Human Cardiomyocytes aconitine resveratrol

Effects of resveratrol on aconitine-induced cardiac arrhythmia in human induced pluripotent stem cell derived cardiomyocytes

R. Printemps, S. Guilbot, M. Le Grand, PhysioStim, Zone Industrielle de Brénas, 81440 Lautrec, France

Resveratrol, a natural ingredient of grape skin, has been demonstrated to exert protective effect on the cardiovascular system. The use of aconitine, the main effective ingredient of a well-known medical herb widely used in traditional Chinese medicine (aconitium), is associated with severe cardiovascular toxicities including tachyarrhythmia and hypotension. The aim of the present work was to evaluate the potential cardioprotective action of resveratrol on aconitine-induced arrhythmia in human induced pluripotent stem cell derived cardiomyocytes (hiPSC-CMs). To this end, aconitine and resveratrol were tested alone first before evaluation of the co-administration of both compounds to finally assess the impact of a post-treatment of resveratrol after aconitine administration or a pre-treatment of resveratrol before aconitine-induced cardiac arrhythmia.

Methods

Human Induced Pluripotent Stem cells derived cardiomyocytes (iPSC-CMs)

  • iCell® Cardiomyocytes2 were supplied by Cellular Dynamics (a Fujifilm company) and seeded on 48 wells plate following manufacturers instruction and analyzed using the xCELLigence RTCA cardio ECR platform (simultaneous measurement of MEA and impedance).
  • 4 to 6 days after seeding, iCell2 cardiomyocytes were exposed to different concentrations of aconitine, resveratrol as well as the combination of both compounds, whether by co-administration or by pre-treatment or post-treatment with resveratrol.

  • Data are expressed as Mean ± SD and the first 2h of treatment are presented. This study is composed of 5 groups and the number of replicates in each group is detailed in the experimental design below.
  • The following parameters were monitored: Cell Index (CI), Amplitude of contraction, Beat rate (BR) , Beating period (BP) , Individual Beating Duration (IBD), Field Potential Duration (FPD) and FPD corrected by Fridericia (FPDc), Spike amplitude, Beating Rhythm Irregularity (BRI).

Experimental design

Results

  • After 30min, the cells stopped beating with aconitine 0.3µM, 1µM and 3µM.
  • Several concentrations of resveratrol were evaluated (0.1µM, 1µM, 10µM and 30µM), no major modification of all the parameters recorded was observed (data not shown).
  • Aconitine 0.1µM will be the concentration selected to induce arrhythmia.
  • Resveratrol 30µM will be evaluated for its antiarrhythmic capacity to reverse aconitine’s effects.

  • Tested alone, resveratrol 30µM had no effect on hiPSC-CMs while aconitine 0.1µM time-dependently ↘ amplitude of contraction (-40% after 30min) and FPDc (-60% after 30min) and ↗ Beat rate (+318% after 30min) and BRI (x5 times vs resveratrol).
  • Compared to aconitine 0.1µM alone, co-administration of aconitine 0.1µM + resveratrol 30µM had no effect on FPDc and BRI and only slightly lowered effects on amplitude of contraction (∆= -6% after 30min) and Beat rate (∆= -29% after 30min).
  • Post-addition of resveratrol 30µM after 30-min exposition to aconitine 0.1µM did not reduce aconitine 0.1µM effects (Amplitude of contraction: ∆= -4%, Beat rate: ∆= -11%, FPDc: ∆= +1% after 30min respectively).
  • A 30 min pre-treatment with resveratrol 30µM ↘ aconitine’s effects on all parameters (amplitude of contraction ∆= -17%, beat rate ∆= -93%, FPDc ∆= -15%, BRI ÷3.8 after 15min) and the occurrence of aconitine’s normal effects are delayed by 30min.

conclusion

  • Exposition of hiPSC-CMs to aconitine induce a massive increase of beating frequency and a decrease of amplitude of contraction.
  • In the same time, resveratrol was not able to suppress aconitine-induced arrhythmia in hiPSC-CMs. Resveratrol was only able to slightly reduce and also delay the occurrence of the maximum effects observed in the presence of aconitine when administrated as a pre-treatment.
  • This multi-parametric assay is particularly relevant for the evaluation of drugs’ anti-arrhythmic properties in a model of cardiac arrhythmia in Human iPSC derived cardiomyocytes.

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