Figure 4.

Enclysis Forms β-Catenin-Rich Vesicles and Is Distinct from Entosis and Suicidal Emperipolesis

(A) Huh-7 cells (CMFDA, green) were co-cultured with Jurkat T cells (BMQC, blue), fixed, stained for anti-E-cadherin (anti-mouse Alexa Fluor 594, red), and imaged by confocal microscopy. The arrow indicates an E-cadherin⁺ entotic vesicle containing another Huh-7 cell.

(B) A binucleate Huh-7 cell containing a T cell in a vesicle devoid of E-cadherin staining.

(C) Huh-7 cell containing a T cell in a β-catenin⁺ enclytic vesicle.

(D) Orthogonal confocal image of a 30-μm-thick section from formalin-fixed, paraffin-embedded tissue from a cirrhotic liver explant stained for β-catenin and the CD4⁺ T cell transcription factor Tbet. Immunohistochemistry reveals autofluorescence, which helps visualize hepatocyte cytoplasm (blue) and,typical for the liver, cell debris that autofluoresces in all channels (white).

(E) Pre-treatment of Huh-7 cells with wortmannin, which inhibits suicidal emperipolesis, did not affect enclysis. However, the myosin-II and macropinocytosis inhibitor blebbistatin (50 μM) reduced T cell capture, as measured by confocal microscopy.

(F) Entosis inhibitor treatment of Huh-7 cells with the ROCK inhibitors Y-27632 and H1152 (10 μM) did not affect enclysis.

(G) Enclysis was perturbed by Huh-7 pre-treatment with the actin organization inhibitors latrunculin A (0.1 μM) and cytochalasin D (1 μM).

Data were derived from three independent experiments. ^∗∗p < 0.01, ^∗∗∗p < 0.001, Student’s non-parametric paired t test (Wilcoxon). Scale bars represent 10 μm.

See also Figures S4–S6.