is likely to be generic to other tumours.