Prevention of antibiotic-associated metabolic syndrome in mice by intestinal alkaline phosphatase

Abstract

Aims: To examine whether co-administration of intestinal alkaline phosphatase (IAP) with antibiotics early in life may have a preventive role against metabolic syndrome (MetS) in mice.

Methods: A total of 50 mice were allocated to four treatment groups after weaning. Mice were treated with azithromycin (AZT) ± IAP, or with no AZT ± IAP, for three intermittent 7-day cycles. After the last treatment course, the mice were administered a regular chow diet for 5 weeks and subsequently a high-fat diet for 5 weeks. Body weight, food intake, water intake, serum lipids, glucose levels and liver lipids were compared. 16S rRNA gene pyrosequencing was used to determine the differences in microbiome composition.

Results: Exposure to AZT early in life rendered mice susceptible to MetS in adulthood. Co-administration of IAP with AZT completely prevented this susceptibility by decreasing total body weight, serum lipids, glucose levels and liver lipids to the levels of control mice. These effects of IAP probably occur as a result of changes in the composition of specific bacterial taxa at the genus and species levels (e.g. members of Anaeroplasma and Parabacteroides).

Conclusions: Co-administration of IAP with AZT early in life prevents mice from susceptibility to the later development of MetS. This effect is associated with alterations in the composition of the gut microbiota. IAP may represent a novel treatment against MetS in humans.

Keywords: azithromycin; intestinal alkaline phosphatase; metabolic syndrome; mice.

Figure 1

Schematic diagram summarizing the timeline of experimental procedures in this study. Fifty male wild-type C57BL/6 mice (IAP group: 12 mice, Vehicle group: 14 mice, AZT+IAP: 10 mice and AZT+Vehicle: 14 mice) were used. FC:Fecal collections, Seq:Microbiome sequencing, IAP:Intestinal Alkaline Phosphatase, LFD:Low-fat diet, HFD:High-fat diet.

Figure 2

Food intake (A), water intake (B), body (C–E), visceral (F) and subcutaneous (G) fat weights among the experimental treatment groups (n=10–14 per experimental group; see legend of Figure 1).Panel C depicts the average body weight gain of the mice in each group during the HFD period (11–15 weeks). Body (D), visceral (F) and subcutaneous (G) fat weights were measured at the end of the study (at 15 weeks).IAP:Intestinal Alkaline Phosphatase, HFD:High-fat diet, Az:Azithromycin, Veh:Vehicle.*p<0.05;**p<0.01;***p<0.001.

Figure 3

Serum lipid profile (A–C) and serum glucose (D–F) among the experimental treatment groups (n=10–14 per experimental group; see legend of Figure 1) at the end of the study (at 15 weeks). IAP:Intestinal Alkaline Phosphatase, LDL:low density lipoprotein, HDL:high density lipoprotein, GTT:glucose tolerance test. *p<0.05;**p<0.01;***p<0.001.

Figure 4

Histological specimens of liver tissue stained with oil-red-O in the IAP (A), Vehicle (B), Azithromycin+IAP (C) and Azithromycin+Vehicle (D) groups (n=10–14 per experimental group; see legend of Figure 1). Liver histopathologic scores (E), triglycerides (F) and total lipids (G) among the experimental treatment groups. Hepatic steatosis was graded based on the number and size of stained fat droplets: 0(none/minimal); 1+(mild); 2+(moderate); 3+(moderate/marked); 4+(marked).IAP:intestinal alkaline phosphatase. *p<0.05;**p<0.01;***p<0.001.

Figure 5

(A) and (B) Species richness (as estimated using the Chao1 estimator) (A) and Shannon diversity (B) within the microbiome data for each group (n= 5 mice per group). Operational taxonomic units (OTUs) are defined at 97% 16S rRNA gene sequence similarity. Mean values for each group are indicated by solid lines, and shading indicates standard error around estimates of the mean. Confidence intervals are shown for Chao1 estimates from individual mice (A). (C) and (D) Principle coordinates analysis of unweighted (C) and weighted (D)UniFrac distances for sequences obtained from treatment groups. Percentage values on axes indicate the amount of variability in the data explained by each of the first two principal coordinates. Contributions of individual taxa to the segregation of samples are indicated by labeled vectors. IAP:Intestinal Alkaline Phosphatase, AZT:Azithromycin.