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. Author manuscript; available in PMC: 2019 Apr 3.
Published in final edited form as: Nat Biomed Eng. 2018 Apr 16;2(5):279–292. doi: 10.1038/s41551-018-0221-2

Figure 6. TRAF6i-HDL therapy shows no adverse immune effects or toxic effects in mice and non-human primates.

Figure 6.

(a) Apoe−/− mice were intravenously injected with TRAF6i-HDL at 5 mg/kg or PBS (n=5 per group). Serum was collected 24 hours after injection. We found no signs of systemic immune activation. Interleukin 6 (IL-6), tissue necrosis factor α (TNFα), chemokine (C-C motif) ligand 2 (CCL2), interleukin 1β (IL-1β), and serum amyloid P-component (SAP) levels were not increased. Bars represent means and standard errors of the mean. (b) In vitro LPS stimulation test in 3x-κBluc plasmid transfected RAW264.7 cells, showing that TRAF6i-HDL did not affect NFκB activation. Bars represent means and standard errors of the mean. (c) In vitro LPS and FGK45 stimulation test in bone marrow derived macrophages showing that TRAF6i-HDL did not affect CCL2 expression. Bars represent means and standard errors of the mean. (d to f) Six non-human primates were infused with either control (n=3) or 1 mg/kg TRAF6i-HDL (n=3). Blood was collected at multiple time points and the animals were sacrificed 72 hours after infusion.(d) Complete blood counts showed no effects of TRAF6i-HDL therapy on lymphocytes, erythrocytes and platelets. Means and standard deviations at each timepjoint are shown. (e) IL-6, TNFα and CCL2 were not affected by TRAF6i-HDL therapy. Means and standard deviations at each timepjoint are shown. (f) Extensive blood chemistry analysis showed no toxic effects of TRAF6i-HDL infusion on hepatic, renal, pancreatic or muscle cell biomarkers. Lipids, glucose, protein (albumin and globulin) and electrolytes were also unaffected. Bars represent means and standard deviations. (g) Specimens from liver, kidneys and spleen were sectioned and stained (H&E) for histological analysis and evaluated by a pathologist (10x magnification is shown). No signs of tissue damage or disturbances in tissue architecture were found in any of the tissues (single experiment). For all figures P-values were calculated with Mann-Whitney U tests (two sided). HCT : Hematocrit, MCV : Mean corpuscular volume, MCH : Mean corpuscular hemoglobin, MCHC : Mean corpuscular hemoglobin concentration, HGB : Hemoglobin, ALB : Albumin, GLOB : Globulin, BUN : Blood urea nitrogen, LDH : Lactate dehydrogenase.