Tumor necrosis factor-alpha promotes survival in methotrexate-exposed macrophages by an NF-kappaB-dependent pathway

Abstract

IntroductionMethotrexate (MTX) induces macrophage apoptosis in vitro, but there is not much evidence for increased synovial macrophage apoptosis in MTX-treated patients. Macrophage apoptosis is reported, however, during clinical response to anti-tumor necrosis factor-alpha (TNF-α) treatments. This implies that TNF-α promotes macrophage survival and suggests that TNF-α may protect against MTX-induced apoptosis. We, therefore, investigated this proposal and the macrophage signaling pathways underlying it.MethodsCaspase-3 activity, annexin-V binding/7-aminoactinomycin D (7-AAD) exclusion and cell-cycle analysis were used to measure steps in apoptosis of primary murine macrophages and cells of the RAW264.7 macrophage cell line that had been exposed to clinically-relevant concentrations of MTX and TNF-α.ResultsMTX induces apoptosis in primary murine macrophages at concentrations as low as 100 nM in vitro. TNF-α, which has a context-dependent ability to increase or to suppress apoptosis, efficiently suppresses MTX-induced macrophage apoptosis. This depends on NF-κB signaling, initiated through TNF Receptor Type 1 ligation. Macrophage colony stimulating factor, the primary macrophage survival and differentiation factor, does not activate NF-κB or protect macrophages from MTX-induced apoptosis. A weak NF-κB activator, Receptor Activator of NF-κB Ligand (RANKL) is likewise ineffective. Blocking NF-κB in TNF-α-exposed macrophages allowed pro-apoptotic actions of TNF-α to dominate, even in the absence of MTX. MTX itself does not promote apoptosis through interference with NF-κB signaling.ConclusionsThese findings provide another mechanism by which TNF-α sustains macrophage numbers in inflamed tissue and identify a further point of clinical complementarity between MTX and anti-TNF-α treatments for rheumatoid arthritis.

Involvement of calmodulin and calmodulin kinase II in tumor necrosis factor alpha-induced survival of bone marrow derived macrophages

“…Contrarily to the actions of this cytokine in TDMs, in bone marrow-derived macrophages (BMDMs) TNFα exerts an unambiguous pro-survival effect [7; 8] (see also Discussion). Yet, similar to TDMs, survival signaling in BMDMs also exhibits an obligatory requirement for constitutive Ca 2+ influx [11] (and see Supplementary figure I).…”

“…In TDMs however, TNFα-induced survival signaling is the result of a compensatory response of the macrophages against the pro-apoptotic actions of the cytokine. In more recent work using murine bone marrow-derived macrophages (BMDMs), in which TNFα exerts an unambiguous pro-survival effect [7; 8], we found, once again, that a Ca 2+ influx dependent mechanism exists and that TRPC3, a member of the TRPC family of Ca 2+ -permeable cation channels [9; 10], is the channel responsible for mediating constitutive Ca 2+ influx in these cells [11]. However, if a CAM/CAMKII-dependent mechanism also operates in BMDMs remains to be determined.…”

Expression of 4-1Bb and Its Ligand on Blasts Correlates with Prognosis of Patients with Aml

“…In chronic lymphatic leukemia (CLL), GITR expression is also documented, shown to induce TNF production, and considered a survival factor when stimulated 12. Studies suggest that GITR is also expressed on myeloid cells such as macrophages with TNF induction13 promoting macrophage-related survival 14. GITR stimulation in mice suffering from carcinoma leads to improved overall survival (OS) due to enhanced host-related immune reactivity 15.…”

Prospects of Apoptotic Cell-Based Therapies for Transplantation and Inflammatory Diseases

“…The interactions of such therapy with other treatments received by patients; for example, steroids (which increase apoptotic cell removal by macrophages [128,129]) or methotrexate (which induces macrophage apoptosis [130] and in theory may delay apoptotic cell removal); The possible autoimmunity due to delayed or impaired apoptotic cell clearance;…”