The consumption of lard oil during pregnancy and postpartum periods has negative effects on cognitive function by altering the fatty acid profile and activating neuroinflammation via calcium signaling pathway in the maternal mice brain

Abstract

It has been suggested that dietary intake of lipids and fatty acids may influence cognitive function, however, the effect of lard intake during pregnancy and postpartum periods on cognitive function of mother remains to be elucidated. We investigated the effect and mechanism of consuming soybean oil (SO), the mixed oil of lard and soybean oil at the ratio of 1:1 (LS) and lard oil (LO) during the pregnancy and postpartum periods on cognitive function of the maternal mice. All pregnant C57BL/6JNifdc mice were fed with soybean oil diet during day 0-10 (the day when vaginal plugs appeared in female mice was recorded as day 0), and then randomly assigned to SO, LS and LO groups (n = 10) from day 11 to day 44. The time in center zone and the number of times to enter in center zone were significantly higher in the SO group than in the LO group detected by the open-field test. The levels of neuroglial cells, NOD-like receptor family pyrin domain containing 3 (NLRP3) inflammasome complex and pyroptosis related proteins in brain of the LO group were significantly higher than those in the SO group. RNA-sequencing results showed that the calcium signaling pathway related genes in brain, including Adcy8, Ntsr1, Trhr, Oxtr, Htr5b and Camk2d levels significantly higher in the LO group than in the SO group. Lipidomic analysis indicated that PG 18:2_18:2, PG 20:5_22:6, and CL 12:0_16:0_22:3_22:5 of glycerophospholipid metabolism in brain significantly connected with Htr5b of calcium signaling pathway. In conclusion, the intake of lard during the pregnancy and postpartum periods is detrimental to the cognitive function of maternal mice, which probably due to changes in the composition of fatty acid in the brain, thereby activating neuroinflammation via calcium signaling pathway in brain.

Keywords: Lard; Neuroinflammation; Omega-3 fatty acid; Postpartum cognitive function.

Graphical abstract

Fig. 1

Effect of dietary oils on cognitive function in the maternal mice. (A) The distance traveled (cm), (B) time in zone (seconds)-center, (C) entries in zone-center, the representative traveled paths of the (D) SO group, (E) LS group and (F) LO group mice in the open-field test. (G) The Y-maze alternation triplet, (H) total arm entries, and (I) alternation triplet (%) in Y-maze test. (J) The escape latency (s) of visible platform trial, hidden platform trial and probe test, (K) entries in zone-platform, and (L) time in zone (%)-quadrant platform of probe test in Morris water maze test. Data are presented as mean ± SD (n = 8). * (P < 0.05) and ** (P < 0.01) indicate the significant difference between groups. SO, soybean oil; LS, mixed oil of lard oil and soybean oil at the ratio of 1:1; LO, lard oil.

Fig. 2

Effect of dietary oils on histopathology and the levels of inflammatory cytokines in the maternal mice brain. (A) The H&E staining diagrams of cortex and hippocampus in maternal mice (scale bar = 20 μm). (B) The percent of normal neurons (%) per field in maternal mice determined by the H&E staining. (C) The Nissl staining diagrams of cortex and hippocampus in maternal mice (scale bar = 20 μm). (D) The percent of normal neurons (%) per field in maternal mice determined by the Nissl staining. (E) The levels of TNF-α, (F) IL-6, (G) IL-1β and (H) IL-18 in maternal mice brain (pg/g). Data are presented as mean ± SD (n = 3). * (P < 0.05) and ** (P < 0.01) indicate the significant difference between groups. SO, soybean oil; LS, mixed oil of lard oil and soybean oil at the ratio of 1:1; LO, lard oil.

Fig. 3

Effect of dietary oils on the activation of neuroglial cells in the maternal mice brain. (A) The GFAP expression of cortex and hippocampus determined by immunofluorescence (scale bar = 20 μm). (B) The GFAP expression of brain determined by western blotting as SO, LO, and LS (n = 3). (C) The IBA1 expression of cortex and hippocampus determined by immunofluorescence (scale bar = 20 μm). (D) The IBA1 expression of brain determined by western blotting as SO, LO, and LS (n = 3). Data are presented as mean ± SD. * (P < 0.05) and ** (P < 0.01) indicate the significant difference between groups. SO, soybean oil; LS, mixed oil of lard oil and soybean oil at the ratio of 1:1; LO, lard oil.

Fig. 4

Effect of dietary oils on the expression of NLRP3 inflammasome complex and pyroptosis related proteins in the maternal mice brain. (A) The representative bands of NLRP3 inflammasome complex and pyroptosis related proteins detected by western blotting as SO, LO, and LS. The relative proteins levels of (B) NLRP3, (C) ASC, (D) Cleaved-Caspase 1, (E) GSDMD, and (F) GSDMD-N. Data are presented as mean ± SD (n = 3). * (P < 0.05) and ** (P < 0.01) indicate the significant difference between groups. SO, soybean oil; LS, mixed oil of lard oil and soybean oil at the ratio of 1:1; LO, lard oil.

Fig. 5

The genes expression related to calcium signaling pathway in the maternal mice brain. (A) The relative FPKM level of genes detected by RNA-sequencing. (B) The relative mRNA expression levels of genes detected by quantitative real time PCR. Data are presented as mean ± SD (n = 3). * (P < 0.05) and ** (P < 0.01) indicate the significant difference compared with LO group. SO, soybean oil; LS, mixed oil of lard oil and soybean oil at the ratio of 1:1; LO, lard oil.

Fig. 6

Effect of dietary oils on fatty acid profile in the brain and its correlation with open-field test data. (A) The proportion of fatty acid with different saturations (%), (B) the proportion of fatty acid monomers (%) and (C) the ratio of n-3/n-6 PUFA in the maternal mice. Person's correlation of (D) SFA (% of total fatty acids), (E) DHA (% of total fatty acids), (F) n-3 PUFA (% of total fatty acids), and (G) the ratio of n-3/n-6 PUFA with time in center (seconds) of the open-field test. Person's correlation of (H) SFA (% of total fatty acids), (I) DHA (% of total fatty acids), (J) n-3 PUFA (% of total fatty acids) and (K) the ratio of n-3/n-6 PUFA with entries in center (times) of the open-field test. Data are presented as mean ± SD (n = 6). * (P < 0.05) and ** (P < 0.01) indicate the significant difference between groups. SO, soybean oil; LS, mixed oil of lard oil and soybean oil at the ratio of 1:1; LO, lard oil.

Fig. 7

Network analysis between significantly differentially expressed lipid molecules and differentially expressed genes of calcium signaling pathway in (A) LO vs SO and (B) LO vs LS. SO, soybean oil; LS, mixed oil of lard oil and soybean oil at the ratio of 1:1; LO, lard oil.

Fig. 8

Pearson correlation analysis between significantly differentially expressed lipid molecules of glycerophospholipid metabolism and differentially expressed genes of calcium signaling pathway in (A) LO vs SO and (B) LO vs LS. * (P < 0.05), ** (P < 0.01) and *** (P < 0.001) indicate the significant difference between groups.