Heterogeneity in cellular and cytokine profiles from multiple samples of tissue surrounding revised hip prostheses

Abstract

Previous studies have attempted to define the biologic properties of the bone-implant interface using a single specimen harvested from the periprosthetic tissues. The purpose of this study was to examine the heterogeneity in cellular and cytokine profiles of multiple samples taken from the tissues surrounding revised hip prostheses. Clinical and radiographic data for nine patients undergoing surgical revision was gathered prospectively. Three tissue samples were taken systematically from the acetabular and/or femoral bed. Morphologic characteristics of the tissues were assessed using hematoxylin and eosin-stained sections. Immunohistochemical staining was performed using monoclonal antibodies to identify macrophages (EMB11 and CD68); activated macrophages (Leu M3); total T lymphocytes (Leu 4 and T11); T-helper lymphocytes (Leu 3A and CD4); cytotoxic/suppressor T lymphocytes (Leu 2A and CD3); and fibroblasts (5B5). In situ hybridization was used to identify the mRNA for specific proteins: interleukin (IL)1 alpha and -beta, IL-2, IL-6, transforming growth factor beta, tumor necrosis factor alpha (TNF alpha), platelet-derived growth factor alpha (PDGF alpha), and interferon gamma. A quantitative assessment was performed for each section by calculating the percentage of positively staining cells using a light microscope and grid-counting technique. A random effect analysis of variance was calculated to determine both the variance between samples within each patient and the variance between different patients. Standard deviations contributed by sampling variance and patient variance were calculated and an F test was applied. Tissue samples taken from different regions of the bone-prosthesis interface showed marked heterogeneity in cellular and cytokine profiles. Critical F values indicating a statistically significant degree of variance between different tissue samples were exceeded for macrophages, cytotoxic/suppressor T lymphocytes, and T-helper lymphocytes. The cytokine profile was significantly different for IL-2, PDGF alpha, and TNF alpha. This tissue heterogeneity may be due to different mechanical and biologic environments along the bone-prosthesis interface. Thus, caution must be exercised in defining the biologic properties of the tissue surrounding revised prostheses according to a single biopsy.

Do Tissues From THA Revision of Highly Crosslinked UHMWPE Liners Contain Wear Debris and Associated Inflammation?

“…Our findings for the conventional cohort are similar to those reported in other immunohistochemical studies of periprosthetic tissues from uncemented, gamma air-sterilized UHMWPE liners [3,5,12,29,40,41]. In general, the colocalization of wear debris and macrophages/histiocytes is well-established [28,41], and several studies have identified the presence of T cells in periprosthetic tissues, with the incidence ranging from categorically absent to 30% [28,40,41].…”

“…The regional and patient differences in the number of macrophages and T cells are supported by previous investigations of immune cells and cytokines in tissues from loosened conventional prostheses [8,29,70]. In agreement with prior immunohistochemical [3,5,12,28,36,40,41] or histologic [42,59] studies, the predominant cells in both cohorts were macrophages and T cells.…”

Cellular profile and cytokine production at prosthetic interfaces

“…The slide is then counterstained so that the cells that have not reacted with the monoclonal antibody are visualised. 20,21 Thick 6 m frozen sections were fixed in absolute acetone at -20°C overnight. The primary mouse antihuman monoclonal antibodies are detailed in Table II.…”

“…We first examined the stained sections to determine the general histomorphological features and then performed a quantitative assessment for each antibody or probe by calculating the percentage of positively staining cells using light microscopy and a grid-counting technique. 21 We classified 50 to 75 cells in each of the four quadrants of the tissue section which yielded a minimum of 200 cell counts per stain or probe. The first quadrant for cell counting was chosen randomly on the slide and labelled the 12 o'clock position; the second, third and fourth quadrants for counting were then automatically designated according to the 3, 6 and 9 o'clock positions.…”

“…The data were analysed using an analysis of variance; intergroup comparisons were made for each antibody or probe The T-lymphocyte monoclonal antibody CD4 was found to bind very weakly to macrophages which were much larger and stained very lightly compared with the T-lymphocytes which were much smaller and stained more intensively. 21 The macrophages were not counted as positive cells.…”