Human milk secretory immunoglobulin a and lactoferrin N-glycans are altered in women with gestational diabetes mellitus

Abstract

Very little is known about the effects of gestational diabetes mellitus (GDM) on lactation and milk components. Recent reports suggested that hyperglycemia during pregnancy was associated with altered breast milk immune factors. Human milk oligosaccharides (HMOs) and N-glycans of milk immune-modulatory proteins are implicated in modulation of infant immunity. The objective of the current study was to evaluate the effect of GDM on HMO and protein-conjugated glycan profiles in breast milk. Milk was collected at 2 wk postpartum from women diagnosed with (n = 8) or without (n = 16) GDM at week 24-28 in pregnancy. Milk was analyzed for HMO abundances, protein concentrations, and N-glycan abundances of lactoferrin and secretory immunoglobulin A (sIgA). HMOs and N-glycans were analyzed by mass spectrometry and milk lactoferrin and sIgA concentrations were analyzed by the Bradford assay. The data were analyzed using multivariate modeling confirmed with univariate statistics to determine differences between milk of women with compared with women without GDM. There were no differences in HMOs between milk from women with vs. without GDM. Milk from women with GDM compared with those without GDM was 63.6% lower in sIgA protein (P < 0.05), 45% higher in lactoferrin total N-glycans (P < 0.0001), 36-72% higher in lactoferrin fucose and sialic acid N-glycans (P < 0.01), and 32-43% lower in sIgA total, mannose, fucose, and sialic acid N-glycans (P < 0.05). GDM did not alter breast milk free oligosaccharide abundances but decreased total protein and glycosylation of sIgA and increased glycosylation of lactoferrin in transitional milk. The results suggest that maternal glucose dysregulation during pregnancy has lasting consequences that may influence the innate immune protective functions of breast milk.

Conflict of interest statement

Author disclosures: J. T. Smilowitz, S. M. Totten, J. Huang, D. Grapov, H. A. Durham, C. J. Lammi-Keefe, C. Lebrilla, and J. B. German, no conflicts of interest.

Figures

FIGURE 1

(A) O-PLS-DA from the final model based on the 12 selected variables demonstrate clear visual separation between women with and without GDM. (B) O-PLS-DA variable loadings for the LV1 display the relative levels of milk N-glycans from women with vs. without GDM. CH, complex hybrid; F, fucose; FS, fucose and sialic acid; GDM, gestational diabetes mellitus; HM, high mannose; LF, lactoferrin; LV, latent variable; O-PLS-DA, orthogonal signal corrected partial least-squares discriminant analysis; S, sialic acid; sIgA, secretory IgA; 5411, N-glycan containing 5 hexoses, 4 N-acetyl-hexosamines, 1 fucose, 1 sialic acid.

FIGURE 2

Concentrations of human milk lactoferrin (LF) and sIgA from women with compared with those without GDM. Means ± SEMs for human milk LF and sIgA concentrations from 500 μL of milk from women with GDM (n = 8) vs. non-GDM (n = 16). ANOVA, different from non-GDM, *P < 0.05. GDM, gestational diabetes mellitus; LF, lactoferrin; sIgA, secretory IgA.

FIGURE 3

Correlation network among milk glycans from women with, compared with those without, GDM selected by O-PLS-DA. Using Spearman’s ρ, P < 0.001, relations and strength of the relations (thickness of the lines; thickest lines, ρ = 0.97; thinnest lines, ρ = 0.5) among the milk components, represented by vertices, are annotated to show the relative difference and importance (vertex size, variable loading on LV1) of the glycans in milk from women with vs. without GDM (non-GDM). CH, complex hybrid; F, fucose; FS, fucose and sialic acid; GDM, gestational diabetes mellitus; HM, high mannose; LF, lactoferrin; LV, latent variable; O-PLS-DA, orthogonal signal corrected partial least-squares discriminant analysis; S, sialic acid; sIgA, secretory IgA; 5411, N-glycan containing 5 hexoses, 4 N-acetyl-hexosamines, 1 fucose, 1 sialic acid.