Downregulation in GATA4 and Downstream Structural and Contractile Genes in the db/db Mouse Heart

Abstract

Reduced expression of GATA4, a transcriptional factor for structural and cardioprotective genes, has been proposed as a factor contributing to the development of cardiomyopathy. We investigated whether the reduction of cardiac GATA4 expression reported in diabetes alters the expression of downstream genes, namely, atrial natriuretic peptide (ANP), B-type natriuretic, peptide (BNP), and α- and β-myosin heavy chain (MHC). db/db mice, a model of type 2 diabetes, with lean littermates serving as controls, were studied. db/db mice exhibited obesity, hyperglycemia, and reduced protein expression of cardiac GLUT4 and IRAP (insulin-regulated aminopeptidase), the structural protein cosecreted with GLUT4. Hearts from db/db mice had reduced protein expression of GATA4 (~35%) with accompanying reductions in mRNA expression of ANP (~40%), BNP (~85%), and α-MHC mRNA (~50%) whereas expression of β-MHC mRNA was increased by ~60%. Low GATA4 was not explained by an increased ligase or atrogin1 expression. CHIP protein content was modestly downregulated (27%) in db/db mice whereas mRNA and protein expression of the CHIP cochaperone HSP70 was significantly decreased in db/db hearts. Our results indicate that low GATA4 in db/db mouse heart is accompanied by reduced expression of GATA4-regulated cardioprotective and structural genes, which may explain the development of cardiomyopathy in diabetes.

Figure 1

Body weight (a), plasma glucose (b), glycated haemoglobin (c), heart weight (d), and the heart-weight-to-body-weight ratio (e) in control and db/db mice. Values are expressed as mean ± SEM for 10–12 mice in each group. db/⁺, control mice; db/db, diabetic mice. *P < 0.05.

Figure 2

Cardiac GLUT4 and IRAP mRNA and protein expression in control and db/db mice. Values are expressed as mean ± SEM obtained from 2 separate experiments each performed with 5 hearts. db/⁺, control mice; db/db, diabetic mice. *P < 0.05, **P < 0.01, ***P < 0.001.

Figure 3

GATA4 is localized in the cell nuclei of cardiomyocytes of 2-day-old rats (a). GATA4 mRNA is expression is similar to that seen in control hearts (b). However, as illustrated in (c), protein expression of GATA4 is downregulated in db/db hearts (c). GATA4 was present in nuclei of the cells expressing cardiomyocyte marker, troponin C (d). Hearts from db/db mice showed a tendency toward a reduced weight (d), but cardiomyocyte surface was increased in db/db hearts (Figure 3e2) compared with control hearts (Figure 3e1). Figure (e3) shows that cardiomyocyte surface area, expressed as μm², was significantly increased in db/db hearts compared with control hearts. Values are expressed as mean ± SEM obtained from 2 separate experiments each performed with 5 hearts. db/⁺, control mice; db/db, diabetic mice. *P < 0.05.

Figure 4

Cardiac ANP and BNP mRNA expression in control and db/db mice. Values are expressed as mean ± SEM obtained from 2 separate experiments each performed with 5 hearts. db/⁺, control mice; db/db, diabetic mice. *P < 0.05, **P < 0.01.

Figure 5

Cardiac α-MHC mRNA expression (a) β-MHC mRNA expression (b), and protein expression (c) in control and db/db mice. Protein expression of β-MHC was not detected in control hearts, whereas in db/db hearts, β-MHC expression was elevated. Values are expressed as mean ± SEM obtained from 2 separate experiments each performed with 5 hearts. db/⁺, control mice; db/db, diabetic mice. *P < 0.05.

Figure 6

Cardiac CHIP mRNA and protein expression (a, b), atrogin1 mRNA expression (c), and MuRF1 mRNA expression (d) in control and db/db mice. Values are expressed as mean ± SEM obtained from 2 separate experiments each performed with 5 hearts. db/⁺, control mice; db/db, diabetic mice.

Figure 7

Cardiac HSP70 mRNA and protein expression in control and db/db mice. Values are expressed as mean ± SEM obtained from 2 separate experiments each performed with 5 hearts. db/⁺, control mice; db/db, diabetic mice. *P < 0.05, **P < 0.01.

Figure 8

Cardiac Nab-1 mRNA and protein expression in control and db/db mice. Values are expressed as mean ± SEM obtained from 2 separate experiments each performed with 5 hearts. db/⁺, control mice; db/db, diabetic mice. **P < 0.01.