Methods for Evaluation of TNF-α Inhibition Effect

Tumor necrosis factor superfamily (TNFSF) ligands and receptors have distinctive structural characters that link them to cell growth, cell survival, or cell death. Some of these can activate both inflammatory and apoptotic pathways, depending on target cell types and other extrinsic stimuli. Many of the TNF receptor superfamily molecules are expressed in cells of the immune system, which may be central to autoimmune and inflammatory diseases as well as cancer. However, the function of TNFSF members is not just restricted to immune cells. Members of TNFSF have been linked to an array of pathophysiologies, including cancer, neurologic, cardiovascular, pulmonary, autoimmune, and metabolic diseases. TNF-α of TNFSF is a pro-inflammatory cytokine produced by macrophages/monocytes, widely implicated in the pathogenesis of inflammatory disorders. In view of these facts, TNF-α has been recommended as an important target for discovering drugs for autoimmune and inflammatory diseases and cancer. Various cell-based assays to understand the role of TNF-α in inflammation and to estimate the concentrations of TNF-α levels in body fluids such as plasma, synovium, etc., are being followed by researchers. In this chapter, methods of cell viability assay, ELISA assay, RT-PCR, and western blot analysis for estimating LPS-induced TNF-α protein expressions are described in detail.