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Related Experiment Videos

Tumor necrosis factor: specific binding and internalization in sensitive and resistant cells.

M Tsujimoto, Y K Yip, J Vilcek

    Proceedings of the National Academy of Sciences of the United States of America
    |November 1, 1985
    PubMed
    Summary

    Recombinant human tumor necrosis factor (TNF) binds to high-affinity receptors on both sensitive and resistant cells. Resistance to TNF cytotoxicity is not due to receptor issues but likely other cellular mechanisms.

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    Area of Science:

    • Immunology
    • Cell Biology
    • Molecular Biology

    Background:

    • Tumor Necrosis Factor (TNF) is a key cytokine involved in inflammation and immunity.
    • Understanding TNF receptor interactions is crucial for developing targeted therapies.
    • Cellular resistance to TNF's cytotoxic effects requires investigation into its binding and processing.

    Purpose of the Study:

    • To investigate the receptor binding, internalization, and degradation of recombinant human TNF.
    • To compare TNF interactions in cells sensitive versus resistant to its cytotoxic action.
    • To elucidate the mechanisms underlying cellular resistance to TNF.

    Main Methods:

    • Radiolabeling of recombinant human TNF with 125I.
    • Incubation with murine L929 (sensitive) and human FS-4 (resistant) cells.

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  • Scatchard analysis to quantify receptor binding.
  • Assessment of TNF internalization and degradation at 37°C.
  • Inhibition studies using chloroquine.

    • Main Results:

    • 125I-labeled TNF specifically bound to high-affinity receptors on both L929 and FS-4 cells.
    • FS-4 cells exhibited more TNF binding sites (7500) than L929 cells (2200).
    • TNF was rapidly internalized and degraded in both cell types, with degradation inhibited by chloroquine, suggesting lysosomal involvement.

    Conclusions:

    • TNF resistance in FS-4 cells is not attributed to a lack of TNF receptors or impaired internalization/degradation pathways.
    • Both sensitive and resistant cells possess functional TNF receptors and can process the cytokine.
    • The basis for TNF resistance likely lies in downstream signaling events rather than initial receptor interaction or degradation.