We are analyzing https://link.springer.com/article/10.1186/s13075-015-0794-z.

Title:
Conditional deletion of caspase-8 in macrophages alters macrophage activation in a RIPK-dependent manner | Arthritis Research & Therapy
Description:
Introduction Although caspase-8 is a well-established initiator of apoptosis and suppressor of necroptosis, recent evidence suggests that this enzyme maintains functions beyond its role in cell death. As cells of the innate immune system, and in particular macrophages, are now at the forefront of autoimmune disease pathogenesis, we examined the potential involvement of caspase-8 within this population. Methods Cre LysM Casp8 fl/fl mice were bred via a cross between Casp8 fl/fl mice and Cre LysM mice, and RIPK3 −/− Cre LysM Casp8 fl/fl mice were generated to assess the contribution of receptor-interacting serine-threonine kinase (RIPK)3. Immunohistochemical and immunofluorescence analyses were used to examine renal damage. Flow cytometric analysis was employed to characterize splenocyte distribution and activation. Cre LysM Casp8 fl/fl mice were treated with either Toll-like receptor (TLR) agonists or oral antibiotics to assess their response to TLR activation or TLR agonist removal. Luminex-based assays and enzyme-linked immunosorbent assays were used to measure cytokine/chemokine and immunoglobulin levels in serum and cytokine levels in cell culture studies. In vitro cell culture was used to assess macrophage response to cell death stimuli, TLR activation, and M1/M2 polarization. Data were compared using the Mann–Whitney U test. Results Loss of caspase-8 expression in macrophages promotes onset of a mild systemic inflammatory disease, which is preventable by the deletion of RIPK3. In vitro cell culture studies reveal that caspase-8–deficient macrophages are prone to a caspase-independent death in response to death receptor ligation; yet, caspase-8–deficient macrophages are not predisposed to unchecked survival, as analysis of mixed bone marrow chimeric mice demonstrates that caspase-8 deficiency does not confer preferential expansion of myeloid populations. Loss of caspase-8 in macrophages dictates the response to TLR activation, as injection of TLR ligands upregulates expression of costimulatory CD86 on the Ly6ChighCD11b+F4/80+ splenic cells, and oral antibiotic treatment to remove microbiota prevents splenomegaly and lymphadenopathy in Cre LysM Casp8 fl/fl mice. Further, caspase-8–deficient macrophages are hyperresponsive to TLR activation and exhibit aberrant M1 macrophage polarization due to RIPK activity. Conclusions These data demonstrate that caspase-8 functions uniquely in macrophages by controlling the response to TLR activation and macrophage polarization in an RIPK-dependent manner.
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Keywords {🔍}

mice, flfl, casp, cre, lysm, caspase, tlr, cells, ripk, bmdms, cell, activation, fig, control, myeloid, deletion, levels, caspasedeficient, pubmed, response, article, data, populations, macrophages, splenic, figure, additional, death, file, numbers, macrophage, compared, expression, usa, google, scholar, aged, increased, cas, analysis, systemic, cdbf, results, fas, nec, disease, bone, vivo, total, studies,

Topics {✒️}

carbobenzoxy-valyl-alanyl-aspartyl-[o-methyl]-fluoromethylketone casp8 fl/fl + z-ietd-fmk bmdms casp8 fl/fl + z-ietd-fmk gro-α growth-regulated oncogene-α wt + casp8 fl/fl mice receptor-interacting serine-threonine kinase keratinocyte chemoattractant/growth-regulated oncogene-α 1β-converting enzyme-inhibitory protein predesigned fluorescein-labeled primer/probes casp8 fl/fl mice antigen-specific t-cell proliferation full size image female casp8 fl/fl autoimmune mrl-lpr/lpr mice cd19+b220+cd138+cd21/35−cd23− z-ietd z-ile-glu cd11b+f4/80+ splenic populations casp8 fl/fl ly6chighcd11b+f4/80+ splenic population regulatory t-cell numbers periodic acid–schiff stain cd4−cd8−cd3+b220+ splenic cd11b+f4/80+ly6chigh produce m-csf-stimulated bmdms macrophage colony-stimulating factor cd11cintermediatepdca-1+b220+ plasmacytoid dcs cre lysm mice ly6clow cd11b+f4/80+ cells fluorescence-activated cell sorting cd4+ t-cell proliferation facs-sorted splenic neutrophils 1/ot-ii/rag −/− mice cd8+ t-cell numbers cd11b+f4/80−ly6g+ neutrophils ova-specific splenic cd4+ maintains cell type–specific caspase-8–deficient splenic populations ly6chighcd11b+f4/80+ splenic cells caspase-8–deficient bmdm cultures myeloid cell–specific caspase-8 reduced il-12/il-23p40 transcription 1/ot-ii/rag −/− cd4+ purify antigen-presenting cells defective signal-transduction pathways ly6chigh cd11b+f4/80+ cells enzyme-linked immunosorbent assays cell death–independent activities goat anti-rat immunogloblulin myeloid cell–specific deletion article download pdf

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         description:Although caspase-8 is a well-established initiator of apoptosis and suppressor of necroptosis, recent evidence suggests that this enzyme maintains functions beyond its role in cell death. As cells of the innate immune system, and in particular macrophages, are now at the forefront of autoimmune disease pathogenesis, we examined the potential involvement of caspase-8 within this population. Cre LysM Casp8 fl/fl mice were bred via a cross between Casp8 fl/fl mice and Cre LysM mice, and RIPK3 −/− Cre LysM Casp8 fl/fl mice were generated to assess the contribution of receptor-interacting serine-threonine kinase (RIPK)3. Immunohistochemical and immunofluorescence analyses were used to examine renal damage. Flow cytometric analysis was employed to characterize splenocyte distribution and activation. Cre LysM Casp8 fl/fl mice were treated with either Toll-like receptor (TLR) agonists or oral antibiotics to assess their response to TLR activation or TLR agonist removal. Luminex-based assays and enzyme-linked immunosorbent assays were used to measure cytokine/chemokine and immunoglobulin levels in serum and cytokine levels in cell culture studies. In vitro cell culture was used to assess macrophage response to cell death stimuli, TLR activation, and M1/M2 polarization. Data were compared using the Mann–Whitney U test. Loss of caspase-8 expression in macrophages promotes onset of a mild systemic inflammatory disease, which is preventable by the deletion of RIPK3. In vitro cell culture studies reveal that caspase-8–deficient macrophages are prone to a caspase-independent death in response to death receptor ligation; yet, caspase-8–deficient macrophages are not predisposed to unchecked survival, as analysis of mixed bone marrow chimeric mice demonstrates that caspase-8 deficiency does not confer preferential expansion of myeloid populations. Loss of caspase-8 in macrophages dictates the response to TLR activation, as injection of TLR ligands upregulates expression of costimulatory CD86 on the Ly6ChighCD11b+F4/80+ splenic cells, and oral antibiotic treatment to remove microbiota prevents splenomegaly and lymphadenopathy in Cre LysM Casp8 fl/fl mice. Further, caspase-8–deficient macrophages are hyperresponsive to TLR activation and exhibit aberrant M1 macrophage polarization due to RIPK activity. These data demonstrate that caspase-8 functions uniquely in macrophages by controlling the response to TLR activation and macrophage polarization in an RIPK-dependent manner.
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         Imiquimod
         Macrophage Polarization
         TLR9 Ligation
         Oral Antibiotic Treatment
         Rheumatology
         Orthopedics
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