Sialic acid utilisation and synthesis in the neonatal rat revisited

Abstract

Background: Milk is the sole source of nutrients for neonatal mammals and is generally considered to have co-evolved with the developmental needs of the suckling newborn. One evolutionary conserved constituent of milk and present on many glycoconjugates is sialic acid. The brain and colon are major sites of sialic acid display and together with the liver also of synthesis.

Methodology/principal findings: In this study we examined in rats the relationship between the sialic acid content of milk and the uptake, utilization and synthesis of sialic acid in suckling pups. In rat milk sialic acid was found primarily as 3' sialyllactose and at highest levels between 3 and 10 days postpartum and that decreased towards weaning. In the liver of suckling pups sialic acid synthesis paralleled the increase in milk sialic acid reaching and keeping maximum activity from postnatal day 5 onwards. In the colon, gene expression profiles suggested that a switch from sialic acid uptake and catabolism towards sialic acid synthesis and utilization occurred that mirrored the change of sialic acid in milk from high to low expression. In brain sialic acid related gene expression profiles did not change to any great extent during the suckling period.

Conclusions/significance: Our results support the views that (i) when milk sialic acid levels are high, in the colon this sialic acid is catabolized to GlcNAc that in turn may be used as such or used as substrate for sialic acid synthesis and (ii) when milk sialic acid levels are low the endogenous sialic acid synthetic machinery in colon is activated.

Figure 1. Gene expression levels of GNE and Slc17a5.

A) Relative GNE (sialic acid synthesis) and Slc17a5 (sialic acid uptake) expression levels in the central nervous system (CNS), liver and small and large intestine throughout development and aging in the mouse, rat and human. (E, embryonic; d, day; w, week; y, year; dashed line, not determined). B) GNE and Slc17a5 expression in the intestine of 17 and 21 day old rat pups. The data was extracted from the Gene Expression Omnibus repository at http://www.ncbi.nlm.nih.gov/sites/GDSbrowser?acc=GDS1273. Mean and SD are shown; N = 3, except PND17 colon, N = 2.

Figure 2. Rat milk lactose, sialyllactose and sialic acid content throughout lactation.

Lactose (upper panel), the sialyllactoses (middle panel; 3SL left scale bar; 6SL right scale bar) and total sialic acid (lower panel) contents were measured in non-stimulated rat milk over time of lactation. Amounts are expressed in g per 100 g dried milk. Mean and SD are shown (N = 5).

Figure 3. Schematic representation of sialic acid metabolism and a summary of gene expression levels in the colon.

The pathways depicted are based on the KEGG aminosugar reference pathway. Full names of the indicated gene/protein are given in the text. The relative gene expression level and milk sialic acid content during first and third week of lactation are indicated by symbols and color code. (*Gene not in array or not expressed; ‘?’ no data that pgm1 can catalyse this step.)

Figure 4. Colon gene expression levels in suckling rat pups.

A–F) Filled symbols apply to the left scale bar and open symbols to the right scale bar. Mean and SD are shown (N = 6, except PND-1 N = 5). The bar above each x-axis indicates the relative level of sialic acid in milk (low, open and high, filled).

Figure 5. Colon gene expression levels in suckling rat pups.

A–C) Filled symbols apply to the left scale bar and open symbols to the right scale bar. Mean and SD are shown (N = 6, except PND-1 N = 5). The bar above each x-axis indicates the relative level of sialic acid in milk (low, open and high, filled).

Figure 6. Sialyllactose and sialic acid levels in plasma and urine of suckling rat pups.

A) Plasma 3′sialyllactose (3SL) and B) urine 3SL and free sialic acid (NeuAc) levels were determined in pups throughout the suckling period. Mean and SD are shown (N = 6). The bar above each x-axis indicates the relative level of sialic acid in milk (low, open and high, filled).

Figure 7. GNE activity in the livers of suckling rat pups.

GNE epimerase activity was determined in liver extracts throughout the suckling period. Mean and SD are shown (N = 6). The bar above the x-axis indicates the relative level of sialic acid in milk (low, open and high, filled).

Figure 8. Brain gene expression levels in suckling rat pups.

A–F) Filled symbols apply to the left scale bar and open symbols to the right scale bar. Mean and SD are shown (N = 6). The bar above each x-axis indicates the relative level of sialic acid in milk (low, open and high, filled).