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Proinflammatory T Cell Polarization Investigated in Early Knee Osteoarthritis

By LabMedica International staff writers
Posted on 01 Feb 2021
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Image: The MACSQuant Analyzer 10 flow cytometer has three lasers (405, 488, 638 nm), two scatter (FSC, SSC) and seven fluorescent channels (Photo courtesy of Miltenyi Biotec)
Image: The MACSQuant Analyzer 10 flow cytometer has three lasers (405, 488, 638 nm), two scatter (FSC, SSC) and seven fluorescent channels (Photo courtesy of Miltenyi Biotec)
In osteoarthritis (OA), the cartilage in the knee joint gradually wears away. As the cartilage wears away, it becomes frayed and rough, and the protective space between the bones decreases. This can result in bone rubbing on bone, and produce painful bone spurs. Osteoarthritis develops slowly and the pain it causes worsens over time.

Assessment of early OA is indispensable in the search for biomarkers as a diagnostic tool. OA pathology has a temporal pattern, and cartilage, bone, and synovial matrix biomarkers show a positive association with the progression of knee OA. Synovial inflammation has been identified as an independent factor significantly contributing to OA pathology.

Orthopedic Specialists at the University Hospital Heidelberg (Heidelberg, Germany) and their colleagues enrolled 40 patients (29 women, 11 men) with arthroscopic or MRI findings of early osteoarthritis of the knee. The mean age of the study population was 41.7 ± 14.3 years. Synovial fluid (SF), synovial membrane (SM), and peripheral blood (PB) were collected at the time of surgery. Mononuclear cells were isolated from heparin anti-coagulated whole blood, SF, and SM cell suspensions using Ficoll-Paque PLUS (GE Healthcare, Waukesha, WI, USA) density gradient centrifugation. T cells were isolated from PB, SF, and SM mononuclear cells by CD3 MACS bead separation (Miltenyi Biotec, Bergisch Gladbach, Germany).

Samples were analyzed by flow cytometry for surface markers and cytokines, which are preferentially expressed by distinct T cell subsets (Th1, Th2, Th17, regulatory T cells). Furthermore, the team analyzed SF and PB supernatants using the Miltenyi Biotec MACSPlex for multiple cytokine expression profiles. Flow analysis was performed using a Miltenyi Biotec MACSQuant Analyzer, which is a 7-channel flow cytometer. Native SF and PB sera were analyzed by the Miltenyi Biotec MACSPlex 12 Kit.

The scientists reported that SF and SM showed a distinct infiltration of CD4+ T lymphocytes, with significantly increased expression of chemokine receptors CXCR3/CCR5, cytokine IFN-γ which is preferentially expressed by Th1 cells, and CD161 which is preferentially expressed by interleukin-17 (IL-17) producing Th17 cells compared to PB. Furthermore, the percentage of CD4+ T cells polarized to regulatory T cells (Treg) was significantly increased in SM compared to SF and PB. No significant differences were observed for CCR3 and CCR4which are preferentially expressed by Th2 cells, although IL-4 values were significantly higher in SM and SF compared to PB. Cytokine analysis showed comparable results between PB and SF, with only IL-6 being significantly increased in SF.

The authors concluded that early OA joints show already significant inflammation through CD4+ T cell infiltration, with predominant Th1 cell polarization. Inflammation seems to be driven by direct proinflammatory cell interaction. Cytokine signaling seems to be negligible at the site of inflammation in early OA, with only IL-6 being significantly increased in SF compared to PB. The study was published on January 22, 2021 in the journal Arthritis Research & Therapy.

Related Links:
University Hospital Heidelberg
GE Healthcare
Miltenyi Biotec

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