Figure 6.

Mitochondrial Ca²⁺ buffering reduces intracellular ROS levels elicited by Akkermansia muciniphila conditioned medium

(A) Confocal images of STC-1 cells transfected with mitochondrial parvalbumin (PV) expression and control vectors showing the expression and mitochondrial localization of targeted PV-MTS-GFP and MTS-GFP fusion proteins. Scale bar = 10 μM.

(B) Representative changes in mitochondrial Ca²⁺ signals over time are shown. Cells transfected with the indicated vectors were loaded with Rhod-2/AM and stimulated with 1%–10% unconditioned (BHI) or conditioned medium (BHI-CM) (arrow). Ca²⁺ signals were attenuated in cells expressing PV in mitochondria.

(C) Peak Ca²⁺ signals were observed in three separate experiments for STC-1 cells transfected with MTS-GFP, and cells transfected with PV-MTS-GFP. ∗p < 0.05 by unpaired Student's t-test.

(D) Representative changes in intracellular ROS levels over time are shown. Cells transfected with the indicated vectors were loaded with DHE and induced by 1/10% unconditioned (BHI) or conditioned medium (BHI-CM) (arrow). DHE fluorescence intensity was significantly reduced in cells expressing PV-MTS-GFP fusion protein.

(E) Peak ROS signals were observed in three separate experiments for STC-1 cells transfected with MTS-GFP, and cells transfected with PV-MTS-GFP stimulated with each represented condition. ∗p < 0.05 by unpaired Student’s t-test. Data in (B) and (D) represent a representative tracing recorded from one individual STC-1 cell of each group. Data in (C) and (E) represented as mean ± SEM of three independent experiments in which at least 25 individual cells were analyzed.