title: Standardisation of uterine natural killer (uNK) cell measurements in the endometrium of women with recurrent reproductive failure.
abstract: Considerable work is being carried out on endometrial NK cells to determine whether they play a role in successful pregnancy outcome. In addition there is debate about whether measurements of uNK should be included in the clinical assessment for women with recurrent implantation failure or recurrent miscarriage. A hindrance to taking this forward is the fact that the density of uNK cells reported by different centres is very different. The aim of this study was to determine the reason for these differences and to develop a standardised method. Three centres participated in the study. Each centre exchanged five formalin fixed, wax embedded sections of endometrium from five women. Sections were immunostained for CD56. Images were taken of 10 random fields at ×400 magnification; total stromal and uNK cells were counted using Image J. Results were expressed as % positive uNK cells and the variation in counts obtained in each centre was compared. After initial analysis a standardised protocol was agreed and the process repeated. Significant variation was seen in the counts obtained after initial analysis (Centre A vs.B, mean difference=-0.72 P<0.001; A vs.C mean difference=-0.47 P<0.001; B vs.C, mean difference=0.25 P=0.085). Analysis suggested that differences may be due to duration of tissue fixation, the embedding and sectioning processes, selection of areas for assessment, definition of immunopositive cells and inclusion or exclusion of blood vessels. Adoption of a standardised protocol reduced the variation (Centre A vs.B mean difference=-0.105 P=0.744; A vs.C mean difference=0.219 P=0.150; B vs.C mean difference=0.32 P=0.031). Use of a standardised method is needed to establish a normal range for uNK cells and to develop a meaningful clinical test for uNK cell measurements.
authors.affiliation: Reproductive and Vascular Biology Group, Institute of Cellular Medicine, Newcastle University, Newcastle upon Tyne, UK.
authors.forename: Gendie E
authors.initials: GE
authors.lastname: Lash
authors.affiliation: Reproductive and Vascular Biology Group, Institute of Cellular Medicine, Newcastle University, Newcastle upon Tyne, UK.
authors.forename: Judith N
authors.initials: JN
authors.lastname: Bulmer
authors.affiliation: Jessop Wing, Sheffield Teaching Hospitals, Sheffield, UK; Prince of Wales Hospital, Chinese University of Hong Kong, Hong Kong, China.
authors.forename: Tin Chiu
authors.initials: TC
authors.lastname: Li
authors.affiliation: Reproductive and Vascular Biology Group, Institute of Cellular Medicine, Newcastle University, Newcastle upon Tyne, UK.
authors.forename: Barbara A
authors.initials: BA
authors.lastname: Innes
authors.affiliation: Biomedical Research Centre, Sheffield Hallam University, City Campus, Sheffield S1 1WB, UK.
authors.forename: Najat
authors.initials: N
authors.lastname: Mariee
authors.affiliation: Biomedical Research Unit in Reproductive Health, Division of Reproductive Health, Warwick Medical School, University of Warwick, Warwick, UK.
authors.forename: Gnyaneshwari
authors.initials: G
authors.lastname: Patel
authors.affiliation: ScHARR, University of Sheffield, Regent Court, Sheffield, UK.
authors.forename: Jean
authors.initials: J
authors.lastname: Sanderson
authors.affiliation: Biomedical Research Unit in Reproductive Health, Division of Reproductive Health, Warwick Medical School, University of Warwick, Warwick, UK.
authors.forename: Siobhan
authors.initials: S
authors.lastname: Quenby
authors.affiliation: Biomedical Research Centre, Sheffield Hallam University, City Campus, Sheffield S1 1WB, UK. Electronic address: S.M.Laird@shu.ac.uk.
authors.forename: Susan M
authors.initials: SM
authors.lastname: Laird
mesh_terms: D000026:Abortion, Habitual / Q000175:diagnosis / Q000276:immunology*; D000328:Adult; D019002:CD56 Antigen / Q000378:metabolism; D004717:Endometrium / Q000473:pathology*; D005260:Female; D006801:Humans; D007150:Immunohistochemistry; D007694:Killer Cells, Natural / Q000276:immunology*; D018655:Lymphocyte Count; D016612:Paraffin Embedding; D011247:Pregnancy; D011256:Pregnancy Outcome; D012015:Reference Standards; D014599:Uterus / Q000473:pathology*
chemical_list: D019002:CD56 Antigen
keywords: Assessment protocol; Endometrium; Recurrent implantation failure; Recurrent miscarriage; Uterine natural killer cells