Figure 6. Colonic epithelial cells engulf apoptotic cells within the colon.
(A) Silencing of BAI1 by siRNA in colonic epithelial cell line HCT-116 (normalized to HPRT1).
(B) HCT-116 cells transfected with control-siRNA and BAI1-siRNA for 48 hours were assessed for engulfment of apoptotic MLE-12 cells labeled with CypHer5E.
(C) HCT-116 cells transfected with GFP, BAI1-WT, and BAI1-AAA were assessed for engulfment of apoptotic MLE-12 cells labeled with CypHer5E.
(D) Engulfment by Caco-2 cells was assessed with apoptotic BEAS-2B or apoptotic MLE-12 cells labeled with CypHer5E. Cytochalasin D (blocks uptake but not binding), was used to ensure that the signal is due to corpse engulfment.
(E) Schematic of staining of apoptotic cells (TUNEL, type I DNA break) and engulfed cells (DNase II-mediated type II break within phagocytes) after DSS (3 %) treatment.
(F and G) Analysis of apoptotic (DNase type I cleavage, yellow) and engulfed corpses (DNase type II cleavage, green) within the proximal colon of control (n=6; 6F) and Vil1-cre, BAI1Tg (n=3; 3F) mice after DSS (3 %) treatment. In the enlarged images, the arrows indicate signal for engulfed corpses within EpCAM+ epithelial cells, and arrow heads indicate double positive cells (signal for apoptotic and engulfed). Hoechst33342 used to normalize between sections; the number of engulfed cells in control animals was set to 100 %. Scale bars=100 µm.
(H) Analysis of apoptotic cell numbers in proximal colon of control (n=6; 6F) and Vil1-cre, BAI1Tg (n=3; 3F) mice after DSS (3 %) treatment.
(I) z-stack image of engulfed cells (DNase II positive, green) within colonic epithelial cells. Scale bars=5 µm.
Data from at least 2–3 independent experiments were combined. Error bars indicate s.e.m. *P<0.05. F, female. See also Figure S6.