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CD36 and CD14 immunoreactivity of Reiter cells in inflammatory synovial fluids
  1. ENRICO SELVI,
  2. STEFANIA MANGANELLI,
  3. RENATO DE STEFANO,
  4. ELENA FRATI,
  5. ROBERTO MARCOLONGO
  1. Institute of Rheumatology, Loc Le Scotte University of Siena, 53100 Siena, Italy
  1. Dr Selvi

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Reiter cells are macrophages containing ingested polymorph nuclei that are commonly found in most inflammatory synovial fluids. Available data indicate that CD36 and CD14 on human monocyte derived macrophages are adhesion molecules involved in several biological processes.1 Of interest, their role in the process of adhesion and phagocytosis of apoptotic cells has been recently demonstrated.2-5

Jones and colleagues demonstrated reduced Reiter cells in the synovial fluids from patients with rheumatoid arthritis. This observation is consistent with the hypothesis that Reiter cells play a regulatory part in preventing autolysis of polymorphonuclear neutrophils (PMN) and thus local tissue damage.6

The purpose of this study was to evaluate by histochemical technique whether Reiter cells express CD36 and CD14 in inflammatory synovial fluids.

We analysed the synovial fluids obtained from the knee joints of 10 patients suffering from inflammatory joint diseases of recent onset (< 6 weeks). Three patients had seropositive active rheumatoid arthritis, four patients had seronegative spondyloarthritis (two reactive arthritis, one psoriatic arthritis, one enteroarthritis) and three patients had crystal induced arthritis (two cases of acute gout and one case of acute pseudogout). Synovial fluids were processed within one hour of aspiration. Two slides were stained with May-Grunwald-Giemsa (MGG) reagent. Reiter cells were counted on the basis of the first 500 cells encountered on MGG stained slides. In addition, two cytocentrifuge monolayer preparations were processed for immunohistochemistry using the monoclonal anti-human-CD36 antibody (Boehringer Mannheim-Germany) diluted to 3.5 mg/ml and the monoclonal antihuman monocyte CD14 antibody (DAKO-Denmark) diluted 1:10 in TRIS-HCL buffer. In brief, specimens were air dried, fixed with acetone and then stored at −70°C until processing. The specimens were incubated for 60 minutes at room temperature with the primary antibody. For the conjugation of peroxidase an En Vision+TM Kit (Dako) was used. The monolayers were then incubated for five minutes with a prediluted diamino-benzidine solution (DAKO) and countercoloured with Mayer's haematoxylin. All incubation steps were preceded by washes in 0.1 M PBS (five minutes × three). The slides were examined at 400 × magnification.

Omission of primary antiserum, use of normal rabbit serum, or one of subsequent steps in the staining method were included as controls for specificity.

Macrophages as well as Reiter cells could be observed on MGG stained slides. Reiter cells were more abundant in synovial fluids from patients with seronegative spondyloarthritides and crystal induced arthritis compared with synovial fluids from RA (table 1). On immunohistochemistry preparations, numerous mononuclear cells showed a CD36 positive reaction, while all the Reiter cells observed displayed a positivity for the thrombospondin receptor. CD14+ mononuclear cells outnumbered CD36+ cells; similarly, all the Reiter cells observed were immunoreactive for the anti-CD14 antibody (fig 1A, 1B).

Table 1

The number of Reiter cells calculated on the first 500 cells encountered on May-Grünwald-Giemsa stained slides

Figure 1

(A) CD36+ mononuclear phagocytes (arrowhead) and Reiter cell (arrow) in cytospin preparation of an inflammatory synovial effusion (enteroarthritis). (B) CD14+ Reiter cell observed in a knee synovial fulid sample from a patient with monoarticular acute gout.

Our findings show that Reiter cells do express both CD36 and CD14 adhesion molecules.

CD36 expression on Reiter cells seems to support the notion of the involvement of this receptor in the clearance of apoptotic PMN during synovial inflammation. In vitro data have shown that thrombospondin receptor and CD14 are some of the most important adhesion molecules involved in cell clearance. The expression of the thrombospondin receptor turns an amateur phagocyte into a professional one.7 It has been hypothesised that dysregulation of this receptor and the ensuing impairment of inflammatory cell elimination could play a part in inducing chronicity as well as tissue damage and scarring.8 Recently, CD14 has been demonstrated to mediate recognition and phagocytosis of apoptotic cells. This interaction depends on a region of CD14 that is supposed to be identical to a region that binds bacterial lipopolysaccharide,5triggering the release of proinflammatory cytokines from macrophages. On the other hand, the interaction with self components acts as an initial step leading to apoptotic cell elimination. A major role for CD36 in the uptake of apoptotic neutrophils has been recently hypothesised, but it seems likely that microenvironmental modifications could promote the switch from a CD36 dependent pathway to pathways using other adhesion molecules such as CD14.9 The removal of inflammatory PMN is mediated by several surface molecules and modulated by microenvironmental modifications; it seems to be a crucial, although only partially understood event for the control and resolution of inflammation. Our results suggest that CD14 and CD36 could be involved in the adhesion of the macrophage to the apoptotic cell, the first step of a process leading to cell clearance. However, as CD14 and CD36 are known to play a part in different biological processes, the demonstration of these multifunctional adhesion molecules on Reiter cells is not a definitive evidence concerning their role for apoptotic cell clearance in the synovial fluid. Additional functional investigations are required to establish the exact role of CD14 and CD36 in the clearance of the PMN in synovial effusions.

Acknowledgments

We thank Dr Nicolo Pipitone for reviewing the manuscript, and Ms Eleonora Franceschini for technical assistance.

References

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