Azeliragon ameliorates Alzheimer's disease via the Janus tyrosine kinase and signal transducer and activator of transcription signaling pathway

Abstract

Objectives: TTP488, an antagonist of the receptor for advanced glycation end-products, was evaluated as a potential treatment for patients with mild-to-moderate Alzheimer's disease (AD). However, the mechanism underlying the protective action of TTP488 against AD has not yet been fully explored.

Methods: Healthy male rats were exposed to aberrant amyloid β (Aβ) 1-42. Lipopolysaccharide (LPS) and the NOD-like receptor family pyrin domain containing 1 (NLRP1) overexpression lentivirus were injected to activate the NLRP1 inflammasome and exacerbate AD. TTP488 was administered to reverse AD injury. Finally, tofacitinib and fludarabine were used to inhibit the activity of Janus tyrosine kinase (JAK) and signal transducer and activator of transcription (STAT) to prove the relationship between the JAK/STAT signaling pathway and TTP488.

Results: LPS and NLRP1 overexpression significantly increased the NLRP1 levels, reduced neurological function, and aggravated neuronal damage, as demonstrated by the impact latency time of, time spent by, and length of the platform covered by, the mice in the Morris water maze assay, Nissl staining, and immunofluorescence staining in rats with AD.

Conclusions: TTP488 administration successfully reduced AD injury and reversed the aforementioned processes. Additionally, tofacitinib and fludarabine administration could further reverse AD injury after the TTP488 intervention. These results suggest a new potential mechanism underlying the TTP488-mediated alleviation of AD injury.

Conflict of interest statement

No potential conflict of interest was reported.

Figures

Figure 1. (A) Western blotting assay for detecting the expression of NLRP1 and NeuN. (B, C) Quantification of the expression of NLRP1 and NeuN. (D) NLRP1/NeuN ratio. (E) Immunofluorescence assay for detecting the expression of NLRP1 and NeuN (×400) and (F) NLRP1/NeuN (+) cells in samples from the sham, Aβ1-42, and Aβ1-42+LPS groups. Protein levels were normalized to those of β-actin (Aβ1-42 vs. sham group, **p<0.05; Aβ1-42+LPS vs. Aβ1-4 group, ## p<0.05, n=6 per group. All data were represented as the mean±standard error).

Figure 2. (A) The latency time of, (B) the time spent by, and (C) the length covered by, the mice from the sham, Aβ1-42+Vehicle, Aβ1-42+5 mg/d TTP488, Aβ1-42+20 mg/d TTP488, and Aβ1-42+50 mg/d TTP488 groups in the platform quadrant in the Morris water maze. (D) Immunofluorescence assay for detecting the expression of NLRP1 and NeuN (×400) and (E) NLRP1/NeuN (+) cells. (F) Nissl staining and (G) Nissl (+) cells in samples from the sham, Aβ1-42+Vehicle, and Aβ1-42+20 mg/d TTP488 groups (Aβ1-42+Vehicle vs. Sham group, **p<0.05; Aβ1-42+20 mg/d TTP488 vs. Aβ1-4+Vehicle group, ## p<0.05, n=6 per group).

Figure 3. (A) The latency time of, (B) the time spent by, and (C) the length covered by, the mice in the platform quadrant in the Morris water maze. (D) TUNEL Immunofluorescence assay (×400). (E) TUNEL (+) cells in samples from the sham, Aβ1-42, Aβ1-42+LPS, and Aβ1-42+LPS+TTP488 groups. (F) The latency time, (G) the time, and (H) the length covered in the platform quadrant in the Morris water maze. (I) TUNEL immunofluorescence assay (×400) and (J) TUNEL (+) cells in samples from the sham, Aβ1-42, Aβ1-42+control oeNLRP1, Aβ1-42+oeNLRP1, and Aβ1-42+oeNLRP1+TTP488 groups (Aβ1-42+LPS or Aβ1-42+oeNLRP1 vs. Aβ1-42 group, **p<0.05; Aβ1-42+LPS+TTP488 or Aβ1-42+oeNLRP1+TTP488 vs. Aβ1-42+LPS or Aβ1-42+oeNLRP1 group, ## p<0.05, n=6 per group).

Figure 4. (A) Western blot assay for detecting the expression of caspase-1, IL-1β, and IL-18. (B) Quantification of the expression levels of caspase-1, IL-1β, and IL-18. (C) ELISA assay for quantifying the expression levels of caspase-1, IL-1β, and IL-18 in the sham, Aβ1-42, Aβ1-42+LPS, and Aβ1-42+LPS+TTP488 groups. (D) Western blot assay for detecting the expression of caspase-1, IL-1β, and IL-18. (E) Quantification of the expression levels of caspase-1, IL-1β, and IL-18. (F) ELISA assay for quantifying the expression levels of caspase-1, IL-1β, and IL-18 in the sham, Aβ1-42, Aβ1-42+control oeNLRP1, Aβ1-42+oeNLRP1, and Aβ1-42+oeNLRP1+TTP488 groups. Protein levels were normalized to those of β-actin (Aβ1-42+LPS or Aβ1-42+oeNLRP1 vs. Aβ1-42 group, **p<0.05; Aβ1-42+LPS+TTP488 or Aβ1-42+oeNLRP1+TTP488 vs. Aβ1-42+LPS or Aβ1-42+oeNLRP1 group, ## p<0.05, n=6 per group).

(A) The latency time of, (B) the time spent by, and (C) the length covered by, mice in the platform quadrant in the Morris water maze. (D) Western blot assay for detecting the expression of caspase-1, IL-1β, and IL-18. (E-G) Quantification of the levels of caspase-1, IL-1β, and IL-18. (H-J) ELISA assay for quantifying the expression levels of caspase-1, IL-1β, and IL-18 in the sham, Aβ1-42, Aβ1-42+oeNLRP1, Aβ1-42+oeNLRP1+TTP488, Aβ1-42+oeNLRP1+TTP488+Tofacitinib, and Aβ1-42+oeNLRP1+TTP488+Fludarabine groups. Protein levels were normalized to those of β-actin (Aβ1-42+oeNLRP1 vs. Aβ1-42 group, **p<0.05; Aβ1-42+oeNLRP1+TTP488 vs. Aβ1-42+oeNLRP1 group, ## p<0.05; Aβ1-42+oeNLRP1+TTP488+Tofacitinib or Aβ1-42+oeNLRP1+TTP488+Fludarabine vs. Aβ1-42+oeNLRP1+TTP488 group, && p<0.05, n=6 per group).