Location
Philadelphia, PA
Start Date
17-4-2026 1:30 PM
End Date
17-4-2026 2:30 PM
Description
Background: Trimethylamine N-oxide (TMAO) and its precursor trimethylamine (TMA) are metabolites derived from dietary sources such as choline and L-carnitine, commonly found in high-cholesterol foods and seafood. TMAO has been implicated in cardiovascular disease, particularly by promoting atherosclerotic plaque formation and vascular inflammation. However, the precise mechanisms by which TMA and TMAO induce cytotoxicity remain unclear, and further studies are needed. Spermidine, a naturally occurring polyamine present in living tissues, has been shown to exert anti-aging effects by inducing autophagy, reducing chronic inflammation, and preventing cell senescence. We hypothesized that spermidine would attenuate the cytotoxicity induced by TMA and TMAO.
Goals/methods: This study aimed to determine whether TMA, TMAO, and spermidine induce cytotoxicity in H9c2 cells. Further, to evaluate whether spermidine pretreatment confers protection against TMA and TMAO-induced cytotoxicity. H9c2 cells were pretreated with varying concentrations of spermidine for 24 hours, followed by the addition of a fixed concentration of either TMA or TMAO. Cell viability was assessed using a colorimetric Cell Counting Kit-8 (CCK-8) assay, and absorbance was measured at 450nm using a Multimode plate reader. Simultaneous cotreatment with spermidine and either TMA or TMAO was also evaluated. Data were expressed as mean±SE of absorbance readings.
Results: TMA and TMAO demonstrated dose-dependent cytotoxic effects. Moreover, TMA exhibited higher potency than TMAO. Untreated control cells had a mean absorbance of 1.67±0.06 (n=27). Treatment with 10 mM TMA resulted in a mean absorbance of 1.80±0.20 (n=5), whereas 50 mM TMA reduced viability to 0.54±0.02 (n=4). In comparison, TMAO required higher concentrations to achieve similar effects: 100 mM TMAO yielded 1.86±0.12 (n=5), while 200 mM TMAO reduced viability to 0.65±0.10 (n=9).
Spermidine slightly increased the cell viability of H9c2 cells. Compared with untreated controls (1.67±0.06, n=27), 10 μm and 100 μm of Spermidine exhibited cell viability of 1.84±0.05 (n=3) and 2.42±0.09 (n=5). Spermidine pretreatment showed a mild to modest increase in cell viability in the presence of TMA or TMAO. Compared with the 100 mM TMA control (0.48±0.03, n=12), pretreatment with 10 μM and 100 μM spermidine yielded higher cell viability of 0.70±0.07 (n=4) and 0.84±0.18 (n=4), respectively. Similarly, compared with the 200 mM TMAO-treated cells (0.65±0.10, n=9), pretreatment with 10 μM and 100 μM spermidine resulted in higher cell viabilities of 1.43±0.33 (n=3) and 1.62±0.64 (n=3), respectively. Cotreatment with spermidine did not demonstrate any protective effects.
Conclusion: Both TMA and TMAO reduced cell viability, with TMA demonstrating greater cytotoxic potency. Although spermidine pretreatment modestly increased cell viability in the presence of TMA or TMAO, additional experiments are needed to achieve statistical significance. Further research will investigate the underlying mechanisms associated with TMA, TMAO, and spermidine.
Embargo Period
6-3-2026
Included in
The Effects of Spermidine on Trimethylamine N-oxide and Trimethylamine – Induced Cytotoxicity on H9c2 Cells
Philadelphia, PA
Background: Trimethylamine N-oxide (TMAO) and its precursor trimethylamine (TMA) are metabolites derived from dietary sources such as choline and L-carnitine, commonly found in high-cholesterol foods and seafood. TMAO has been implicated in cardiovascular disease, particularly by promoting atherosclerotic plaque formation and vascular inflammation. However, the precise mechanisms by which TMA and TMAO induce cytotoxicity remain unclear, and further studies are needed. Spermidine, a naturally occurring polyamine present in living tissues, has been shown to exert anti-aging effects by inducing autophagy, reducing chronic inflammation, and preventing cell senescence. We hypothesized that spermidine would attenuate the cytotoxicity induced by TMA and TMAO.
Goals/methods: This study aimed to determine whether TMA, TMAO, and spermidine induce cytotoxicity in H9c2 cells. Further, to evaluate whether spermidine pretreatment confers protection against TMA and TMAO-induced cytotoxicity. H9c2 cells were pretreated with varying concentrations of spermidine for 24 hours, followed by the addition of a fixed concentration of either TMA or TMAO. Cell viability was assessed using a colorimetric Cell Counting Kit-8 (CCK-8) assay, and absorbance was measured at 450nm using a Multimode plate reader. Simultaneous cotreatment with spermidine and either TMA or TMAO was also evaluated. Data were expressed as mean±SE of absorbance readings.
Results: TMA and TMAO demonstrated dose-dependent cytotoxic effects. Moreover, TMA exhibited higher potency than TMAO. Untreated control cells had a mean absorbance of 1.67±0.06 (n=27). Treatment with 10 mM TMA resulted in a mean absorbance of 1.80±0.20 (n=5), whereas 50 mM TMA reduced viability to 0.54±0.02 (n=4). In comparison, TMAO required higher concentrations to achieve similar effects: 100 mM TMAO yielded 1.86±0.12 (n=5), while 200 mM TMAO reduced viability to 0.65±0.10 (n=9).
Spermidine slightly increased the cell viability of H9c2 cells. Compared with untreated controls (1.67±0.06, n=27), 10 μm and 100 μm of Spermidine exhibited cell viability of 1.84±0.05 (n=3) and 2.42±0.09 (n=5). Spermidine pretreatment showed a mild to modest increase in cell viability in the presence of TMA or TMAO. Compared with the 100 mM TMA control (0.48±0.03, n=12), pretreatment with 10 μM and 100 μM spermidine yielded higher cell viability of 0.70±0.07 (n=4) and 0.84±0.18 (n=4), respectively. Similarly, compared with the 200 mM TMAO-treated cells (0.65±0.10, n=9), pretreatment with 10 μM and 100 μM spermidine resulted in higher cell viabilities of 1.43±0.33 (n=3) and 1.62±0.64 (n=3), respectively. Cotreatment with spermidine did not demonstrate any protective effects.
Conclusion: Both TMA and TMAO reduced cell viability, with TMA demonstrating greater cytotoxic potency. Although spermidine pretreatment modestly increased cell viability in the presence of TMA or TMAO, additional experiments are needed to achieve statistical significance. Further research will investigate the underlying mechanisms associated with TMA, TMAO, and spermidine.