The cardiac response to increased work includes a reactivation of fetal genes. The response to a decrease in cardiac work is not known. Such information is of clinical interest, because mechanical unloading can improve the functional capacity of the failing heart. We compared here the patterns of gene expression in unloaded rat heart with those in hypertrophied rat heart. Both conditions induced a re-expression of growth factors and proto-oncogenes, and a downregulation of the 'adult' isoforms, but not of the 'fetal' isoforms, of proteins regulating myocardial energetics. Therefore, opposite changes in cardiac workload in vivo induce similar patterns of gene response. Reactivation of fetal genes may underlie the functional improvement of an unloaded failing heart.

Relatively little is known concerning the regulation of uncoupling proteins (UCPs) in the heart. We investigated in the adult rodent heart 1) whether changes in workload, substrate supply, or cytokine (TNF-alpha) administration affect UCP-2 and UCP-3 expression, and 2) whether peroxisome proliferator-activated receptor alpha (PPARalpha) regulates the expression of either UCP-2 or UCP-3. Direct comparisons were made between cardiac and skeletal muscle. UCP-2, UCP-3, and PPARalpha expression were reduced when cardiac workload was either increased (pressure overload by aortic constriction) or decreased (mechanical unloading by heterotopic transplantation). Similar results were observed during cytokine administration. Reduced dietary fatty acid availability resulted in decreased expression of both cardiac UCP-2 and UCP-3. However, when fatty acid (the natural ligand for PPARalpha) supply was increased (high-fat feeding, fasting, and STZ-induced diabetes), cardiac UCP-3 but not UCP-2 expression increased. Comparable results were observed in rats treated with the specific PPARalpha agonist WY-14,643. The level of cardiac UCP-3 but not UCP-2 expression was severely reduced (20-fold) in PPARalpha-/- mice compared to wild-type mice. These results suggest that in the adult rodent heart, UCP-3 expression is regulated by PPARalpha. In contrast, cardiac UCP-2 expression is regulated in part by a fatty acid-dependent, PPARalpha-independent mechanism.

Background-Mechanical unloading of the heart results in atrophic remodeling. In skeletal muscle, atrophy is associated with inactivation of the mammalian target of rapamycin (mTOR) pathway and upregulation of critical components of the ubiquitin proteosome proteolytic (UPP) pathway. The hypothesis is that mechanical unloading of the mammalian heart has differential effects on pathways of protein synthesis and degradation. Methods and Results-In a model of atrophic remodeling induced by heterotopic transplantation of the rat heart, we measured gene transcription, protein expression, polyubiquitin content, and regulators of the mTOR pathway at 2, 4, 7, and 28 days. In atrophic hearts, there was an increase in polyubiquitin content that peaked at 7 days and decreased by 28 days. Furthermore, gene and protein expression of UbcH2, a ubiquitin conjugating enzyme, was also increased early in the course of unloading. Transcript levels of TNF-␣, a known regulator of UbcH2-dependent ubiquitin conjugating activity, were upregulated early and transiently in the atrophying rat heart. Unexpectedly, p70S6K and 4EBP1, downstream components of mTOR, were activated in atrophic rat heart. This activation was independent of Akt, a known upstream regulator of mTOR. Rapamycin treatment of the unloaded rat hearts inhibited the activation of p70S6K and 4EBP1 and subsequently augmented atrophy in these hearts compared with vehicle-treated, unloaded hearts. Conclusions-Atrophy of the heart, secondary to mechanical unloading, is associated with early activation of the UPP. The simultaneous activation of the mTOR pathway suggests active remodeling, involving both protein synthesis and degradation.

CD4+Foxp3+ regulatory T (Treg) cells were shown to control all aspects of immune responses. How these Treg cells develop is not fully defined, especially in neonates during development of the immune system. We studied the induction of Treg cells from neonatal T cells with various TCR stimulatory conditions, because TCR stimulation is required for Treg cell generation. Independent of the types of TCR stimulus and without the addition of exogenous TGF-β, up to 70% of neonatal CD4+Foxp3− T cells became CD4+Foxp3+ Treg cells, whereas generally <10% of adult CD4+Foxp3− T cells became CD4+Foxp3+ Treg cells under the same conditions. These neonatal Treg cells exert suppressive function and display relatively stable Foxp3 expression. Importantly, this ability of Treg cell generation gradually diminishes within 2 wk of birth. Consistent with in vitro findings, the in vivo i.p. injection of anti-CD3 mAb to stimulate T cells also resulted in a >3-fold increase in Treg cells in neonates but not in adults. Furthermore, neonatal or adult Foxp3− T cells were adoptively transferred into Rag1−/− mice. Twelve days later, the frequency of CD4+Foxp3+ T cells converted from neonatal cells was 6-fold higher than that converted from adult cells. Taken together, neonatal CD4+ T cells have an intrinsic “default” mechanism to become Treg cells in response to TCR stimulations. This finding provides intriguing implications about neonatal immunity, Treg cell generation, and tolerance establishment early in life.