Semaglutide Normalizes Increased Cardiomyocyte Calcium Transients in a Rat Model of High Fat Diet-Induced Obesity

Figures

• 

  Figure 1. Overview of the experimental design and the different treatment groups. Rats in the HFD/Sema group (n = 8) were fed a high‐fat, high‐fructose diet (HFD) for 8 weeks, followed by 8 weeks of treatment with subcutaneous semaglutide (120 μg/kg/day); during this treatment period, they had access to either the HFD or a low‐fat diet (LFD). The HFD/Veh group (n = 8) was managed similarly to the HFD/Sema group, except they received saline instead of semaglutide. Rats in the HFD group (n = 3) were given the high‐fat, high‐fructose diet, and the LFD group (n = 3) received the low‐fat diet for the entire duration of 16 weeks. Control rats (CON, n = 5) were maintained on a standard chow diet throughout the 16 weeks. Created with BioRender.com.

• 

  Figure 2. Sarcomeric and cytosolic Ca2+ dynamics transients in cardiomyocytes from controls (CON), non‐treated (HFD/Veh), and semaglutide‐treated high‐fat, high‐fructose diet (HFD) rat hearts. Cardiomyocytes were paced at 0.3 Hz under normal Tyrode conditions (NT), with 30 nM isoprenaline (Iso) at 0.3 Hz and 3 Hz, and during the washout (WO) phase at 0.3 Hz. (Top panels) Sarcomere length plotted to intracellular [Ca2+]c. (Mid panels) Diastolic and systolic sarcomere lengths. (Bottom panels) Diastolic and systolic intracellular [Ca2+]c. The washout (WO) phase refers to the time following the cessation of 3 Hz stimulation, still in the presence of isoprenaline, with stimulation resumed at 0.3 Hz frequency for 240 seconds.

• 

  Figure 3. Sarcomeric and cytosolic Ca2+ dynamics in cardiomyocytes from controls (CON), and non‐treated (HFD/Veh) and semaglutide‐treated high‐fat, high‐fructose diet (HFD) rat hearts. Cardiomyocytes were paced at 0.3 Hz and exposed to physiological stress conditions with 30 nM Isoprenaline and an increased stimulation rate to 3 Hz for 3 min. Panel (A) displays diastolic and systolic sarcomere lengths. Panel (B) illustrates percentage of sarcomere shortening. Panel (C) shows time to 50% and 90% sarcomere re‐lengthening. Panel (D) presents diastolic and systolic intracellular [Ca2+]c. Panel (E) illustrates the [Ca2+]c transient amplitude. Panel (F) shows the time to (TT) 50% and 90% [Ca2+]c decay. Control rat cardiomyocytes (n = 51 cardiomyocytes from n = 3 hearts). HFD/Sema cardiomyocytes were isolated from HFD rats treated with semaglutide (n = 120 cardiomyocytes from n = 5 hearts). HFD/Veh cardiomyocytes were isolated from HFD rats administered saline (n = 100 cardiomyocytes from n = 3 hearts). Washout phase (WO) refers to the time following the cessation of 3 Hz stimulation, with stimulation resumed at 0.3 Hz frequency. Statistical significance: # P < 0.05 HFD/Sema versus HFD/Veh…

• 

  Figure 4. Sarcoplasmic reticulum (SR) Ca2+ load in cardiomyocytes from non‐treated (HFD/Veh) and semaglutide‐treated high‐fat, high‐fructose diet (HFD) hearts. (A) Original tracings showcase caffeine‐induced increases in cytosolic [Ca2+]c, serving as an indicator of SR Ca2+ load, at 1 Hz pacing and 10 mM caffeine. Panel (B) Displays the amplitude of [Ca2+]c changes (Δ[Ca2+]c). Panel (C) Illustrates the tau (τ) of [Ca2+]c removal at 1 Hz, indicative of SR Ca2+‐ATPase (SERCA) activity, and during caffeine application, representing Na+/Ca2+‐exchanger (NCX) activity. Panel (D) reveals the time to peak (TPP) [Ca2+]c at 1 Hz and during caffeine application. HFD/Sema cardiomyocytes were isolated from HFD rats treated with semaglutide (n = 42 cardiomyocytes from n = 3 hearts). HFD/Veh cardiomyocytes were isolated from HFD rats administered saline (n = 49 cardiomyocytes from n = 3 hearts). Sal, Saline; Sema, Semaglutide. Statistical significance: #P < 0.05 (one‐way ANOVA); *P < 0.05, **P < 0.01, ****P < 0.0001.

• 

  Figure 5. L‐Type Ca2+ channel activity through patch‐clamp electrophysiology. Panel (A) presents baseline activity of L‐Type Ca2+ channel, and (B) its activity during the application of 30 nM Isoprenaline, with representative protocols illustrated in the insets. Panel (C) outlines the Ca2+‐induced inactivation tau (τ) kinetics of the L‐type Ca2+ channel. HFD/Sema cardiomyocytes were isolated from HFD rats treated with semaglutide (n = 33 cardiomyocytes from n = 3 hearts). HFD/Veh cardiomyocytes were isolated from HFD rats administered saline (n = 29 cardiomyocytes from n = 3 hearts). Sal, Saline; Sema, Semaglutide. Statistical significance: #P < 0.05 (two‐way ANOVA); *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.

• 

  Figure 6. Sarcomeric and cytosolic Ca2+ dynamics in cardiomyocytes from low‐fat (LFD) and high‐fat, high‐fructose diet (HFD) rat hearts. Cardiomyocytes were paced at 0.3 Hz and exposed to physiological stress conditions with 30 nM Isoprenaline and an increased stimulation rate to 3 Hz for 3 min. Panel (A) displays diastolic and systolic sarcomere lengths. Panel (B) illustrates percentage of sarcomere shortening. Panel (C) shows time to 50% and 90% sarcomere re‐lengthening. Panel (D) presents diastolic and systolic intracellular [Ca2+]c. Panel (E) illustrates the [Ca2+]c transient amplitude. Panel (F) shows the time to (TT) 50% and 90% [Ca2+]c decay. LFD (16 weeks), cardiomyocytes isolated from rats maintained on a low‐fat diet for 16 weeks (n = 79 cardiomyocytes from n = 3 hearts). HFD (16 weeks), cardiomyocytes isolated from rats subjected to a high‐fat, high‐fructose diet for 16 weeks (n = 70 cardiomyocytes from n = 3 hearts). WO, washout phase, refers to the time following the cessation of 3 Hz stimulation, with stimulation resumed at 0.3 Hz frequency. Statistical significance: # P < 0.05 HFD versus LFD (two‐way ANOVA).

• 

  Figure 7. Sarcomeric and cytosolic Ca2+ dynamics transients in cardiomyocytes from low‐fat (LFD) and high‐fat, high‐fructose diet (HFD) rat hearts. Cardiomyocytes were paced at 0.3 Hz under normal Tyrode conditions (NT), with 30 nM isoprenaline (Iso) at 0.3 Hz and 3 Hz, and during the washout (WO) phase at 0.3 Hz. (Top panels) Sarcomere length plotted to intracellular [Ca2+]c. (Mid panels) Diastolic and systolic sarcomere lengths. (Bottom panels) Diastolic and systolic intracellular [Ca2+]c. The washout (WO) phase refers to the time following the cessation of 3 Hz stimulation, still in the presence of isoprenaline, with stimulation resumed at 0.3 Hz frequency for 240 s.