[Show abstract][Hide abstract] ABSTRACT: We have used an oligonucleotide microarray to identify genes that are affected by congestive heart failure and those influenced by treatment with DITPA and DITPA in combination with captopril using a rat postinfarction model. The most striking result when comparing heart failure to sham operation was that all of the mitochondrial and metabolic enzymes affected were down regulated. When comparing heart failure with DITPA treatment, most of the down regulated metabolic genes were returned toward normal. When comparing heart failure with heart failure animals treated with DITPA and captopril, metabolic enzymes were no longer significantly downregulated. DITPA treatment and the combination of DITPA and captopril show that the metabolic enzymes were no longer down regulated. This represents a substantial improvement in the energy- generating capacity of the heart. These results indicate that the actions of DITPA and the combination of DITPA and captopril in heart failure can be partially explained by differences in gene activation.
[Show abstract][Hide abstract] ABSTRACT: The heart is an important target of thyroid hormone actions. Only a limited number of cardiac target genes have been identified, and little is known about their regulation by T(3) (3,3',5-triiodothyronine) and thyroid hormone analogs. We used an oligonucleotide microarray to identify novel cardiac genes regulated by T(3) and two thyroid hormone analogs, 3,5-diidodothyropropionic acid (DITPA) and CGS 23425 [N-[3,5-dimethyl-4-(4'-hydroxy-3'-isopropylphenoxy)-phenyl]-oxamic acid]. DITPA binds with lower affinity than T(3) to thyroid hormone receptor alpha1 and beta1 isoforms, whereas CGS 23425 binds selectively to beta1. Fluorescent-labeled cDNA was prepared from cultured heart cells maintained in medium stripped of thyroid hormone ("hypothyroid" control) or treated with T(3), DITPA, and CGS 23425 at concentrations 5 times their respective K(d) values for 48 h. The arrays were scanned and analyzed using an analysis of variance program. Sixty-four genes were identified that were >1.5 times up- or down-regulated by one of the treatments with P < 0.05. The genes regulated by T(3) and DITPA were nearly identical. Thirteen genes were differentially regulated by CGS 23425. Genes encoding contractile proteins, Ca(2+)-ATPase of sarcoplasmic reticulum and several proteins of mitochondrial oxidative phosphorylation, were up-regulated by T(3) and DITPA but not by CGS 23425. These results indicate that some, but not all, of the actions of thyroid hormone analogs can be explained by differences in gene activation.
Journal of Pharmacology and Experimental Therapeutics 10/2004; 311(1):164-71. DOI:10.1124/jpet.104.069153 · 3.97 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: In the myocardium, myocyte cell division is irreversibly blocked shortly after birth. The signal that initiates cell cycle withdrawal is unknown. The purpose of this study was to relate changes in expression of beta1 integrin and its associated alpha subunits to cardiomyocyte cell cycle progression during the fetal-to-neonatal developmental transition in rat.
The developmental expression pattern and function of beta 1 integrin and several of its associated alpha subunits were examined using reverse transcription (RT) polymerase chain reaction (PCR) and beta 1 blocking antibodies. During the fetal to neonatal transition, a dramatic shift occurred in the levels of beta1 and alpha isoforms. At the 17-day fetal stage only beta 1A was present, which remained relatively constant until immediately after birth then decreased by 30% at the adult stage. By contrast, beta 1D appeared at fetal day 18, increased at neonatal day 2, and afterwards remained constant. This resulted in a ratio of beta 1A to beta 1D of about 1:1 in the adult heart. The integrin beta 1-associated subunits, alpha 3, alpha 6, and alpha 7, were expressed at extremely low levels in 17-day fetal cardiomyocytes. After birth alpha 3 and alpha 6 transiently increased at the 2-day neonatal stage, while alpha 7 isoforms B, C, and X2 progressively increased to the adult stage. Unlike skeletal muscle cells, fluorescence-activated cell sorting analysis (FACS) showed no down regulation of the alpha 5 beta 1 fibronectin receptor during cell cycle withdrawal. Treatment of cultured cardiomyocytes with beta1 blocking antibody inhibited the cell cycle in fetal but not in neonatal cells.
These results suggest that progression through the cardiomyocyte cell cycle may be dependent upon cell attachment via integrin beta1 and correlate with changes that occur in beta1 spliced variants and their respective alpha isoforms.
Cardiovascular Research 10/2000; 47(4):715-25. · 5.94 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Cardiomyocyte terminal differentiation was examined by studying the interaction of retinoblastoma protein (pRb) family members with E2F during the developmental transition from 17-day fetal to 2-day neonatal. Additionally, the expression pattern of cyclins, cyclin-dependent kinases (CDKs), and CDK inhibitors responsible for modulating the phosphorylation of pRb were studied. p107, pRb, and p130 are regulators of cellular proliferation, differentiation, and cell cycle exit and entry, respectively. The active, underphosphorylated form of these proteins targets the E2F family of transcriptional factors that play a critical role in the control of genes associated with DNA synthesis. Electromobility shift analyses demonstrated E2F complexed with p107 in proliferating fetal cardiomyocytes, whereas in 2-day neonatal cells, E2F was principally associated with p130 and a low level of pRb. At the 2-day neonatal stage, decreased protein levels were observed for cyclins D2, D3, and E, and CDK2 and CDK4. No changes were observed in the mRNA levels of the D-cyclins in neonatal cells; however, the transcripts for cyclins A and E and CDK4 were diminished. In skeletal myoblasts, differentiation is associated with induction of p21, a CDK inhibitor, by a MyoD-dependent pathway. Although heart cells lack MyoD, CDK assays demonstrated that the activity of CDKs 2, 4, and 6 were downregulated in 2-day neonatal cells, and CDC2 was increased. RT-PCR indicated that p21 mRNA was induced 1.4-, 2.0-, and 3.1-fold in the 2-day neonatal, 7-day neonatal, and adult stages, respectively, compared to the 17-day fetal stage. At the protein level, p21 also increased at the 2-day neonatal stage. Kinase inhibitory immunodepletion assays showed that CDK inhibitory activity was markedly increased in the 2-day neonate. Although mRNA levels of the p27 CDK inhibitor were unchanged, its protein level and inhibitory effect on CDK2 and CDK4 were increased. Thus, cardiomyocytes retain the capacity to proliferate until the early neonatal period when a series of changes occur, including a switch in pRb partners, a decrease in CDK levels and induction of CDK inhibitory activity, which is associated with terminal differentiation.
Journal of Molecular and Cellular Cardiology 03/1998; 30(3):563-78. DOI:10.1006/jmcc.1997.0620 · 4.66 Impact Factor