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Acute cardiotoxicity evaluation of the marine biotoxins OA, DTX-1 and YTX.

Ferreiro SF, Carrera C, Vilariño N, Louzao MC, Santamarina G, Cantalapiedra AG, Botana LM - Toxins (Basel) (2015)

Bottom Line: Food safety authorities from several countries have regulated the content of DSPs and YTXs in shellfish to protect human health.The results demonstrated that these toxins do not exert acute effects on hERG channel activity.Additionally, in vivo experiments showed that these compounds do not alter cardiac biomarkers and ECG in rats acutely.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Farmacología, Facultad de Veterinaria, Universidad de Santiago de Compostela, 27002 Lugo, Spain. sara.fernandez.ferreiro@usc.es.

ABSTRACT
Phycotoxins are marine toxins produced by phytoplankton that can get accumulated in filter feeding shellfish. Human intoxication episodes occur due to contaminated seafood consumption. Okadaic acid (OA) and dynophysistoxins (DTXs) are phycotoxins responsible for a severe gastrointestinal syndrome called diarrheic shellfish poisoning (DSP). Yessotoxins (YTXs) are marine toxins initially included in the DSP class but currently classified as a separated group. Food safety authorities from several countries have regulated the content of DSPs and YTXs in shellfish to protect human health. In mice, OA and YTX have been associated with ultrastructural heart damage in vivo. Therefore, this study explored the potential of OA, DTX-1 and YTX to cause acute heart toxicity. Cardiotoxicity was evaluated in vitro by measuring hERG (human èter-a-go-go gene) channel activity and in vivo using electrocardiogram (ECG) recordings and cardiac damage biomarkers. The results demonstrated that these toxins do not exert acute effects on hERG channel activity. Additionally, in vivo experiments showed that these compounds do not alter cardiac biomarkers and ECG in rats acutely. Despite the ultrastructural damage to the heart reported for these toxins, no acute alterations of heart function have been detected in vivo, suggesting a functional compensation in the short term.

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Effects of OA, DTX-1 and YTX on human èter-a-go-go gene (hERG) channel activity. Automated patch clamp experiments were performed using a CHO cell line stably expressing hERG. (A) Voltage clamp protocol for the activation of hERG; (B) Representative current trace obtained after YTX addition. Currents were monitored for 5 min after toxin addition. COI: current of interest; (C) No effect of OA on hERG currents. Current magnitude is expressed as percentage of pre-treatment current that remained after 5 min of exposure to 10 µM OA or carrier alone (mean ± SEM; n = 4); (D) No effect of YTX on hERG currents. Current magnitude is expressed as percentage of pre-treatment current that remained after 5 min of exposure to 10 µM YTX or carrier (mean ± SEM; n = 4).
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toxins-07-01030-f002: Effects of OA, DTX-1 and YTX on human èter-a-go-go gene (hERG) channel activity. Automated patch clamp experiments were performed using a CHO cell line stably expressing hERG. (A) Voltage clamp protocol for the activation of hERG; (B) Representative current trace obtained after YTX addition. Currents were monitored for 5 min after toxin addition. COI: current of interest; (C) No effect of OA on hERG currents. Current magnitude is expressed as percentage of pre-treatment current that remained after 5 min of exposure to 10 µM OA or carrier alone (mean ± SEM; n = 4); (D) No effect of YTX on hERG currents. Current magnitude is expressed as percentage of pre-treatment current that remained after 5 min of exposure to 10 µM YTX or carrier (mean ± SEM; n = 4).

Mentions: The ability to block hERG channel currents is one of the required tests for preliminary evaluation of cardiotoxicity in drug development. Therefore, the effects of OA and YTX (Figure 1) on hERG activity were explored using automated patch clamp for the measurement of hERG currents in a CHO cell line stably expressing this channel. HERG channels were activated with the voltage protocol shown in Figure 2A. After stabilization of hERG currents, the cells were exposed to 10 µM OA or YTX, or an equivalent concentration of DMSO (carrier) for 5 min. The Ionflux system warrants an immediate change of extracellular solution and therefore no time is needed to allow for drug diffusion. For routine screening of hERG inhibiting drugs it is common to test concentrations in this range that will cause a clear and fast inhibition if the drug has high to medium blocking potency. A representative image of the current displayed by hERG CHO cells after YTX addition appears in Figure 2B. OA (Figure 2C, grey bar) and YTX (Figure 2D, grey bar) did not alter IKr amplitude after 5 minutes of exposure to a toxin concentration of 10 µM, when compared to the current in the same cells just before toxin addition. Similar results were obtained in cells exposed to carrier alone; no difference of IKr before and after addition of DMSO was observed (Figure 2C,D, white bars). DTX-1 (Figure 1) is an analog of OA and it has been described to have similar potency [1,31,32], thereby it was not tested in this assay to save the limited amount available for in vivo studies, which will provide information about potential cardiotoxicity generated by several possible mechanisms. Concentrations higher than 10 µM were not tested considering that compounds with an IC50 between 1 and 100 µM are classified as low potency hERG blockers and therefore would be pathologically irrelevant [33]. An underestimation of blocking potency due to technique limitations is not probable because the IC50 obtained for cisapride, a well-known high-potency hERG blocker, in our experimental conditions was 3 nM [34].


Acute cardiotoxicity evaluation of the marine biotoxins OA, DTX-1 and YTX.

Ferreiro SF, Carrera C, Vilariño N, Louzao MC, Santamarina G, Cantalapiedra AG, Botana LM - Toxins (Basel) (2015)

Effects of OA, DTX-1 and YTX on human èter-a-go-go gene (hERG) channel activity. Automated patch clamp experiments were performed using a CHO cell line stably expressing hERG. (A) Voltage clamp protocol for the activation of hERG; (B) Representative current trace obtained after YTX addition. Currents were monitored for 5 min after toxin addition. COI: current of interest; (C) No effect of OA on hERG currents. Current magnitude is expressed as percentage of pre-treatment current that remained after 5 min of exposure to 10 µM OA or carrier alone (mean ± SEM; n = 4); (D) No effect of YTX on hERG currents. Current magnitude is expressed as percentage of pre-treatment current that remained after 5 min of exposure to 10 µM YTX or carrier (mean ± SEM; n = 4).
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Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4417953&req=5

toxins-07-01030-f002: Effects of OA, DTX-1 and YTX on human èter-a-go-go gene (hERG) channel activity. Automated patch clamp experiments were performed using a CHO cell line stably expressing hERG. (A) Voltage clamp protocol for the activation of hERG; (B) Representative current trace obtained after YTX addition. Currents were monitored for 5 min after toxin addition. COI: current of interest; (C) No effect of OA on hERG currents. Current magnitude is expressed as percentage of pre-treatment current that remained after 5 min of exposure to 10 µM OA or carrier alone (mean ± SEM; n = 4); (D) No effect of YTX on hERG currents. Current magnitude is expressed as percentage of pre-treatment current that remained after 5 min of exposure to 10 µM YTX or carrier (mean ± SEM; n = 4).
Mentions: The ability to block hERG channel currents is one of the required tests for preliminary evaluation of cardiotoxicity in drug development. Therefore, the effects of OA and YTX (Figure 1) on hERG activity were explored using automated patch clamp for the measurement of hERG currents in a CHO cell line stably expressing this channel. HERG channels were activated with the voltage protocol shown in Figure 2A. After stabilization of hERG currents, the cells were exposed to 10 µM OA or YTX, or an equivalent concentration of DMSO (carrier) for 5 min. The Ionflux system warrants an immediate change of extracellular solution and therefore no time is needed to allow for drug diffusion. For routine screening of hERG inhibiting drugs it is common to test concentrations in this range that will cause a clear and fast inhibition if the drug has high to medium blocking potency. A representative image of the current displayed by hERG CHO cells after YTX addition appears in Figure 2B. OA (Figure 2C, grey bar) and YTX (Figure 2D, grey bar) did not alter IKr amplitude after 5 minutes of exposure to a toxin concentration of 10 µM, when compared to the current in the same cells just before toxin addition. Similar results were obtained in cells exposed to carrier alone; no difference of IKr before and after addition of DMSO was observed (Figure 2C,D, white bars). DTX-1 (Figure 1) is an analog of OA and it has been described to have similar potency [1,31,32], thereby it was not tested in this assay to save the limited amount available for in vivo studies, which will provide information about potential cardiotoxicity generated by several possible mechanisms. Concentrations higher than 10 µM were not tested considering that compounds with an IC50 between 1 and 100 µM are classified as low potency hERG blockers and therefore would be pathologically irrelevant [33]. An underestimation of blocking potency due to technique limitations is not probable because the IC50 obtained for cisapride, a well-known high-potency hERG blocker, in our experimental conditions was 3 nM [34].

Bottom Line: Food safety authorities from several countries have regulated the content of DSPs and YTXs in shellfish to protect human health.The results demonstrated that these toxins do not exert acute effects on hERG channel activity.Additionally, in vivo experiments showed that these compounds do not alter cardiac biomarkers and ECG in rats acutely.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Farmacología, Facultad de Veterinaria, Universidad de Santiago de Compostela, 27002 Lugo, Spain. sara.fernandez.ferreiro@usc.es.

ABSTRACT
Phycotoxins are marine toxins produced by phytoplankton that can get accumulated in filter feeding shellfish. Human intoxication episodes occur due to contaminated seafood consumption. Okadaic acid (OA) and dynophysistoxins (DTXs) are phycotoxins responsible for a severe gastrointestinal syndrome called diarrheic shellfish poisoning (DSP). Yessotoxins (YTXs) are marine toxins initially included in the DSP class but currently classified as a separated group. Food safety authorities from several countries have regulated the content of DSPs and YTXs in shellfish to protect human health. In mice, OA and YTX have been associated with ultrastructural heart damage in vivo. Therefore, this study explored the potential of OA, DTX-1 and YTX to cause acute heart toxicity. Cardiotoxicity was evaluated in vitro by measuring hERG (human èter-a-go-go gene) channel activity and in vivo using electrocardiogram (ECG) recordings and cardiac damage biomarkers. The results demonstrated that these toxins do not exert acute effects on hERG channel activity. Additionally, in vivo experiments showed that these compounds do not alter cardiac biomarkers and ECG in rats acutely. Despite the ultrastructural damage to the heart reported for these toxins, no acute alterations of heart function have been detected in vivo, suggesting a functional compensation in the short term.

Show MeSH
Related in: MedlinePlus