Research Paper Volume 13, Issue 11 pp 15620—15637

Formulated Chinese medicine Shaoyao Gancao Tang reduces NLRP1 and NLRP3 in Alzheimer’s disease cell and mouse models for neuroprotection and cognitive improvement

Neuroprotective effect of SG-Tang in Aβ-GFP-expressing SH-SY5Y cells inflamed with HMC3 conditioned medium. (A) Experimental flow chart. Aβ-GFP SH-SY5Y cells were plated in medium with retinoic acid (RA, 10 μM) on day 1, and treated with SG-Tang (100 μg/ml) next day for 8 h, followed by doxycycline (Dox, 5 μg/ml) addition to induce Aβ-GFP expression. On day 6, DMEM-F12 medium without retinoic acid addition (− RA) was mixed with HMC3 conditioned medium (CM/±IFN-γ, 1:1 ratio) and added to the cells for 2 days. (B) iNOS, NLRP1, NLRP3, TNF-α, IL-1β and IL-6 levels, (C) ROS production, GFP fluorescence and caspase-1 activity, (D) neurite length, process and branch were assessed on day 8 (n = 3). For normalization, the relative iNOS, NLRP1, NLRP3, TNF-α, IL-1β, IL-6 and ROS levels in uninduced and CM/-IFN-γ stimulated cells were set as 100%. P values: comparisons between induced (Dox+, + CM/-IFN-γ) vs. uninduced (Dox-,+ CM/-IFN-γ) cells (#: P ##: P ###: P &: P &&: P $$: P $$$: P P P P post hoc Tukey test).

Figure 3. Neuroprotective effect of SG-Tang in Aβ-GFP-expressing SH-SY5Y cells inflamed with HMC3 conditioned medium. (A) Experimental flow chart. Aβ-GFP SH-SY5Y cells were plated in medium with retinoic acid (RA, 10 μM) on day 1, and treated with SG-Tang (100 μg/ml) next day for 8 h, followed by doxycycline (Dox, 5 μg/ml) addition to induce Aβ-GFP expression. On day 6, DMEM-F12 medium without retinoic acid addition (− RA) was mixed with HMC3 conditioned medium (CM/±IFN-γ, 1:1 ratio) and added to the cells for 2 days. (B) iNOS, NLRP1, NLRP3, TNF-α, IL-1β and IL-6 levels, (C) ROS production, GFP fluorescence and caspase-1 activity, (D) neurite length, process and branch were assessed on day 8 (n = 3). For normalization, the relative iNOS, NLRP1, NLRP3, TNF-α, IL-1β, IL-6 and ROS levels in uninduced and CM/-IFN-γ stimulated cells were set as 100%. P values: comparisons between induced (Dox+, + CM/-IFN-γ) vs. uninduced (Dox-,+ CM/-IFN-γ) cells (#: P < 0.05, ##: P < 0.01, ###: P < 0.001), induced and inflamed (Dox+, + CM/+IFN-γ) vs. induced (Dox+, + CM/-IFN-γ) cells (&: P < 0.05, &&: P < 0.01), induced and inflamed (Dox+, + CM/+IFN-γ) vs. uninduced (Dox-, + CM/-IFN-γ) cells ($$: P < 0.01, $$$: P < 0.001), or SG-Tang treated (+ CM/+IFN-γ, +SG-Tang) vs. untreated (+ CM/+IFN-γ) cells (*: P < 0.05, **: P < 0.01, ***: P < 0.001). (One-way ANOVA with a post hoc Tukey test).