Elsevier

Biomaterials

Volume 29, Issues 24–25, August–September 2008, Pages 3444-3450
Biomaterials

Extrinsic and intrinsic pathways of apoptosis in aseptic loosening after total hip replacement

https://doi.org/10.1016/j.biomaterials.2008.04.044Get rights and content

Abstract

Particle-induced osteolysis is a major cause of aseptic loosening after total joint replacement. The purpose of the current study was to identify various apoptosis-related pathways in the cellular response to wear debris. Fas receptor, BAK and caspase-3 cleaved were evaluated immunohistochemically in capsules and interface membranes from patients with aseptic hip implant loosening. Moreover, we investigated local cellular proliferation, documented by the presence of Ki-67, to evaluate the proportion of apoptosis in relation to the proliferation in the different cells. We detected a strong expression of caspase-3 cleaved, Fas and BAK in macrophages, giant cells and T-lymphocytes. The fibroblasts showed caspase-3 cleaved and BAK, but no Fas staining. Demonstrated by Ki-67 staining, we found increased proliferation of macrophages and fibroblasts. Statistical analysis showed a significant positive correlation (p < 0.001) between the above mentioned results and the presence of wear debris. The intensity of apoptosis and proliferation differed, depending on the extent of osteolysis. Overall, four different patterns of immunoreactivity were identified. We think, however, that in particle-induced osteolysis apoptosis is pathologically increased – a phenomenon also seen in other diseases. In these instances, the number and degree of apoptotic reactions are so great that the resulting cell remains cannot be completely removed. This leads to an increased excretion of fibrogenic mediators that could be responsible for increased proliferation of fibroblasts in spite of the increased apoptosis. Moreover, it leads to an increased excretion of cytokines which could be responsible for the activation of osteoclasts.

Introduction

Particle-induced osteolysis is a major cause of aseptic loosening after hip joint replacement. Periprosthetic osteolysis is initiated by an aseptic inflammatory response to phagocytosis of implant wear particles resulting in increased proliferation and differentiation of osteoclast precursors into mature osteoclasts [1]. Progressive osteolysis can result in implant instability and failure, eventually requiring revision surgery. The complexity of this topic is reflected by the large number of published studies on particle-induced bio-reactivity. While the osteolytic cascade initiated by cytokine release from macrophages has been studied extensively, the induction of apoptosis in this context has been addressed by only a few authors [2], [3]. Thus far, it has been shown in vitro that ceramic, metal and polyethylene particles can induce apoptosis of macrophages [4], [5]. In a previous study we described apoptosis of macrophages, giant cells and T-lymphocytes in capsules and interface membranes of patients with aseptic hip implant loosening in vivo [6]. In that study we examined members of the Bcl-2 family. In this current study we investigated additional apoptosis-related markers in order to gain more insight into the role of apoptosis in aseptic loosening.

Caspase-3 has been identified as a key mediator of apoptosis of mammalian cells. Caspases are synthesized as inactive proforms and on activation they cleave to aspartate residues [7]. Caspases cleave a number of different substrates in the cytoplasm or nucleus leading to many of the morphological features of apoptotic cell death [7]. Activation of caspase-3 can be initiated at the plasma membrane upon ligation of death receptors (extrinsic pathway) or at the mitochondria (intrinsic pathway). There are therefore two different entry points for caspase-3 activation [8]. The extrinsic and intrinsic pathways merge at caspase-3. Activation of caspase-3 has been shown to be a key regulatory checkpoint that regulates terminal events in the apoptotic cascade [9], [10]. In the apoptotic effector pathway, caspase-3 is probably best correlated with apoptosis [11].

Fas (CD95/APO-1) is a death receptor of the tumour necrosis factor (TNF) receptor super-family. Its stimulation results in activation of the initiator caspase-8 which can propagate the apoptosis signal by direct cleavage of downstream effector caspases such as caspase-3 [12]. Binding of Fas ligand (FasL/CD95L) to the Fas receptor results in clustering of receptors and initiates the extrinsic pathway. This begins outside a cell when conditions in the extracellular environment determine that a cell must die [13].

BAK is a marker of the intrinsic (mitrochondrion) pathway which begins when an injury occurs within the cell. In many instances the apoptotic pathways involve an obligatory propagation through mitochondria. Central to the mitochondrial checkpoint is a complex interplay between members of the Bcl-2 family, which consists of an antiapoptotic subgroup including Bcl-2, and even the pro-apoptotic BAK, BAX and BH3-domain-only subgroups [14].

Beside apoptosis, we investigated cellular proliferation in aseptic loosening as documented by the presence of Ki-67, a cell-cycle-related nuclear protein, expressed by proliferating cells in all phases of the active cell-cycle (G1, S, G2 and M phases). It is absent in resting (G0) cells [15].

The purpose of this study was to identify additional apoptosis-related pathways in the macrophage response to wear debris and to evaluate the proportion of apoptosis in relation to the proliferation in the different cells. We formulated the following questions: Are the extrinsic and intrinsic pathways involved in the apoptotic process to the same degree? Which kinds of cells are involved? Does apoptosis correlate with disease severity, measured by Paprosky's classification of particle-induced osteolytic changes around hip implants? Does wear debris also induce proliferation as well as apoptosis?

Section snippets

Patients

The study was approved by the local Ethics Committee. Three patient groups were distinguished for this study. All three groups consisted of consecutive cases operated at the Department of Orthopaedics of the University of Duisburg-Essen. Group 1 consisted of total hip arthroplasty cases, while Groups 2 and 3 served as controls.

Group 1: The group consisted of 21 patients, 13 females and 8 males. The mean age was 70.3, ranging from 45 to 85 (standard deviation (SD): 11.4). The mean survival time

Histology

Group 1: HE staining of the interface tissue revealed typical features of membranes that form in response to arthroplasty-derived particles, i.e. numerous fibrous cells in a bed of connective tissue. In the capsule tissue HE staining showed the fibrous membrane with collagen fibres, and the synovial membrane with loose connective tissue, fibrous and synovial cells.

Varying quantities of wear debris, including metal and polyethylene particles, were found in the capsule and interface tissues,

Hypotheses

Apoptosis is an active form of cell death which plays an important role in the physiological turnover of normal cells for the maintenance of tissue homeostasis. Apoptosis enables human beings to control cell number in tissues and to eliminate individual cells that threaten the survival of human beings. The purpose of the current study was to identify various apoptosis-related pathways in the cellular response to wear debris.

Causative relation between particles and apoptosis in particle-surrounding macrophages and giant cells

The presence of polyethylene and metal debris in capsules and

Conclusions

In this study, apoptosis of fibroblasts, macrophages, giant cells and T-lymphocytes has been detected in vivo in the capsules and interface membranes of patients with aseptic hip implant loosening. Both the intrinsic and the extrinsic pathways are involved in the activation of apoptosis in these cells, with exception of the fibroblasts, which showed no Fas staining. We saw varying degrees of apoptotic reactions and proliferation of macrophages and fibroblasts in the interfaces. We were able to

Acknowledgements

This study was supported by a grant from the Verein der Freunde und Förderer des Evangelischen Krankenhauses Essen-Werden e.V.

We thank Prof. Dr. K.W. Schmid from the Institute of Pathology and Neuropathology, University of Duisburg-Essen for advice and review of the current study and Mrs. N. Cramer for technical assistants.

References (43)

  • J. Savill et al.

    Phagocyte recognition of cells undergoing apoptosis

    Immunol Today

    (1993)
  • J. Savill et al.

    Granulocyte clearance by apoptosis in the resolution of inflammation

    Semin Cell Biol

    (1995)
  • S. Stea et al.

    Apoptosis in peri-implant tissue

    Biomaterials

    (2000)
  • J.J. Cohen

    Apoptosis

    Immunol Today

    (1993)
  • J.T. Wang et al.

    The role of particulate orthopaedic implant materials in peri-implant osteolysis

  • M.J. Silva et al.

    What's new in orthopaedic research

    J Bone Joint Surg Am

    (2002)
  • O.L. Huk et al.

    Apoptosis in interface membranes of aseptically loose total hip arthroplasty

    J Mater Sci Mater Med

    (2001)
  • D. Granchi et al.

    Cell death induced by metal ions: necrosis or apoptosis?

    J Mater Sci Mater Med

    (1998)
  • A. Degterev et al.

    A decade of caspases

    Oncogene

    (2003)
  • M.O. Hengartner

    The biochemistry of apoptosis

    Nature

    (2000)
  • S. Roy et al.

    Maintenance of caspase-3 proenzyme dormancy by an intrinsic “safety catch” regulatory tripeptide

    Proc Natl Acad Sci U S A

    (2001)
  • Cited by (44)

    • Contributions of human tissue analysis to understanding the mechanisms of loosening and osteolysis in total hip replacement

      2014, Acta Biomaterialia
      Citation Excerpt :

      The control samples involved in almost all studies are synovial tissues from primary osteoarthritis taken before THA/total knee arthroplasty surgery, rarely synovial tissue from around a well-fixed THA, tissues from mandibular or maxilla fractures or biological ingrowth membranes [27,31,38,40,41,48,50,52,53,66,68,88,90–92,101,106,134]. The periprosthetic tissue from deceased patients with THA was also used [5,23,24,61–63,130]. One important factor potentially influencing the results from tissue analysis is the location of the harvested specimen [115].

    • Adiponectin attenuates osteolysis in aseptic loosening of total hip replacements

      2014, Acta Biomaterialia
      Citation Excerpt :

      However, knowledge about the role of apoptosis in this context is limited. Previous studies have demonstrated that apoptosis in macrophages and giant cells during particle engulfment is partially responsible for osteolysis in aseptic loosening of joint implants [2–4]. We demonstrated by means of a murine calvarial model of wear particle-induced osteolysis that inhibition of apoptosis leads to decreasing bone resorption by osteoclasts [5].

    • Dihydroartemisinin exhibits antitumor activity toward hepatocellular carcinoma in vitro and in vivo

      2012, Biochemical Pharmacology
      Citation Excerpt :

      Generally, apoptosis is executed by two pathways, the intrinsic and the extrinsic. The intrinsic pathway, also called mitochondrial pathway, is usually triggered by DNA damage, and the extrinsic pathway, known as the receptor-mediated pathway, is initiated by death receptor activation [18]. DHA has been demonstrated to induce apoptosis through both apoptotic pathways.

    View all citing articles on Scopus
    View full text