Microscopic measurement of inflammation in synovial tissue: inter-observer agreement for manual quantitative, semiquantitative and computerised digital image analysis

Objectives

To evaluate inter‐observer agreement for microscopic measurement of inflammation in synovial tissue using manual quantitative, semiquantitative and computerised digital image analysis.

Methods

Paired serial sections of synovial tissue, obtained at arthroscopic biopsy of the knee from patients with rheumatoid arthritis (RA), were stained immunohistochemically for T lymphocyte (CD3) and macrophage (CD68) markers. Manual quantitative and semiquantitative scores for sub‐lining layer CD3+ and CD68+ cell infiltration were independently derived in 6 international centres. Three centres derived scores using computerised digital image analysis. Inter‐observer agreement was evaluated using Spearman''s Rho and intraclass correlation coefficients (ICCs).

Results

Paired tissue sections from 12 patients were selected for evaluation. Satisfactory inter‐observer agreement was demonstrated for all 3 methods of analysis. Using manual methods, ICCs for measurement of CD3+ and CD68+ cell infiltration were 0.73 and 0.73 for quantitative analysis and 0.83 and 0.78 for semiquantitative analysis, respectively. Corresponding ICCs of 0.79 and 0.58 were observed for the use of digital image analysis. All ICCs were significant at levels of p<0.0001. At each participating centre, use of computerised image analysis produced results that correlated strongly and significantly with those obtained using manual measurement.

Conclusion

Strong inter‐observer agreement was demonstrated for microscopic measurement of synovial inflammation in RA using manual quantitative, semiquantitative and computerised digital methods of analysis. This further supports the development of these methods as outcome measures in RA.Microscopic measurement of inflammation in synovial tissue is employed globally by centres working in the field of arthritis research.¹ Adequate and comparable synovial tissue can be safely obtained using blind‐needle biopsy or rheumatological arthroscopy.^2,3,4 In the acquired samples, various parameters may be examined, including cell populations, vascularity, cytokines and adhesion molecules. In rheumatoid arthritis (RA), many of these have been found to relate to disease activity, severity, outcome, and to exhibit a change after treatment with corticosteroids, disease‐modifying antirheumatic drugs (DMARDs) and biological therapy.^{5,6,7,8,9,10,11,12,13,14,15}Several analysis techniques have been employed to measure these parameters. Semiquantitative analysis is a relatively quick method and therefore may facilitate examining large quantities of tissue.⁷ Quantitative analysis is time‐consuming but more sensitive than semiquantitative scoring to change in individual patients.¹⁶ It has been shown in previous studies that these methods can reflect overall joint inflammation when applied to relatively limited amounts of synovial tissue, even though inflammation may differ widely between individual sites in a single joint.^17,18,19 Computerised digital image analysis has been applied more recently in this area and has been shown to correlate well with conventional methods of measurement.^20,21,22This multi‐centre study was undertaken to standardise and validate the methods mentioned previously by evaluating inter‐observer agreement between multiple examiners in the measurement of selected parameters of inflammation in RA synovial tissue by manual quantitative, semiquantitative and computerised image analysis.