Comparisons of chromosome Y-substituted mouse strains reveal that the male-specific chromosome modulates the effects of androgens on cardiac functions

Abstract

Background: The C57BL/6J.Y^A/J mouse strain is a chromosome-substituted line where the original male-specific portion of chromosome Y (MSY) from C57BL/6J mice was substituted for that from A/J mice. In hearts from male C57BL/6J.Y^A/J and C57BL/6J mice, orchidectomy (ORX) affected in a strictly strain-specific fashion the expression a subset of genes showing enrichment for functional categories, including that of circadian rhythms and cardiac contractility. We further tested whether: (1) there were strain-specific differences in cardiac circadian rhythms; (2) strain-dependent differences in the effects of ORX on contractility genes translated into differences in cardiac functions; and (3) differential contractility responses occurred preferentially at times when circadian rhythms also showed strain-specific differences.

Methods: In hearts from the two above strains, we (1) profiled the expression levels of 15 circadian genes at 4-h intervals across a 24 h period; (2) tested the effects of either ORX or androgen replacement on expression of cardiac contractility genes, and that of ORX on myocardial functional reserve; and (3) verified whether the effects of MSY variants on cardiac contractility-related responses showed synchronicity with differences in circadian rhythms.

Results: Among the 15 tested circadian genes, a subset of them were affected by strain (and thus the genetic origin of MSY), which interacted with the amplitude of their peak of maximal expression at 2:00 PM. At that same time-point, ORX decreased (and androgen supplementation increased) the expression of three contractility-related genes, and decreased myocardial relaxation reserve in C57BL/6J.Y^A/J, but not in C57BL/6J mice. These effects were not detected at 10:00 AM, i.e., at another time-point when circadian genes showed no strain-specific differences.

Conclusions: The results indicate that in mice, androgens have activational effects on cardiac circadian rhythms, contractile gene expression, and myocardial functional reserve. All effects occurred preferentially at the same time of the day, but varied as a function of the genetic origin of MSY. Androgens may therefore be necessary but not sufficient to impart male-specific characteristics to some particular cardiac functions, with genetic material from MSY being one other necessary factor to fully define their range of actions.

Keywords: Androgens; Chromosome Y substituted mouse strains; Circadian genes; Circadian rhythms; Genetic variants; Heart; Male-specific portions of chromosome Y; Myocardial contractile reserve.

Fig. 1

Profiling of expression of 15 circadian genes in hearts from intact C57BL/6J and C57.Y^A/J male mice. Expression levels were measured at 4-h intervals between ZT0 and ZT20. Values at each time-point correspond to mean ± SEM (n = 4). The arrows point to ZT8, which is a time when several circadian genes show some disparity in expression levels. Post-hoc Tukey multiple comparison tests were performed for the three genes where the significance of the strain × time interaction had a P value <0.05 (*P < 0.05; **P < 0.01)

Fig. 2

Profiling of expression of Dbp, Tef, and Hlf in hearts from sham-operated C57BL/6J and C57.Y^A/J male mice. Expression levels were measured at 4-h intervals between ZT0 and ZT12. Values at each time-point correspond to mean ± SEM (n = 7–8). P values for each term of the two-way ANOVA analysis are as indicated. The asterisks correspond to the significance of the differences detected by post-hoc Tukey multiple comparison tests (**P < 0.01). The P values for all terms of the two-way ANOVA tests are as indicated

Fig. 3

Effects of either surgery (a–c) or of chronic treatment with DHT (d–f) on expression levels of Pln, Ttn, and Fhl2 in hearts from C57BL/6J and C57.Y^A/J male mice. Values at each time-point correspond to mean ± SD (n = 6–8). P values correspond to those for the interaction of strain with the effect of surgery. The asterisks (a–d) correspond to the significance of the differences detected by post-hoc Tukey multiple comparison tests (*P < 0.05; **P < 0.01; ***P < 0.001). e, f Significance of differences as detected by t tests between animals implanted with either empty (VEH) or DHT-filled (DHT) silastic tubes (*P < 0.05; ***P < 0.001)

Fig. 4

Effects of surgery on myocardial inotropic (left) and lusitropic (right) reserves. All measurements were performed at ZT8; values are mean ± SD (n = 6–8). Left: only strain had a significant effect; the asterisks correspond to the significance of differences as detected by post-hoc Tukey multiple comparison tests (***P < 0.001; **P < 0.01). Right: there was a significant interaction of strain with the effect of surgery on lusitropic reserve. The asterisks correspond to the significance of the differences detected by post-hoc Tukey multiple comparison tests (**P < 0.01)