Arrhythmogenic mechanisms of interleukin-6 combination with hydroxychloroquine and azithromycin in inflammatory diseases

Figures

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  Figure 1. In-vivo impact of interleukin-6, azithromycin and hydroxychloroquine on the electrocardiogram of guinea pigs. (a) ECG of a guinea pig before, (b) after administration of IL-6(184 μg/kg) alone, (c) IL-6 + AZM(1) + HCQ(0.5), (d) IL-6 + AZM(1) + HC(1) and (e) IL-6 + AZM (1) + HCQ(2). Comparison of individual data points for heart rate (HR), PR, QRS and QTc between baseline and different interventions is illustrated in panels (f–i) using paired t-test versus basal group and one-way repeated measures analysis of variance for multiple comparisons among the drug interventions. Heart rate (j), PR (k), QRS (l) and QTc (m) data from two groups of guinea pigs, one (n = 6) injected with AZM (1) and HCQ (0.5) and the other (n = 6) injected with IL-6 (184 μg/kg) + AZM (1) + HCQ (0.5). AZM (1) = 38.2 mg/kg; HCQ(0.5) = 22.9 mg/kg; HCQ(1) = 45.8 mg/kg; HCQ(2) = 91.6 mg/kg. p values shown are in comparison to basal conditions.

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  Figure 2. In vivo impact of azithromycin and hydroxychloroquine on the IL-6R transcript and proteins levels. (a) mRNA levels of IL6R in untreated guinea pig hearts (control) and in guinea pigs treated with AZM (1) and HCQ (2) for 30 min. (b) Densitometric analysis and (c) the corresponding western blot for IL6Rα proteins untreated guinea pig heart lysate (control) and guinea pigs treated with AZM (1) and HCQ (2) for 30 min. The blots were probed with a monoclonal antibody for IL6Rα (Anti IL6Rα—H7 Santa Cruz). The band at 80 kDa represents IL6Rα. GAPDH indicates internal control. Each experiment was performed in triplicates from 4 guinea pigs (2 controls, lanes 1 & 2 and 2 treated with AZM + HCQ, lanes 3 & 4).

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  Figure 3. In-vivo impact of tocilizumab, interleukin-6, azithromycin and hydroxychloroquine on the electrocardiogram of guinea pigs (a) ECG of a guinea pig before, (b) after administration of TCZ (10 mg/kg) alone, (c) TCZ + IL-6(184 μg/kg), (d) TCZ + IL-6 + AZM(1) + HCQ(0.5), (e) TCZ + IL-6 + AZM(1) + HC(1) and (f) TCZ + IL-6 + AZM (1) + HCQ(2). Comparison of individual data points for heart rate (HR), PR, QRS and QTc between baseline and different interventions is illustrated in panels (g–j). AZM(1) = 38.2 mg/kg; HCQ (0.5) = 22.9 mg/kg; HCQ(1) = 45.8 mg/kg; HCQ(2) = 91.6 mg/kg. p values shown are in comparison to basal conditions.

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  Figure 4. In-vitro impact of interleukin-6, azithromycin and hydroxychloroquine on the electrogram of Langendorff perfused guinea pig hearts. Electrograms of an isolated guinea pig heart before (a) and (b) after perfusions of IL-6 (200 μg/L) alone for 40 min, (c) IL-6 + AZM(1) + HCQ(0.5), (d) IL-6 + AZM(1) + HCQ(1) and (e) IL-6 + AZM (1) + HCQ (2). The comparison of individual data points for heart rate (HR), PR, QRS and QTc between baseline and the different interventions is illustrated in panels (f–i) using paired t-test. Heart rate (j), PR (k), QRS (l) and QTc (m) data from two groups of guinea pigs, one (n = 5) perfused with AZM(1) and HCQ(0.5) and the other (n = 5) perfused with IL-6(200 µg/L) + AZM(1) + HCQ(0.5). AZM(1) = 41.5 mg/L; HCQ(0.5) = 24.9 mg/L. AZM(1) = 41.5 mg/L; HCQ(0.5) = 24.9 mg/L; HCQ(1) = 49.8 mg/L; HCQ(2) = 99.6 mg/L. Paired t-test was used to compare interventions to baseline. (*) indicates p < 0.05.

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  Figure 5. Impact of interleukin-6, azithromycin and hydroxychloroquine on action potential and key ion channels in guinea pig ventricular myocytes. Action potentials (a) and ion currents, ICaL (b), INa (c) and IKr (g–k) recorded from control myocytes (black traces) and from another group of myocytes preincubated with IL-6 for 40 min (red traces), IL-6(200 µg/L) + AZM (1) + HCQ (0.1) (green traces), IL-6 + AZM (1) + HCQ(0.5) (pink traces) and IL-6 + AZM(1) + HCQ(1) (dark red traces). The comparison of individual data points for APD90 (d) and current densities for ICaL (e), INa (f) and IKr (l) during control and after different interventions is presented. HCQ(0.1) = 5.0 mg/L; HCQ(0.5) = 24.9 mg/L; HCQ(1) = 49.8 mg/L; AZM(1) = 41.5 mg/L. Unpaired t-test was used between the control group and the group treated with IL-6 + AZM + HCQ. p values shown are comparison to basal conditions.

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  Figure 6. Impact of azithromycin and hydroxychloroquine with and without interleukin-6 on action potential and key ion channels in guinea pig ventricular myocytes. APD90 (a), AP amplitude (APA, b), ICaL (c) INa (d) and IKr (e) data from two groups of guinea pig myocytes before (basal) and after administrations of AZM(1) + HCQ(0.1); and the other control group without IL-6 and the treated group with IL-6 (200 μg/L), AZM(1) and HQC(0.1). AZM(1) = 41.5 mg/L; HCQ(0.5) = 24.9 mg/L. Paired t-test was used within the same group before (basal) and with (AZM + HCQ). Unpaired t-test was used between control group and the group treated with IL-6 + AZM + HCQ. p values shown are comparison to basal conditions.