Figure 1.
129 mice possess fewer Paneth cells than B6 mice.
(A) Lysozyme (Lyz) staining of ileal tissue shown at 20× magnification from B6 and 129 mice. Inset shows high power view (40×) of single crypt. (B) Flow cytometry was used to quantify Lyz+ cells from the ileal epithelium of B6 and 129 mice. Data are representative of 3 independent experiments. (C) Lyz transcripts were measured using quantitative RT-PCR of ileal RNA extracts from SPF and GF mice. Copy number is normalized to Gapdh, and fold change is relative to the GF B6 group (*P<.0001).
Figure 2.
B6 and 129 mice display distinct patterns of ileal antimicrobial peptide expression.
Ileal transcript levels of (A) α-defensins; (B) angiogenin 4 (Ang4), (C) regenerating islet-derived protein 3 gamma (Reg3γ); and (D) α-defensin related sequence 10 (Defa-rs10) are shown for B6 and 129 GF and SPF mice (n = 6–7 mice/group). Data are shown as means with SEM (*P<.05). Defa-rs10 was not detected (#) in any B6 animal.
Figure 3.
B6 and 129 mice exhibit distinct patterns of ileal α-defensin expression.
(A) mRNA expression of α-defensin isoforms in the ileum of B6 and 129 GF and SPF mice (n = 7/group). Copy number is normalized to Gapdh (*P<.05, # - not detected). (B) AU-PAGE demonstrates peptide expression patterns of α-defensin isoforms in the ileum of B6 and 129 SPF mice. First lane is recombinant Defa4 control; each additional lane represents an individual mouse. Individual bands (based on calculated mass determined via mass spectrometry): 1-Defa5; 2-Defa24, Defa27; 3-Defa20, Defa21; 4-Defa2; 5-Defa22; 6-Defa4 (recombinant); 7-Defa5; 8-Defa2/18v, Defa11, Defa16, Defa21; 9-Defa18, Defa25; 10-Defa4. (C) Representative mass spectrum for band #1. Mass data for all gel bands are summarized in Table S1.
Figure 4.
Host genotype affects the composition of intestinal microbiota.
(A) 4 experimental groups were defined by colonizing GF B6 or 129 mice with the stool microbiota of SPF B6 or 129 mice (Donor→Recipient). Ileal mucosa and stool bacterial communities were analyzed by T-RFLP at 12 wk post-colonization. (B) TRFs derived from these communities were used to generate nonmetric multidimensional scaling plots for ileal tissue and stool samples. In both bacterial compartments, the 129 (blue) and B6 (red) recipient groups were statistically dissimilar based on ANOSIM analysis (P<.05, n = 3–4 mice/group). (C) Percent contribution of T-RFs to each group (top 100% for ILEUM, top 90% for STOOL). Each color represents a unique T-RF indicating a putative bacterial group.
Figure 5.
Host genotype and bacterial source affect abundances of specific bacterial groups within the intestinal microbiota.
Quantification of specific bacterial groups from (A) ileal tissue samples, and (B) cecal contents using quantitative PCR of the 16 S rRNA gene is shown (n = 3–4 mice/group). Copy number is normalized to total 16 S, and fold change is relative to the B6 donor→B6 recipient group (black bars). Values are expressed as mean ± SEM (*P≤.05).
Table 1.
Donor and recipient effects on specific bacterial group levels.
Figure 6.
129 mouse α-defensins exhibit diminished antimicrobial activity against pro-inflammatory E. coli strains.
(A) AU-PAGE was used to generate α-defensin profiles from B6 and 129 mice. The α-defensin region was excised and overlaid onto E.coli-laden agarose. (B) Zones of bacterial inhibition were detected for both NC101 (upper) and LF82 (lower) E. coli strains. Increased growth inhibition was consistently observed from α-defensins from B6 mice.