Two-photon microscopy of Paneth cells in the small intestine of live mice

Abstract

Paneth cells are one of the principal epithelial cell types in the small intestine, located at the base of intestinal crypts. Paneth cells play key roles in intestinal host-microbe homeostasis via granule secretion, and their dysfunction is implicated in pathogenesis of several diseases including Crohn’s disease. Despite their physiological importance, study of Paneth cells has been hampered by the limited accessibility and lack of labeling methods. In this study, we developed a simple in vivo imaging method of Paneth cells in the intact mouse small intestine by using moxifloxacin and two-photon microscopy (TPM). Moxifloxacin, an FDA-approved antibiotic, was used for labeling cells and its fluorescence was strongly observed in Paneth cell granules by TPM. Moxifloxacin labeling of Paneth cell granules was confirmed by molecular counterstaining. Comparison of Paneth cells in wild type, genetically obese (ob/ob), and germ-free (GF) mice showed different granule distribution. Furthermore, Paneth cell degranulation was observed in vivo. Our study suggests that TPM with moxifloxacin labeling can serve as a useful tool for studying Paneth cell biology and related diseases.

Intravital imaging allows real-time characterization of tissue resident eosinophils

“…Intravital imaging for the GI tract, liver, skin, and lymph node were done as previously described 53–57 and reviewed in ref. 58 .…”

Astrovirus replication in human intestinal enteroids reveals multi-cellular tropism and an intricate host innate immune landscape

“…Mock-infected cells served as negative controls. UEA-1 lectin was chosen as it can stain multiple intestinal cell types (e.g., entero-endocrine cells [24], colon adenocarcinoma cells [24], small intestinal epithelial cells [25], goblet cells [26], Paneth cells [27]) in different mammalian species. As anticipated, no viral staining was observed in mock-infected cells (Fig 3A).…”

Paneth cells: Maintaining dynamic microbiome‐host homeostasis, protecting against inflammation and cancer

“…[ 28 ] This influence of PC is mediated by such factors as Wnt‐ligands (Wnt3, Wnt6 and Wnt9b), Notch ligands (Dll‐1 and Dll‐4), as well as epidermal (EGF) and α‐transforming (TGF‐α) growth factors. [ 4,12–20,22–25,27,28,30–60 ] PC affect +LGR5‐cells predominantly through the Wnt signaling pathway. [ 14 ] Wnt proteins are membrane‐bound ligands that are necessary for cell development, migration, and polarization.…”

“…[ 6 ] In addition, PC secrete Notch ligands on their membranes, such as Dll1 and Dll4, which bind the Notch receptors on intestinal stem cells, that leads to an increase in the number of stem cells. [ 2,4–20,22–25,27,28,30–59,61–65 ] Recent studies have shown that there is also a metabolic interdependence between PC and stem cells. [ 7 ] Therefore, remarkably, stem cells receive signals that regulate their activity from their descendants—PC.…”

“…[ 3 ] Since PC are highly specialized cells, they do not synthesize deoxyribonucleic acid and do not replicate. [ 40 ] The location, maturation and differentiation of PC in the crypt are regulated by the Wnt signaling pathway via activation of the β‐catenin mechanism. Wnt signaling controls the constitutive secretion of some PC specific products: the transcription factor (SOX9), necessary for PC differentiation, and regulatory molecule Ephrin type‐B (EphB), which determines the correct location along the crypt‐villus axis.…”