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. 2023 Oct 5;18(11):2123–2137. doi: 10.1016/j.stemcr.2023.09.002

Figure 4.

Figure 4

Effect of carnitine medium supplementation on EHT properties

(A) Effect of carnitine supplementation ΔGlucose per workload (ΔGlucose = Glucose concentration at baseline minus glucose concentration after 24 h of incubation; workload = force × frequency). Nested t test vs. CON, ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001. OCNT2 (+/+) control, n = 27 EHTs from 3 batches; OCNT2 (+/+) + carnitine (2 mM), n = 28 EHTs from 3 batches; OCTN2 (N32S) control, n = 23 EHTs from 3 batches; OCTN2 (N32S) + carnitine (2 mM), n = 23 EHTs from 3 batches; OCTN2 (−/−) control, n = 13 EHTs from 3 batches; OCTN2 (−/−) + carnitine (2 mM), n = 16 EHTs from 3 batches. Data are expressed as mean ± SEM.

(B) Effect of carnitine supplementation on force of spontaneous beating EHTs at the last day of treatment (days 33-42). Values were normalized to last day of treatment of untreated control. Student’s t test vs. CON, ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.001. OCNT2 (+/+) control, n = 54 EHTs from 4 batches; OCNT2 (+/+) + carnitine (2 mM), n = 49 EHTs from 4 batches; OCTN2 (N32S) control, n = 36 EHTs from 3 batches; OCTN2 (N32S) + carnitine (2 mM), n = 33 EHTs from 3 batches; OCTN2 (−/−) control, n = 9 EHTs from 1 batch; OCTN2 (−/−) + carnitine (2 mM), n = 9 EHTs from 1 batch. Data are expressed as mean ± SEM.

(C) Effect of carnitine supplementation on average contraction peaks. Depicted are representative average EHT contraction peaks of OCTN2 (+/+), OCTN2 (N32S), and OCTN2 (−/−). EHTs were electrically paced at 1.5 Hz in standard EHT medium ± carnitine (2 mM). Values were normalized to untreated control. n = 9–16 EHTs per condition from 1 batch.

(D and E) Liquid chromatography-mass spectrometry analysis of acylcarnitines and ceramides. Effect of carnitine supplementation on (D) acylcarnitine and (E) ceramide content of OCNT2 (+/+) and OCTN2 (N32S) EHTs after 33 days of culture and supplementation. Two-way ANOVA followed by Bonferroni’s post test for multiple comparisons, ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001. Data are expressed as mean ± SEM. n = 4 EHT pools (containing 3 EHTs each) per genotype and carnitine supplementation from 1 batch. See also Figure S5.