![]() ![]() Further work is planned to investigate the role of the inflammasome in JSLE, using pharmacological interventions with specific known and novel inhibitors of the NLRP3 inflammasome. The NLRP3 inflammasome is thought to be an important mediator in the pathogenesis of certain inflammatory diseases, and flare episodes associated with JSLE. IF was positive for cleaved caspase-1 in ATP treated Mφs and this was confirmed in lysed cells, using western blotting.Ĭonclusion: Overall, the results indicate that Mφs undergo pyroptosis via the NLRP3 inflammasome when challenged with a two-signal approach of priming and ATP, and that cytokines, nuclear debris and DAMPs may not only trigger, but amplify the inflammatory response of this pathway. Furthermore, a greater increase in LDH activity was observed in Mφs that were incubated with JSLE serum and NET material (3.7x10 -3 ± 0.5x10 -3 and 7.6x10 -3 ± 0.5x10 -3, respectively, compared to 0.00 milliunits/mL n = 3-6 p < 0.05). ATP-treated Mφs showed increased LDH activity compared to controls (3.4x10 -3 ± 0.12x10 -3) compared to 0.00 milliunits/mL, respectively n = 3 p < 0.05). ATP-treated Mφs were collected and either assayed for pyroptosis marker, lactate dehydrogenase (LDH) activity, cleaved caspase-1 with immunofluorescence (IF), or lysed for cleaved caspase-1 using western blotting. ![]() Primed Mφs were tested for cell surface markers, HLA-DR, CD282 (TLR2), and CD68 using flow cytometry. ![]() Some Mφs were incubated with 10% JSLE patient sera or NETosis-derived material (10 ng/ml) from PMA-treated neutrophils. Primed Mφs not treated with ATP were used as a negative control. Methods: THP-1 cell line-derived macrophages (Mφs) were primed using lipopolysaccharide (LPS 10 ng/ml) and interferon (IFN) γ (20 ng/ml) in complete media at 37 ☌ for 24 hours, and subsequently treated with 10 mM Adenosine triphosphate (ATP) for 30 min. Objectives: To investigate the role of pyroptotic cell death in JSLE, which occurs following the assembly and activation of the NLRP3 inflammasome, which may be a promising target in the treatment of JSLE and other inflammatory diseases. Impaired clearance of nuclear debris from cells and upregulated circulating damage-associated molecular patterns (DAMPs) and cytokines are thought to trigger cell death and thus, further increase release of pro-inflammatory molecules following the cytosolic assembly of inflammasomes. Introduction: Juvenile-onset Systemic Lupus Erythematosus (JSLE) is a severe autoimmune disease causing organ damage and long-term morbidity. ![]()
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December 2022
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