Naotaifang III Protects Against Cerebral Ischemia Injury Through LPS/TLR4 Signaling Pathway in the Microbiota–Gut–Brain Axis

Figures & data

Table 1 Drug Composition of Naotaifang III

Chinese Medicine	Family	Weight (g)
Huang Qi (Astragali Radix)	Legume	30
Ge Gen (Puerariae Lobatae Radix)	Legume	24
Di Long (Pheretima)	Euphorbiidae	12
Chuan Xiong (Chuanxiong Rhizoma)	Apiaceae	12
Jiang Can (Bombyx Batryticatus)	Saturniidae	9
Huang Qin (Scutellariae Radix)	Lamiaceae	12
Da Huang (Rhei Radix Et Rhizoma)	Polygonaceae	15
Pu Gongyin (Taraxaci Herba)	Compositae	12
Gan Jiang (Zingibers Rhizoma)	Zingiberaceae	12
Gan Cao (Glycyrrhiza Radix Et Rhizoma)	Legume	3

Figure 1 Establishment of the intestinal dysbacteriosis rat model.

Figure 2 Treatment of each group.

Figure 3 Quality control of Naotaifang III.

Notes: (a) Astragaloside_IV; (b) Emodin; (c) Ferulic_acid; (d) Puerarin.

Figure 4 Effects of intestinal microbiota disorder on neurological function score and cerebral infarction volume at different time after cerebral ischemia.

Notes: (a) Neurological function score; (b and c) Cerebral infarction volume. n = 18. **P < 0.01 vs N-M.

Figure 5 Effects of Naotaifang III on the cerebral injury.

Notes: (a) Neurological function score (n = 15); (b and c) Cerebral infarction volume (n = 3); (d) HE staining (×400, n = 3); (e) Golgi staining (× 400 and × 1000, n = 3). The values are the mean ± SEM. **P < 0.01 vs N-S; ^#P < 0.05 vs N-M; ^##P < 0.01 vs N-M; ^ΔP < 0.05 vs D-M; ^ΔΔP < 0.01 vs D-M.

Figure 6 Effects of Naotaifang III on LPS in blood plasma and brain cortex, IL-1β in brain cortex.

Notes: (a) Levels of LPS in blood plasma; (b) Levels of LPS in brain cortex; (c) Levels of IL-1β in brain cortex. The values are the mean ± SEM. n = 6. *P < 0.05 vs N-S; **P < 0.01 vs N-S; ^#P < 0.05 vs N-M; ^ΔP < 0.05 vs D-M; ^ΔΔP < 0.01 vs D-M.

Figure 7 Effects of Naotaifang III on TLR4 and NF-κB expression in brain cortex (immunofluorescence staining, 400×).

Notes: (a) Immunofluorescence staining of TLR4; (b) Immunofluorescence staining of NF-κB; The values are the mean ± SEM. n = 3. **P < 0.01 vs N-S; ^##P < 0.01 vs N-M; ^ΔΔP < 0.01 vs D-M.

Figure 8 Effects of Naotaifang III on TLR4 and NF-κB expression in brain cortex (Western blot).

Notes: The values are the mean ± SEM. n = 3. *P < 0.05 vs N-S; ^#P < 0.05 vs N-M; ^##P < 0.01 vs N-M; ^ΔP < 0.05 vs D-M.

Figure 9 Effects of Naotaifang III on intestinal microbiota diversity.

Notes: (a) VENN and Upset plot of OTU levels; (b) Intestinal microbiota alpha diversity parameters. (c) Rarefaction curves. (d) PCoA, NMDS, and distance thermal graph. Data are expressed as the mean ± SD, n = 5. ^#P < 0.05 vs N-M; ^ΔΔP < 0.01 vs D-M.

Figure 10 Effects of Naotaifang III on intestinal microbiota structure.

Notes: (a) Heat map of species composition at phylum level; (b) Stack map of species at class level. (c) Ternary phase diagram of species at genus level.

Figure 11 Effects of Naotaifang III on intestinal microbiota function.

Notes: (a) PICRUSt function prediction chart; (b) Differential functional test analysis.