Cardiac Hypertrophy in Transgenic Mice with Overexpressed Small-Subunit of Human Myosin Light Chain Phosphatase

BACKGROUND AND OBJECTIVES: Recent reports have demonstrated that perturbation of the balance between myo-sin light chain (MLC) phosphorylation and the dephosphorylation status is associated with the development of cardiac hypertrophy. Myosin light chain phosphatase (MLCP) is a key enzyme that regulates the phosphorylation status of the MLC, but its functional roles in cardiac muscle have not been well investigated. Especially, the functions of the small-subunit of MLCP in cardiac muscles are not well elucidated. Here, whether the human heart-specific small-subunit (M21) of MLCP is associated with hypertrophic responses in a transgenic mice model were assessed. MATERIALS AND METHODS: The transgenic mice, overexpressing the human M21, were generated from a cardiac-specific transgenic construct. Cardiac tissues from the transgenic mice were subjected to histology for their morphological examination. The echocardiographic parameters of the murine heart were examined with transgenic mice aged 1, 2 and 3 months, and compared with their non-transgenic littermates. To determine whether the transgenic heart was sensitive to stress, the echocardiographic examination was also performed at the baseline, both before and after the administration of isoproterenol, at a dosage of 30 microgram/g/day, for 2 weeks. RESULTS: The histological analysis of the transgenic heart revealed myocyte disarray and nuclear hypertrophy. No significant differences were observed between the transgenic and non-transgenic mice in relation to the echocardiographic determinants, such as the left ventricular dimensions and the wall thickness. Chronic cardiac stress, induced by isoproterenol infusion, also failed to show any significant differences in relation to the same determinants. CONCLUSION: Overexpression of the human M21 in the murine heart induced myocyte hypertrophy. However, the overall cardiac functions were not affected under normal and stressed conditions.

Cell-type specific regulation of thrombospondin-1 expression and its promoter activity by regulatory agents

Thrombospondin-1 (TSP-1), a multifunctional protein that is able to function as a negative regulator of solid tumor progression and angiogenesis, is normally present at a very low level but rapidly elevated in pathological tissues. To understand the cellular regulation of TSP-1 expression, the mode of it's expression in Hep3B, SK-HEP-1, and porcine aortic endothelial (PAE) cells was examined in the presence of all-trans retinoic acid (ATRA), interleukin-6 (IL-6), interferon-gamma (IFN-gamma), or phorbol 12-myristate 13-acetate (PMA). ATRA or IL-6 induced a dose-dependent increase of TSP-1 protein and mRNA levels in PAE cells, while they negatively regulated TSP-1 expression in the Hep3B and SK-HEP-1 cells. In contrast, PMA showed just the opposite effects on the TSP-1 expression in the same cells. IFN-gamma had little effect on TSP-1 level in Hep3B and PAE cells. The TSP-1 expression in SK-HEP-1 cells by these agents showed a close resemblance to that of liver cells rather than that of the endothelial cell line. Possible TSP-1 promoter-mediated responses by ATRA, IL-6, IFN-gamma, or PMA in Hep3B and PAE cells examined with luciferase activity of TSP-LUC reporter plasmid showed that levels of TSP-1 promoter activity were lower than that of the expressed TSP-1 protein and mRNA levels. Transfection of c-Jun and/or RARalpha expression vectors into Hep3B and PAE cells resulted in the enhanced TSP-1 promoter activity as well as the increments of of its protein and mRNA level. These results suggest that regulatory agents-induced TSP-1 expression may be attributed to mRNA stability and/or translational activation in concert with transcriptional activation and TSP-1 expression may be independently controlled via each signal pathway stimulated by PMA or ATRA.