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Comparative Study
. 2002 Nov 26;99(24):15451-5.
doi: 10.1073/pnas.202604299. Epub 2002 Nov 13.

Developmental regulation of intestinal angiogenesis by indigenous microbes via Paneth cells

Thaddeus S Stappenbeck  1 , Lora V Hooper, Jeffrey I Gordon
Affiliations
Comparative Study

Developmental regulation of intestinal angiogenesis by indigenous microbes via Paneth cells

Thaddeus S Stappenbeck et al. Proc Natl Acad Sci U S A. .

Abstract

The adult mouse intestine contains an intricate vascular network. The factors that control development of this network are poorly understood. Quantitative three-dimensional imaging studies revealed that a plexus of branched interconnected vessels developed in small intestinal villi during the period of postnatal development that coincides with assembly of a complex society of indigenous gut microorganisms (microbiota). To investigate the impact of this environmental transition on vascular development, we compared the capillary networks of germ-free mice with those of ex-germ-free animals colonized during or after completion of postnatal gut development. Adult germ-free mice had arrested capillary network formation. The developmental program can be restarted and completed within 10 days after colonization with a complete microbiota harvested from conventionally raised mice, or with Bacteroides thetaiotaomicron, a prominent inhabitant of the normal mouse/human gut. Paneth cells in the intestinal epithelium secrete antibacterial peptides that affect luminal microbial ecology. Comparisons of germ-free and B. thetaiotaomicron-colonized transgenic mice lacking Paneth cells established that microbial regulation of angiogenesis depends on this lineage. These findings reveal a previously unappreciated mechanism of postnatal animal development, where microbes colonizing a mucosal surface are assigned responsibility for regulating elaboration of the underlying microvasculature by signaling through a bacteria-sensing epithelial cell.

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Figures

Fig 1.
Fig 1.
Villus capillary networks in conventionally raised mice. (A–C) Sections prepared from the junction between the middle and distal thirds of the small intestine of a normal P28 mouse. (A) Hematoxylin/eosin (H&E)-stained section of a crypt–villus unit. The villus is located above, and the crypt below the dashed line. (Inset) A higher power view of the crypt showing Paneth cells (arrows). (B) Section from the same animal as in A, but stained with H&E and antibodies to the endothelial marker, von Willebrand's factor (purple; arrows). (C) Single confocal microscopic scan of a 120-μm-thick cryosection. The capillary network is stained with FITC-tagged high-molecular-weight dextran (green), and epithelial nuclei with Syto61 (red). To view a three-dimensional rotating image of a compiled set of serial scans of this type, see Movies 1 and 2, which are published as supporting information on the PNAS web site, www.pnas.org, or go to http://gordonlab.wustl.edu/vasculature. (D) Confocal view of FITC-dextran-labeled vessels in a villus positioned at the junction of the middle and distal thirds of the small intestine of a normal, conventionally raised P14 mouse. (Bars, 25 μm.)
Fig 2.
Fig 2.
Rapid microbial induction of angiogenesis in small intestinal villi of adult ex-germ-free mice. (A–C) Confocal scans of the capillary network present in the upper third of small intestinal villi. Whole mounts are from the junction between the middle and distal thirds of the small intestines of 6-week-old NMRI mice (capillaries, green; nuclei, red). (A) Germ-free (GF) mouse. (B) Age-matched ex-germ-free conventionalized (CONV) mouse killed 10 days after colonization with an unfractionated microbiota harvested from a conventionally raised “donor.” (C) Ex-germ-free mouse 10 days after colonization with B. thetaiotaomicron (B. theta) alone. To view three-dimensional rotating images of the capillary networks shown in A–C, see Movies 3–5, which are published as supporting information on the PNAS web site, or go to http://gordonlab.wustl.edu/vasculature. (D) Quantitation of villus capillary network density. Mean values ± SEM are plotted for each condition. The asterisk indicates that the difference relative to germ-free mice is statistically significant (P < 0.005 according to Student's t test). (Bars in A–C, 25 μm.)
Fig 3.
Fig 3.
Paneth cell and microbial regulation of angiogenesis. (A–D) Confocal scans of 120-μm-thick cryosections showing the upper thirds of villi. (A) Germ-free, Paneth cell-deficient P28 male CR2-tox176 mouse. (B) Age- and gender-matched, germ-free, Paneth-cell-containing normal littermate. (C) Ex-germ-free P28 CR2-tox176 mouse examined 7 days after colonization with B. thetaiotaomicron. (D) P28 nontransgenic mouse killed 7 days after mono-association with B. thetaiotaomicron. To view three-dimensional rotating images of the capillary networks shown in A–D, see Movies 6–9, which are published as supporting information on the PNAS web site, or go to http://gordonlab.wustl.edu/vasculature. (E) Quantitation of capillary network density. Mean values ± SEM for each condition are plotted. The statistical significance of differences between various groups is noted (Student's t test). (Bars in A–D, 25 μm.)
See this image and copyright information in PMC

Comment in

  • Commensal bacteria make a difference.
    Hentschel U, Dobrindt U, Steinert M. Hentschel U, et al. Trends Microbiol. 2003 Apr;11(4):148-50. doi: 10.1016/s0966-842x(03)00038-6. Trends Microbiol. 2003. PMID: 12706986

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