Figure 3
Toll-like receptor 9 (TLR9) stimulation decreases SERCA2 activity in cardiomyocytes.
A RNAi-mediated knockdown of SERCA2 diminished the TLR9-induced AMP-activated protein kinase (AMPK) activation. Different siRNAs targeting SERCA2 similarly inhibited TLR9-induced AMPK activation. Values indicate densitometric ratio of pAMPK to tubulin in immunoblots, mean ± s.e.m. The data were obtained from three independent experiments.
B Ca2+-ATPase activity was assessed with microsome fractions from neonatal cardiomyocytes with or without 30-min treatment of CpG-ODN over two different concentrations of free Ca2+. The data were obtained from 4 (Ca2+; 0.17) or 5 (Ca2+; 7.3) preparations in each group.
C Förster resonance energy transfer (FRET)/CFP emission ratio plots of SR/ER Ca2+ probe in cardiomyocytes treated with control (n = 26) or CpG-ODN (n = 27). CpG-ODN significantly reduced SR/ER Ca2+ in neonatal cardiomyocytes after CpG-ODN. The measurements were normalized to the ratio at time 0.
D Representative FRET/CFP ratiometric pseudocoloured images of SR/ER Ca2+ probe in cardiomyocytes treated with control or CpG-ODN. Scale bar indicates 10 ÎĽm.
E Ca2+ content, as assessed by the amplitude of the caffeine-induced Fluo-4 transient in adult rat cardiomyocytes, was significantly decreased after CpG treatment (n = 28) compared to the control (n = 26).
F Representative traces of cytosolic Fluo-4 transients from the control (black; n = 50) and CpG-ODN (red; n = 51)-treated cardiomyocytes elicited at 0.5 Hz pacing.
G Time to peak and time to 50% decay of the Fluo-4 transients were both delayed by CpG stimulation. Also, the amplitude of the Fluo-4 transients (F/F0) was decreased by CpG stimulation.
H Thapsigargin (500 ng/ml for 0, 15, 30 min) increased pAMPK without the administration of CpG-ODN. The data were obtained from three independent experiments.
I CpG-ODN administration did not induce ER stress that was assessed by the expression level of GRP78 and GRP94 and was measured by real-time PCR in rat cardiomyocytes treated with CpG-ODN for the indicated period.
Data information: * P < 0.05 ** P < 0.01 *** P < 0.001 compared with the control. Error bars indicate s.e.m.
Source data are available online for this figure.
