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. 2023 Jan 18;10:1102209. doi: 10.3389/fmolb.2023.1102209

FIGURE 1.

FIGURE 1

Apical-out intestinal organoids show tight barrier formation. (A) Overview of the protocol for the directed differentiation of H9 embryonic stem cells towards intestinal organoids with apical-out orientation. Scale bar: 100 μm. (B) On the left, a low magnification TEM image of the epithelial sheet of apical-out organoids demonstrating some enterocytes. In the middle image, TEM indicates the presence of apical microvilli (MV) on the outer surface of organoids. Functional ultrastructural features of the intestinal epithelium are indicated in the apical region of the enterocytes: microvilli (arrows) with core actin filaments and glycocalix (stars), vesicles of smooth endoplasmic reticulum (sER) and intercellular junctions: tight junction (TJ), zonula adherens (ZA) and desmosome (De). In the basal region (right image), the indicated structures correspond to the basement membrane (BM), collagen fibers (CF) and the mitochondria (Mit). Scale bars: 1 μm (left) and 500 nm (middle and right). (C) qRT-PCR analysis shows significantly higher expression of the “leak pathway” regulators ZO-1 and OCLN and the “pore pathway” regulators CLDN1, CLDN3, CLDN5 in the apical-out intestinal organoids compared to undifferentiated control stem cells. Error bars indicate mean ± S.E.M. (n = 3).