Mouse Paneth Cell-Enriched Proteome Enabled by Laser Capture Microdissection

Paneth cells are antimicrobial peptide-secreting
cells located
at the base of the crypts of the small intestine. The proteome of
Paneth cells is not well defined because of their coexistence with
stem cells, making it difficult to culture Paneth cells alone in vitro. Using a simplified toluidine blue O method for
staining mouse intestinal tissue, laser capture microdissection (LCM)
to isolate cells from the crypt region, and surfactant-assisted one-pot
protein digestion, we identified more than 1300 proteins from crypts
equivalent to 18,000 cells. Compared with the proteomes of villi and
smooth muscle regions, the crypt proteome is highly enriched in defensins,
lysozymes, and other antimicrobial peptides that are characteristic
of Paneth cells. The sensitivity of the LCM-based proteomics approach
was also assessed using a smaller number of cell equivalent tissues:
a comparable proteomic coverage can be achieved with 3600 cells. This
work is the first proteomics study of intestinal tissue enriched with
Paneth cells. The simplified workflow enables profiling of Paneth
cell-associated pathological changes at the proteome level directly
from frozen intestinal tissue. It may also be useful for proteomics
studies of other spatially resolved cell types from other tissues.