https://orcid.org/0000-0002-4923-1964
Figures & data
Figure 1. Effects of SA and SA+Pro interventions on maternal and neonatal rats.
(a) Schematic representation of the animal study. (b) Effects of SA and SA+Pro interventions on oligosaccharide and free SA levels in rat milk. (c) The β-diversity of milk microbiota represented using NMDS and the relative abundances of bacterial genera in rat milk after intervention with SA and SA+Pro. (d) The relative microbial abundances in the intestinal microbiota and the SCFA compositions in the feces of maternal rats. (e) The abundance of intestinal bacterial genera in neonatal rats. →n was used in figures to differentiate the neonatal groups from the maternal groups; The relative abundances of Lactobacillus spp., L. reuteri, and total and individual SCFA contents were compared among the different groups of neonatal rats. (f) A schematic representation of the OVA-immunization procedure used with neonatal rats fed by different mothers, as well as the serum levels of OVA-specific IgG (OVA-IgG) in neonatal rats before and after OVA-immunization. (g) The levels of sIgA and cytokines in the serum and feces of neonatal rats. (h) Bubble plots representing the results of GO enrichment analysis based on differentially expressed genes in splenic lymphocytes from L. reuteri-treated rats compared with those in untreated lymphocytes. All values are presented as the mean ± SD (n ≥ 5).
Figure 2. SA+Pro intervention in lactating St6gal1± (H) rats compromised allergic responses in neonates by regulating 6′-SL synthesis and the neonatal gut microbiota.
(a) A schematic diagram depicting the cross-feeding procedure used with WT neonates fed by WT, H, and KO maternal rats. (b) Comparison of the major milk oligosaccharides in different rat mothers. (c) Changes in the intestinal microbiota of neonates fed by different maternal rats. (d) The serum levels of cytokines in neonatal rats and changes in lymphocyte subsets in the spleen and neonatal MLNs. (e) A schematic diagram indicating that neonates fed by H maternal rats or H maternal rats received SA+Pro intervention. (f) Changes in the major milk oligosaccharides of the H maternal rats, with or without SA+Pro intervention. (g) The abundance of intestinal bacterial genera and the total and individual SCFA contents in neonatal rats. (h) A schematic diagram representing the establishment of a model of OVA-induced allergic responses in neonates fed by H maternal rats, with or without SA+Pro intervention. (i) Differences in the anal temperatures of neonatal rats following OVA challenge. (j) Hematoxylin and eosin staining and histopathological scores of the intestines in the offspring. (k) Changes in serum anaphylaxis-related cytokine levels, expressed as a percentage of the lymphocyte subpopulations in the spleen and MLNs of offspring. All values are presented as the mean ± SD (n ≥ 3).
Figure 3. The Gpr41-PI3K-Akt-PPAR pathway was involved in regulating 6′-SL biosynthesis in mammary glands by SA+Pro.
(a) Bubble plots representing KEGG enrichment analysis of differentially expressed genes related to signaling pathways in the mammary glands of different groups. The CON group indicated the mammary glands of the WT maternal mice, the H groups indicated the mammary glands of the H maternal mice, and the SA and SA+Pro groups indicated the mammary glands of the maternal mice that received SA and SA+Pro intervention during lactation, respectively. (b) Bubble plots represent GO enrichment analysis of differentially expressed genes in the mammary glands of different groups. (c) Expression of glycosyltransferases in the mammary glands of rats in different groups. (d) Effects of different treatments on the expression of glycosyltransferases in human mammary epithelial cells (MCF-10A). (e) The expression and phosphorylation levels of proteins involved in the PI3K-AKT-mTOR and PPARγ pathways were determined using western blot analysis. GAPDH expression was detected as an internal control. The protein expression levels were quantified using densitometry. (f) Effects of PPARγ and PI3K-AKT inhibitors on ST6Gal-1 expression in MCF-10A cells stimulated with But. (g) ChIP assay results depicting the interaction between the RXRα–PPARγ complex and the promoter region of St6gal1. All values are presented as the mean ± SD (n ≥ 3), ***p < .001.
Table 1. Characteristics of the study participants.
Figure 4. Dynamics of the major HMO fractions and gut microbiota in participants.
(a) The contents of the major HMO fractions in the breast milk of each participant (left) and the average HMO levels (right) in each intervention group during the intervention period. *Mothers who used Abx before giving birth or postpartum. ^Non-secretor mothers. (b) Trends of the major HMOs in the milk of participants who received different interventions. (c) Comparison of the contents of all HMOs, F-HMOs, S-HMOs, and other HMOs (neither fucosylated nor sialylated) in the major HMOs fractions in all milk samples from the M1 and M2 groups at three time points. (d) The contents of all HMOs, F-HMOs, S-HMOs, and other HMOs (neither fucosylated nor sialylated) in the milk of secretory and non-secretory VB and CS mothers in the M1 or M2 groups. (e, f) The bacterial α-diversity (indicated using the Shannon index) and the relative abundances of intestinal microbial genera of mothers in each group before and after the intervention. (g, h) The α-diversity of the intestinal microbiota of mothers (indicated using the Shannon index) and the relative abundances of bacterial genera in the gut of secretory and non-secretory VB and CS mothers in the M1 and M2 groups. All values are presented as the mean ± SD. The study size is indicated in the panels. Multiple comparisons were performed by using one-way ANOVA. All values are presented as the mean ± SD (n ≥ 20).
Table 2. Structures of the major HMOs.
Figure 5. Changes in the intestinal microbiota of infants fed by different mothers.
(a, b) The bacterial α-diversity (indicated using the Shannon index) and the relative abundances of the intestinal microbial genera in infants fed by different mothers before and after the intervention. (c) The dominant microbial communities that exhibited significant differences between the groups of infants fed by different mothers were analyzed using LEfSe. The values among groups were compared by Kruskal-Wallis rank sum test. (d) The relative abundances of intestinal Bifidobacterium spp. in the infants of N1 and N2 groups. (e) The relative abundances of bacterial genera in infants who were exclusively or not exclusively breastfed. (f, g) Effects of maternal Abx use and the delivery mode on the α-diversity and relative abundances of intestinal microbiota in infants. All values are presented as the mean ± SD (n ≥ 6). The study size is indicated in the panels.
Figure 6. Analysis of data from the 20 paired mothers and infants.
(a, b) The contents of all HMOs, F-HMOs, S-HMOs, and other HMOs and the relative abundances (%) of HMOs in milk samples from 20 mothers in each group. (c) Comparison of the major HMOs fractions in milk samples from 20 mothers in each group. Multiple comparisons were performed by using one-way ANOVA. (d) A line chart depicting the content trends of all HMOs and the major S-HMOs in milk samples from both groups. (e, f) The α-diversity (indicated using the Shannon index) and longitudinal changes in the mean relative abundances of bacterial genera in 20 mother – infant pairs. The participating infants were further divided into groups fed by mothers in the M1 and M2 groups who did or did not use Abx before giving birth or postpartum. Thus, the groups studied included the N1 and N1 Abx groups (infants), the M1 Abx and M2 Abx groups (mothers who received Abx before or after giving birth), and the N1 and N1 Abx group (infants), as displayed under the histogram. (g) The β-diversity of the intestinal microbiota in the infants was analyzed using NMDS. (h) Changing patterns in the relative abundances of the major intestinal bacterial genera in the infants of different groups. (i) The bacterial groups that exhibited significant differences among infants from different groups were analyzed using LEfSe. The values among groups were compared by Kruskal-Wallis rank sum test. (j) Correlations between maternal HMO levels and the gut microbial genera of infants before and after intervention in the mothers. +p < .05, *p ≤ .01, ￥p ≤ .001, #p ≤ .0001. All values are presented as the mean ± SD (n = 20). The study size is indicated in the panels.
Table 3. Primers used for q-PCR detection of glycosyltransferase genes expressed in mammary epithelial cells.
Supplemental material